bioacid

Transcription

bioacid

BIOACID
A proposed national ‘Verbundprojekt’ of the Federal Ministry of Education and Research
Coordinator:
Prof. Dr. Ulf Riebesell
Forschungsbereich Marine Biogeochemie
Leibniz-Institut für Meereswissenschaften an der Universität Kiel
Düsternbrooker Weg 20
24105 Kiel
Tel. 0431-600-4444
Email: [email protected]
Partners:
Alfred-Wegener-Institut für Polar- und Meeresforschung, Bremerhaven
Carl von Ossietzky Universität Oldenburg
Christian-Albrechts-Universität zu Kiel
Forschungszentrum TERRAMARE, Wilhelmshaven
GKSS-Forschungszentrum Geesthacht GmbH
Heinrich-Heine-Universität, Düsseldorf
Jacobs – University, Bremen
Leibniz-Institut für Gewässerökologie und Binnenfischerei, Berlin
Leibniz-Institut für Meereswissenschaften IFM-GEOMAR, Kiel
Leibniz-Institutes für Ostseeforschung Warnemünde
Ludwig-Maximilians-Universität München
Max-Planck-Institut für Marine Mikrobiologie, Bremen
PreSens Precision Sensing GmbH, Regensburg
Ruhr-Universität Bochum
Universität Bremen
Universität Hamburg
Universität Rostock
Westfälische Wilhelms - Universität Münster
Zentrum für Marine Tropenökologie (ZMT), Bremen
BIOACID: Biological Impacts of Ocean Acidification
Zusammenfassung…………………………………………………………………...
4
Summary…………………………………………………………………………….
5
1. Project background…………………………………………………………….
6
2. Scientific and technological objectives………………………………………. .
8
3. Relevance to National Priorities and the National Funding Policy…………..
9
4. Structure of the Project and Consortium Building……………………….…..
10
5. Overview of Themes 1-5………………………………………………………...
13
5.1. Theme 1: Primary production, microbial processes
and biogeochemical feedbacks ………………………………..... 13
5.2. Theme 2: Performance characters: reproduction, growth and
behaviours in animal species ………………………………….... 16
.
5.3. Theme 3: Calcification - sensitivities across phyla and ecosystems....…... 19
5.4. Theme 4: Species interactions and community structure
in a changing ocean.......................................................................
22
5.5. Theme 5: Integrated assessment – sensitivities and uncertainties….…....
25
6. Management structure and procedures ………………………………….……. 28
7. Data management and dissemination…………………………………………..
31
8. Infrastructure development, training and transfer of know-how………….…
32
9. International and National Cooperation…………………………………….....
33
10. Summary Budget………………………………………………………………...
38
11. Detailed descriptions of Themes and Projects………………………………....
39
11.1: Theme 0: Overarching activities………………………………………....
41
0.1 Project coordination……………………………..…………………………..
0.2 BIOACID data management…………………………………………….…..
0.3 Infrastructure development…………………………………….....................
0.4 Training and transfer of knowhow……………………………........……..…
41
44
49
63
11.2: Theme 1: Primary production, microbial processes
and biogeochemical feedbacks…………………………….…..
67
1.1: Acclimation versus adaptation in autotrophs……………………………….
1.2: Turnover of organic matter……………………………………………....….
1.3: Modelling biogeochemical feedbacks of the organic carbon pump……..….
70
85
98
11.3: Theme 2: Performance characters: reproduction, growth and
behaviours in animal species……………………………….....
2.1: Effects on grazers and filtrators……………………………………….…....
2.2: Long-term physiological effects on different life stages
of benthic crustaceans....................................................................................
2.3: Effects on top predators (fishes, cephalopods)……………………………..
2
105
108
121
134
11.4: Theme 3: Calcification - sensitivities across phyla and ecosystems…..
3.1: Cellular mechanisms of calcification………………………………………..
3.2: Calcification under pH-stress: Impacts on ecosystem and
organismal levels…………………………………………………………....
3.3: Ultra-structural changes and trace element / isotope partitioning in
calcifying organisms…………………………………………………… …...
3.4: Microenvironmentally controlled (de-)calcification mechanisms…………...
3.5: Impact of present and past ocean acidification on metabolism,
biomineralization and biodiversity of pelagic and neritic calcifiers…………
145
148
168
183
193
207
11.5: Theme 4: Species interactions and community structure in
a changing ocean........................................................................
219
4.1: OA impacts on interactions in and structure of benthic communities……….
224
4.2: OA effects on food webs and competitive interactions
in pelagic ecosystems ……………………………………………………...
236
11.6: Theme 5: Integrated assessment – sensitivities and uncertainties…….
247
5.1: Impact of Alkalinity fluxes from the Wadden Sea on the carbon cycle
and the primary production in the North Sea…………………………………
5.2: Evaluating and optimising parameterisations of pelagic calcium
carbonate production in global biogeochemical ocean models……………….
5.3: Viability-method for the impact assessment of ocean acidification
under uncertainty……………………………………………….………….….
12. Appendices……………………………………………………………………….
249
255
261
264
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Zusammenfassung
Neben der Atmosphäre ist der Ozean die zweitgrößte Senke für anthropogenes Kohlendioxid.
Das vom Ozean aufgenommene CO2 reagiert mit dem Meerwasser und bildet Hydrogenkarbonat
und Säure, bei gleichzeitiger Zehrung von Karbonationen. Das Ergebnis dieses Prozesses, der als
Ozeanversauerung bezeichnet wird, ist ein Anstieg der Konzentrationen von CO2 und
Hydrogenkarbonat und eine Abnahme des pH Wertes (Zunahme des Säuregrades) und der
Karbonationenkonzentration. Bei ungebremst fortschreitenden CO2 Emissionen wird sich die
Chemie des Meerwassers bis zum Ende dieses Jahrhundert in einer Weise verändern, wie es die
meisten der heute im Ozean lebenden Organismen während ihrer jüngeren Evolution nicht erlebt
haben. Dies könnte Folgen für die Konkurrenzfähigkeit pH/CO2 sensibler Arten haben. Bei
zunehmender Ozeanversauerung droht dadurch der Verlust an Biodiversität, ökologische und
funktionale Veränderungen in den marinen Lebensgemeinschaften und eine reduzierte Kapazität
des Ozeans zur weiteren Aufnahme von anthropogenem CO2.
Trotz der Risiken, die diese Entwicklung in sich birgt, fehlt es uns an einem grundlegenden
Verständnis der möglichen Konsequenzen einer Ozeanversauerung. Um diese Lücke zu schließen
und eine System basierte Abschätzung der hiermit verbundenen Risiken und Unsicherheiten zu
erlangen, wird BIOACID die Expertise von Molekular- und Zellbiologen, Biochemikern,
Pflanzen- und Tierphysiologen, Meeresökologen, marinen Biogeochemikern und
Ökosystemmodellierern in einem integrierenden Ansatz kombinieren. Über Disziplin-, Themenund Projektgrenzen hinweg werden die BIOACID Partner gemeinschaftlich Experimente
durchführen, Labor übergreifend Ausrüstung und Messkapazitäten nutzen, Probenmaterial und
Fachkompetenz austauschen und die umfangreichen Datensätze in Richtung auf ökosystemare
Modelle der Ozeanversauerung analysieren und synthetisieren. Dies wird ergänzt durch
Trainingsworkshops zu zentralen Forschungsinhalten und -methoden von BIOACID Experten für
alle Mitglieder des Konsortiums. Die übergeordneten Fragestellungen des Projektes sind:
Welche Auswirkungen hat die Ozeanversauerung auf die marinen Organismen und ihre
Lebensräume, was sind die zu Grunde liegenden Mechanismen und möglichen
Anpassungen auf der Ebene von Populationen und Gemeinschaften, in welchem Maße
werden die Auswirkungen durch andere Stressfaktoren beeinflusst und welche
Konsequenzen ergeben sich daraus für die marinen Ökosysteme, biogeochemischen
Kreisläufe und mögliche Rückkopplungen auf das Klimasystem?
Um diese Fragen über einen weiten Bereich von potentiell sensitiven biologischen Prozessen
anzuwenden, sind die in BIOACID geplanten Forschungsaktivitäten nach Schlüsselkomponenten
und funktionalen Gruppen der marinen Lebensgemeinschaften strukturiert. Aus den diversen
experimentellen Ansätzen und Feldbeobachtungen gewonnene Erkenntnisse sollen auf der Basis
einer integrierenden Datensynthese dazu beitragen, Unsicherheiten bezüglich der
prognostizierten Entwicklung zu identifizieren und mögliche Schwellenwerte unumkehrbarer
Veränderungen zu erkennen. Ein zusätzlicher Mehrwert für das Projekt soll durch enge
Kooperation mit verwandten nationalen und internationalen Projekten mit Schwerpunkt auf
Ozeanversauerung und Ozeanerwärmung erzielt werden.
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Summary
Next to the atmosphere, the ocean is the second largest sink for anthropogenic carbon dioxide. As
fossil fuel CO2 enters the surface ocean, it reacts with seawater to form bicarbonate and protons,
thereby consuming carbonate ions. The net result of this process, termed ocean acidification, is
an increase in CO2 and bicarbonate concentrations and a decrease in seawater pH and carbon ion
concentration. If CO2 emissions continue to rise at current rates, before the end of this century the
resulting changes in seawater chemistry will expose marine organisms to conditions which they
have not experienced during their recent evolutionary history and which may pose a threat to the
competitive fitness of pH/CO2 sensitive species and groups. Thus, as the ocean continues to
acidify, there is an increasing risk of biodiversity losses, of profound ecological and functional
shifts, and of a reduced capacity of the ocean to buffer further CO2 increase.
Despite this emerging problem and the risks it may involve, surprising little is know about the
possible impacts of ocean acidification. To close the many gaps in our understanding and to
allow a systems-based assessment of the risks and uncertainties associated with ocean
acidification, BIOACID will take an integrated approach combining the expertise of molecular
and cell biologists, biochemists, plant and animal physiologists, marine ecologists, ocean
biogeochemists and ecosystem modellers. The interaction between BIOACID scientists across
disciplines, research themes and projects will include joint experiments, collaborative use of
equipment and measurement capacity, exchange of samples and expertise, and the analysis and
synthesis of comprehensive data sets towards an ecosystem model of ocean acidification. These
activities will be complemented by training workshop offered by BIOACID experts to all
members of the consortium. Following this approach, the overarching questions of BIOACID
are:
What are the effects of ocean acidification on marine organisms and their habitat, what are
the underlying mechanisms of responses and possible adaptations on the level of
populations and communities, how are they modulated by other environmental stressors,
and what are the consequences for marine ecosystems, ocean biogeochemical cycles, and
possible feedbacks to the climate system?
To address these questions for a wide range of potentially sensitive biological processes, research
activities will be structured according to key ecosystem components and functional groups.
Information gained in a variety of experimental approaches and field assays will be synthesized
to obtain an integrated assessment of sensitivities and uncertainties and to identify the potential
thresholds associated with ocean acidification. BIOACID will benefit from close interactions
with related national and international project focussing on ocean acidification and ocean
warming.
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1.
Project background
The world’s oceans help moderating climate change thanks to their extensive capacity to store
anthropogenic carbon dioxide. Since pre-industrial times, the oceans have sequestered nearly half
of the fossil fuel CO2 released into the atmosphere and presently take up approximately 30% of
current CO2 emissions (Sabine et al. 2004). As CO2 enters the surface ocean it reacts with
seawater and generates changes in carbonate chemistry already measurable today (Figure 1). In
case of unabated CO2 emissions, the resulting changes in seawater chemistry will, in the course
of this century, expose marine organisms to conditions which they may not have experienced
during their recent evolutionary history (Raven et al., 2005). This raises concerns regarding the
biological, ecological, biogeochemical, and societal implications of ocean acidification.
Fig. 1:
Measured changes in surface ocean CO2 partial pressure (pCO2) and pH at the European Station for
Time-series in the Ocean off the Canary Islands (ESTOC), the Hawaii Ocean Time-series Station
(HOT) and the Bermuda Atlantic Times Series station (BATS). Since 1750, surface ocean pH has
decreased by 0.12 units (calculated). Since 1980, pH has decreased by 0.02 units per decade (measured).
Source: IPCC 4th Assessment Report (2007)
In its Special Report “The Future Oceans – Warming up, Rising High, Turning Sour”, the
German Advisory Council on Global Change (WBGU, Berlin 2006) states: “Because of the
importance of the consequences of ocean acidification, research in this area should be intensified
considerably. As long as there is no general scientific consensus about the tolerable limit for the
effects of acidification, a margin of safety according to the precautionary principle should be
observed. The suggestion of the WBGU to prevent a pH decrease of more than 0.2 is oriented
toward the goal of avoiding an aragonite undersaturation in the ocean surface layer. If it is found
that other intolerable damages already occur before reaching aragonite undersaturation, then the
guard rail will have to be adjusted accordingly.” (see Fig. 2)
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Fig. 2:
Mean surface ocean pH values during glacial a pre-industrial times (calculated from reconstructed
atmospheric pCO2), measured pH values in the present ocean, and projected future seawater pH for an
atmospheric CO2 concentration of approx. 750 ppm. The red line indicates the WBGU guard rail. Source:
WBGU Special Report (2006) after IMBER (2005)
As stated in the report, the tolerable window for ocean acidification defined by WBGU presently
relies on an extremely small data base. In fact, rather than using the limited data on observed
biological consequences of ocean acidification, the WBGU reaches its recommendation on the
basis of projected changes in water chemistry (aragonite saturation state). While this is an
appropriate approach in view of the scarcity of biological information, there is a clear need to
establish a reliable data base on tolerance levels for ocean acidification in key groups of pH/CO2
sensitive marine organisms in order to reach a more informed recommendation.
Our present knowledge on the effects of ocean acidification (OA) on the marine biota is largely
based on experimental work with single organisms or strains maintained in short-term
incubations often exposed to abrupt and extreme changes in carbonate chemistry. Based on
presently available data little is known about OA induced habitat change, synergetic effects from
other stressors, such as ocean warming, the sensitivity of genetically diverse populations, and the
ability of organisms to undergo long-term physiological and genetic adaptations. Hence, it is
difficult to predict how the responses of key species will affect the population, community and
ecosystem level, for example by the replacement of OA-sensitive with OA-tolerant species. In
view of these uncertainties, it is presently impossible to define critical thresholds (tipping points)
for tolerable pH decline or to predict the pathways of ecosystem changes where threshold levels
have been surpassed.
BIOACID will take the challenge to address the substantial gaps in the understanding of the
consequences of ocean acidification. To achieve this, BIOACID will take an integrated approach
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combining the expertise of molecular and cell biologists, biochemists, plant and animal
physiologists, marine ecologists, ocean biogeochemists and ecosystem modellers. Close
collaboration will be established with other European national and EU projects on ocean
acidification to complement the expertise and research capacities of the marine science
community in Germany and to benefit from the emerging synergies. Most notably, BIOACID
will profit from existing links to the European Project on Ocean Acidification (EPOCA) and
intends to develop close interactions and possibly joint activities with the UK programme on
ocean acidification presently developed in the framework of the Natural Environment
Research Council’s (NERC) new five-year strategy.
References
Fabry VJ, Seibel BA, Feely RA, Orr JC (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICES J Mar Sci 65:
414–432.
Feely RA, Sabine CL, Hernandez-Ayon JM, Ianson D, Hales B (2008) Evidence for upwelling of corrosive "acidified" water onto the
continental shelf. Science 320: 1490 – 1492.
Raven et al. Royal Society, Ocean Acidification Due to Increasing Atmospheric Carbon Dioxide (Policy Document 12/05, Royal Society,
London, 2005).
Sabine, C.L., R.A. Feely, N. Gruber, R.M. Key, K. Lee, J.L. Bullister, R. Wanninkhof, C.S. Wong, D.W.R. Wallace, B. Tilbrook, F.J. Millero,
T.-H. Peng, A. Kozyr, T. Ono, and A.F. Rios (2004), The Oceanic sink for Anthropogenic CO2, Science, 305, 367-371.
WBGU Special Report “The Future Oceans – Warming up, Rising High, Turning Sour”, the German Advisory Council on Global Change,
Berlin 2006
2.
Scientific and technological objectives
BIOACID will assess uncertainties, risks and thresholds related to the emerging problem of
ocean acidification (OA) at molecular, cellular, organismal, population, community and
ecosystem scales. It will
contribute to determining the impacts of OA on marine biota and their potential for
acclimation and adaptation,
improve our understanding of OA-related ecosystem changes, including both pelagic and
benthic habitats, synthesize information for ecosystem modelling and determine critical
thresholds,
assess the consequences of OA-induced biological responses on elemental cycling and
biogeochemical feedbacks to the climate system.
To achieve this goal, BIOACID will (i) employ a suite of observational, experimental and
modelling approaches leading to an integrated assessment of potential risks and thresholds
associated with ocean acidification (see Section 4 and 10), and (ii) coordinate with major national
and international projects and programmes (see Section 8). The project aims to deliver scientific
information and knowledge that directly addresses important global environmental issues and
that is relevant to national policy issues relating to energy policy, the biodiversity agenda, and
adaptation to global change. Major deliverables will include:
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high-profile scientific contributions to IGBP’s international science programmes of IMBER
(Integrated Marine Biogeochemical and Ecosystem Research) and SOLAS (Surface Ocean
Lower Atmosphere Study) based on Germany’s national marine expertise and infrastructure
strengthened scientific basis for policy-relevant assessments, including input into the IPCC
(Intergovernmental Panel on Climate Change) 5th Assessment Report and providing the
scientific foundation for assessments of changes in marine biodiversity, e.g. in the framework
of DIVERSITAS.
3.
Relevance to National Priorities and the National Funding Policy
BIOACID addresses topics of global change research that are central to the BMBF’s mission
(see below) and which are of immediate relevance with regard to the “Positionspapier für eine
kohärente deutsche Forschungsstrategie zum Globalen Wandel” developed by the Nationales
Komitee für Global Change Forschung to guide national research over the next years. BIOACID
will take the challenge of providing a sound data base on which to make recommendations
regarding critical thresholds for ocean acidification as requested in the Special Report “The
Future Oceans – Warming up, Rising High, Turning Sour” by the German Advisory Council on
Global Change (WBGU, Berlin 2006). BIOACID’s combination of experimental approaches,
field studies and modelling activities to assess different scenarios of future CO2 levels, targeting
in a coordinated manner all relevant entities from the cell to the ecosystem and from short to
long-term effects, is a unique and ambitious approach to the emerging problem of ocean
acidification. Research carried out under BIOACID is also relevant as a basis for informed
decisions on adaptation and mitigation strategies developed in the context of the
“Aktionsprogramm Forschung zum Klimawandel”.
In particular, BIOACID directly addresses goals of the BMBF’s Earth System and Marine
Research programmes by focussing on the response of marine organisms and ecosystems to a
changing global environment. The project is designed to assess the sensitivity of key species,
communities, and habitats to variable forcing including anthropogenic change such as
elevated CO2 levels. In this way, the project will improve the understanding of mechanisms by
which the ocean and its ecosystems react to human and natural impacts, including feedbacks of
such changes on biogeochemical cycles, the atmospheric composition and climate. The project
will therefore contribute to a better understanding of the role of the ocean in the climate
system and an improved description of the effects of global change on sensitive marine
ecosystems. Such understanding is a pre-requisite for prediction and assessment of the impacts of
human activity including climate-protection measures, on climate, global ecosystems and marine
natural resources.
Due to its multi-disciplinary nature, BIOACID also contributes directly to at least three Priority
Research Themes of the BMBF’s Geotechnology Programme, namely: (1) The Coupled System
Earth – Life, (2) Global Climate Change: Causes and Effects and (3) Biogeochemical
Cycles: Links between Geosphere and Biosphere. Furthermore, BIOACID research is of
immediate relevance to the new BMBF framework programme “Research for Sustainability”
and will make significant contributions to risk assessments on the loss of marine biodiversity,
including input to the Global Biodiversity Information Facility.
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4.
Structure of the Project and Consortium Building
4.1.
Scientific programme
In order to achieve the objectives of BIOACID, five thematic areas have been identified which
cover the range of processes from the base of the marine food chain to the community and
ecosystem level, and of mechanisms from the sub-cellular to the whole organism level. In view
of their distinct sensitivities to ocean acidification, calcification and carbonate dissolution
processes will be the focal point of a separate theme. According to these research priorities the
scientific programme of BIOACID is structured under the following five themes:
Theme 1:
Primary production, microbial processes
& biogeochemical feedbacks
Theme 2:
Performance characters:
Reproduction, growth & behaviours in animal species
Theme 3:
Calcification: Sensitivities across phyla and ecosystems
Theme 4:
Species interactions and community structure in a changing ocean
Theme 5:
Integrated assessment: Sensitivities and uncertainties
BIOACID will pursue a multidisciplinary approach, involving scientists with expertise in cell and
molecular biology, microbiology, physiology, evolutionary biology, marine ecology, marine
chemistry, biogeochemistry and numerical modelling. BIOACID will employ a wide range of
scientific approaches and methodologies, extending from field monitoring of OA-sensitive areas
and ecosystems to joint mesocosm perturbation experiments, combined with closely coordinated
ecosystem and biogeochemical modelling activities using parameterizations of observed
biological responses and their biogeochemical implications.
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Theme 1
Project coordination
Data management
Primary production,
microbial processes
and biogeochemical
feedbacks
Theme 3
Calcification:
Sensitivities across
phyla and ecosystems
Infrastructure
development
Training and transfer
of knowhow
Fig. 3:
Theme 5
Theme 2
Performance characters:
reproduction, growth
and behaviours in
animal species
Theme 4
Species interactions
and community
structure in a
changing ocean
Integrated
assessment:
Sensitivities and
uncertainties
Structure of the project: research themes and overarching activities. Joint activities and the multiple
links between themes and projects are described in detail in section 11.
During the development of the research programme strong emphasis was put on linking the subprojects and projects, both within and between themes. These links range from joint experiments,
joint use of equipment and measurement capacity, to exchange of samples, exchange of expertise,
and training workshop offered by BIOACID experts to all members of the consortium (see 8.2.
Training and transfer of know how). For detailed information on the multiple linkages developed
in BIOACID see Table 0.1 under 11. Detailed Description of Themes and Projects and the
specific links described in each of the individual projects. Over-arching activities include not
only project coordination and logistics, outreach and data management but also the development
and application of joint infrastructure (CO2 sensors, benthic and pelagic mesocosm systems,
NMR, NanoSIMS).
4.2.
Consortium building
The BIOACID consortium was built based on a bottom-up, open competition approach among all
interested German institutes and universities conducting marine-oriented research. Starting with a
presentation of the project idea to representatives of the Projektträger Jülich (PTJ) in
Warnemünde in May 2007, followed by a positive signal from the PTJ, a planning group (Table
1) was formed and first met at the MPI in Bremen July 2007.
11
Table 1: Composition of the planning group.
Planning group
Members
Chairs:
Ulf Riebesell (IFM-GEOMAR), Hans-Otto Pörtner (AWI)
Members:
Antje Boetius (MPI Bremen), Gerd Graf* (U Rostock), Thorsten Reusch
(U Münster), Maren Voss (IOW)
* during the pre-proposal writing stage Gerd Graf (U Rostock) asked to withdraw from the
planning group; Maren Voss (IOW) was invited as a replacement
In the following months the planning group developed a tentative structure for a German national
programme on ocean acidification and prepared a core proposal which was submitted to the
German Ministry for Education and Research (BMBF) in November 2007. An oral presentation
of the core proposal to representatives of the BMBF (Referat 725 System Erde and Referat 723
Globaler Wandel) and of the PTJ took place in Bonn in December 2007. Based on this
presentation and the follow-up discussions the planning group was invited to prepare a full
proposal for international peer review.
In January 2008 a call for project contributions was made public and sent to potential partner
universities/institutes, requesting a 2-page description of proposed contributions to the
programme by February 15, 2008. Over 50 pre-proposals were received and screened by the
planning group during a group meeting in Kiel in February 2008 based on quality and
complementarity of the proposed research. The principle investigators (PI) of all selected preproposals were invited to participate in a workshop and at the same time informed about the
tentative budget allocated to their project contributions, as recommended by the planning group.
A 2-day workshop of all invited PIs was held in Kiel in April 2008, during which the consortium
was founded and the structure of the programme, including the themes, projects and sub-projects
were developed. During the workshop theme and project leaders (see Tables 2 and 3) were
chosen and responsibilities for the preparation of the full-proposal were assigned. It followed a
10-week period of intense communication and writing activities by all partners involved. The full
proposal was completed in June and proudly handed over to the PTJ on July 2, 2008.
Time line of consortium building and proposal preparation:
(1)
Presentation of project idea at PTJ Warnemünde
May
2007
(2)
Planning group established
July
2007
(3)
Planning group finalizes core proposal
Nov.
2007
(4)
Presentation of project idea to BMBF and PTJ in Bonn
Dec.
2007
(5)
Call for project contributions to potential partner universities/institutes Jan.
2008
(deadline for submission: February 15, 2008)
(6)
Screening of proposed contributions based on quality and
complementarity, invitation of project partners
12
Febr.
2008
(7)
joint workshop of all partners, revision of core proposal,
Apr.
2008
April - June
2008
July
2008
development and structuring of themes, projects, and sub-projects
(8)
preparation of full proposal by all project partners
(9)
submission of full proposal
5.
Overview of Themes 1-5
5.1.
Theme 1:
Primary production, microbial processes and
biogeochemical feedbacks
5.1.1. Theme summary
Overarching questions
1. How do marine primary producers and heterotrophic bacteria of diverse taxonomic
groups respond to ocean acidification (OA) and increase in CO2 concentration on?
Which groups/species (e.g. calcifying vs. non calcifying species) are negatively impacted
and which benefit?
2. To what extent will key phytoplankton species and bacteria be able to acclimate to OA?
What is the potential for evolutionary adaptation and concomitant genetic changes
within 100s to 1000s of generations?
3. What are the consequences of ocean acidification for the turn-over and export of
organic matter?
4. What are the combined effects of CO2 and temperature changes on the marine soft
tissue pump and DOC export and the air-sea exchange of CO2?
Primary producers in the marine realm encompass phylogenetically very diverse groups
(Falkowski et al. 2004), differing widely in their photosynthetic apparatus and carbon enrichment
systems (Giordano et al. 2005). Preliminary data reveal that species with effective carbon
concentration mechanisms (CCMs) are less sensitive to increased CO2 levels than those lacking
efficient CCMs, analogous to findings in terrestrial vegetation. Currently, our ignorance of the
metabolic diversity of oceanic autotrophy and microbial heterotrophs hampers any projection of
total marine primary production and regeneration in response to increased carbonation. A focus
of Theme 1 will therefore be to identify critical physiological traits that determine the sensitivity
of key groups of primary producers and bacteria to increases ocean acidification and carbonation.
Most of the biological oceanic carbon uptake is driven by regenerated production and only ~20%
by new nitrogen input to surface waters (Laws et al., 2000) which drives the export of organic
carbon from surface waters (Eppley and Peterson 1979). While the export of carbon, nitrogen,
and phosphorus is generally considered to be closely coupled in marine biogeochemical cycling,
recent work in mesocosms shows increasing C:N and C:P ratios during primary production with
increasing CO2 concentrations (Riebesell et al., 2007). Excess carbon assimilation partially ends
up as dissolved organic carbon and may increase the export of organic matter through the
formation of gel particles that enhance particles aggregation, such as transparent exopolymer
13
particles (Arrigo, 2007; Engel et al., 2004). These processes, if representative for pelagic
autotrophic communities, could give rise to a biologically driven feedback to the climate system.
What controls the release of dissolved organic matter by either phytoplankton or bacteria, and
how these substances affect the nutrition and aggregation of pelagic organisms needs further
investigation in order to improve the description of biogeochemical turnover processes and their
sensitivities to increasing pCO2 (Figure 4).
Theme 1 will study plankton communities in controlled lab experiments on several relevant time
scales, from short-term physiological adjustment, to acclimation, to longer-lasting evolutionary
adaptation. Treatment levels, experimental set-ups and response variables were chosen
concordantly among diverse target groups in order to ensure full comparability of results in the
subprojects.
Fig. 4: Compartments and interactions studied in Theme 1
Theme 1 will combine laboratory-based work with field campaigns to assess the responses of
natural plankton communities. Given the importance of coastal areas to fisheries and other
marine resources and services, the coastal ecosystems constitute an important target region. Insitu experiments and sampling will be carried out in the Baltic Sea, an enclosed sea with high
nutrient input and anthropogenic pressures.
14
An improved implementation of possible impacts of ocean acidification and sea surface warming
influence on the marine soft-tissue pump and the cycling (and export) of dissolved organic
carbon in global marine carbon cycle models, like the model PICES, is urgently needed. As part
of the modeling component of Theme 1 new parameterizations will be incorporated based on
empirical results generated under this theme.
5.1.2. Progress expected
Theme 1 aims at a better understanding of complex species interactions in communities
comprising primary producers and bacteria and associated biogeochemical dynamics under
present pCO2 concentrations, twice (projected for 2100) and three times the present
concentration. Laboratory and field experiments will deliver insight into the sensitivity of key
plankton species to high CO2 concentrations. Synergistic effects due to temperature increase and
changes in nutrient concentrations will be considered in some experimental setups.
Expected changes in the formation, sedimentation, and export of biogenic particles due to
changing mineral ballast will be assessed by means of experiments in pressure controlled
chambers. This will be complemented by determining CO2 effects on the regulation of DOM
release from primary producers and its impact on the bacterial community.
Acclimation will be described in terms of physiological responses. The genomic changes in
response to long-term exposure will be investigated to identify the time scales on which
adaptations can be achieved.
Data on responses of organisms will be built into models. This involves (1) an improvement of
the description of the soft-tissue pump in a global biogeochemical model and (2) integrating
ocean acidification sensitivities at the organism level into ecosystem modelling.
5.1.3. Projects under this Theme
Project 1.1: Acclimation versus adaptation in autotrophs (Thorsten Reusch)
Project 1.2: Turnover of organic matter (Anja Engel)
Project 1.3: Modelling biogeochemical feedbacks of the organic carbon pump (Birgit
Schneider)
References
Arrigo KR, (2007) Carbon Cycle Marine manipulations, Nature, 450: 491-492.
Engel A, Thoms S, Riebesell U, Rochelle-Newall E,Zondervan I, (2004) Polysaccharide aggregation as a potential sink of marine dissolved
organic carbon, Nature, 428: 929-932.
Eppley RW, Peterson BJ, (1979) Particulate organic matter flux and planktonic new production in the deep ocean, Nature, 282: 677-680.
Falkowski P, Barber RT, Smetacek V, (1998) Biogeochemical Controls and Feedbacks on Ocean Primary Production, Science, 281,(5374):
200-205; DOI: 10.1126/science.281.5374.200.
Giordano M, Beardall J, Raven JA (2005) CO2 concentrating mechanisms in algae: mechanisms, environmental modulation, and evolution.
Annu Rev Plant Biol 56: 99-131
Laws EA, Falkowski PG, Smith WO, Ducklow H, McCarthy JJ, (2000) Temperature effects on export production in the open ocean, Global
Biogeochem. Cycles, 14,(4): 1231-1246.
Riebesell U, Schulz KG, Bellerby RGJ, Botros M, Fritsche P, Meyerhöfer M, Neill C, Nondal G, Oschlies A, Wohlers J, Zöllner E, (2007)
Enhanced biological carbon consumption in a high CO2 ocean, Nature 450: 545-548
15
5.2.
Theme 2
Performance characters: Reproduction, growth and
behaviours in animal species
1. Which physiological mechanisms define sensitivity or tolerance of marine
animals to ocean acidification and how do they set or modify performance levels
and fitness?
2. Can acclimation capacity (gene expression capacity) for such mechanisms
explain physiological plasticity?
3. How does acclimation or adaptation to new levels of CO2 and temperature
affect organism performance?
4. In a comparison of species and their populations from temperate to polar
climates, do they differ in their sensitivity or capacity to resist ocean
acidification through acclimation or evolutionary adaptation? How do these
findings relate to differences in temperature and associated ocean
physicochemistry?
5. Which life stages of functionally important marine organisms are most sensitive
to ocean acidification and how does the level of sensitivity relate to the ontogeny
of physiological mechanisms?
Ecosystem effects of ocean acidification include those on metazoan life. However, while
ecosystem effects of warming trends have clearly been identified, those of ocean acidification are
still equivocal. Within the next decades, elevated CO2 levels are expected to affect marine water
breathing animals directly through effects on the physiology and performance of the individual
organism and indirectly through changes in food web structure. Emerging knowledge indicates
that sensitivity to elevated CO2 levels differs between animal phyla and species. It may also differ
depending on geographical latitude and associated climate conditions. Effects may be large and
potentially detrimental especially in life forms with a low metabolic rate, for examples among
calcifying benthic macroorganisms (Wood et al., 2008) or in the deep sea. This hypothesis is in
line with recent observations in habitats contaminated by natural CO2 emissions, e.g. in volcanic
areas around Ischia (Hall-Spencer et al., 2008). Initial findings suggest decreased growth and
enhanced mortality of sensitive species such as among molluscs or echinoderms in response to a
doubling of CO2 from pre-industrial levels to 560 ppm (Shirayama and Thornton 2005), a value
which is likely surpassed during this century. As effects of ocean acidification are expected on
top of those of ocean warming, studying ecosystem effects of OA will thus need to consider both,
the direct influence of ocean physicochemistry on individual organisms and species, and also the
CO2 dependent modulation of responses to temperature in particular.
For an in-depth cause and effect understanding, it is essential to unravel the physiological
mechanisms that define whole organism sensitivity to ocean acidification (e.g. Pörtner et al.
2004, Fabry et al. 2008) and especially those, which synergistically interact with temperature
effects on marine organisms (cf. Pörtner et al. 2005). A current hypothesis emphasizes a key role
for the capacity of acid-base regulation in defining sensitivity (Pörtner, 2008 for review).
16
Available data indicate that deviations of extracellular pH from its setpoint mediate several of the
observed whole organism effects. Theme 2 will investigate how and to what extent these
disturbances affect whole animal performance and how acclimation to various CO2 levels can
alleviate some of these effects. It will also address to what extent sensitivity to ocean
acidification interacts with thermal stresses and is shaped by the specialization of organisms on
ambient climate conditions according to latitude.
Fig. 5: Effects of ocean acidification and warming on marine ectotherms
– Theme 2 concepts and their implementation
Effects of anthropogenic ocean acidification on animal communities are expected on medium to
long time scales, due to the progressive accumulation of CO2 and due to long generation times. In
variable environments (e.g. upwelling areas, Feely et al., 2008) not only the drift in mean
physicochemical parameters but also enhanced amplitudes will require consideration in analyses
of CO2 effects. For an analysis of the interaction between specialization on various climates on
the one hand and sensitivity to ocean acidification on the other hand, physiological studies (of
e.g. performance and acid-base regulation) as well as investigations of gene expression patterns
and population structure will be carried out in species and populations living in a climate gradient
across latitudinal clines. Comparisons of fertilized eggs, juvenile and adult life stages will be
essential to identify the bottlenecks of sensitivity throughout ontogeny as well as their
physiological background. The physiological principles shaping performance may also control
calcification, and thus the shell growth of bivalves and other calcifiers over time. Performance
has been shown to link climate change to ecosystem effects of warming (Pörtner and Knust
2007). Performance characters like growth or foraging capacity are likely also involved in multistep processes affecting marine food webs. Here, species-specific responses and sensitivities
17
cause various species of an ecosystem to be affected differently, resulting in changes in species
interactions, population adaptability, food web structure and associated carbon fluxes. The work
will include species of coastal areas and relevant to fisheries and other marine services. Theme 2
also includes approaches to test and further develop mechanistic concepts and models of effects
of ocean acidification. This includes kinetic modelling of the mechanisms of ion and acid-base
regulation for an improved quantitative understanding of effects, to generate a basis for a more
comprehensive, mechanism based modelling approach.
Theme 2 aims at a better understanding of the physiological underpinning of specific and
combined CO2 and temperature effects on marine water breathing animals and on ecosystem
function. Lab and mesocosm experiments will provide insight into specific sensitivities,
acclimation and adaptation of marine invertebrates and fish to CO2 and temperature under present
CO2 concentrations as well as under levels twice (as projected for 2100) and three times the
present concentration. We expect to complement our knowledge of the physiological mechanisms
responding to changing CO2 levels and to changing body fluid and tissue acid-base variables,
considering the specific patterns of tissue functioning and its regulation. As a result, a more
comprehensive picture is expected of the mechanisms involved in shaping sensitivity,
acclimation and adaptation to various CO2 accumulation scenarios. Furthermore, consideration of
the projected temperature increase in our experiments will lead to a better understanding of the
synergistic effects of warming and acidification as well as the underlying mechanisms. The
generalized view of physiological principles should also help to identify the master variables
mediating the effects of ocean acidification (such as e.g. extracellular pH) and to comprehend
their integrating role at various levels of biological organization.
Mechanisms contributing to the set-points of acid-base regulation will be identified and their
contribution quantified in experimental and kinetic modelling studies. Acclimation will be
described in terms of the longer term adjustments in the functioning of mechanisms and
processes as well as in terms of the underlying genomic changes involving transcriptional and
translational processes. The time scales of acclimation and adaptation should become accessible
from long-term exposures and from comparisons of species and populations in various
temperatures and thus CO2 concentration regimes along a latitudinal cline between temperate and
polar areas.
Insight in the responses at organism level throughout ontogeny as well as at population level will
build the basis for identifying the CO2 and temperature induced modulation of key processes such
as recruitment and calcification which are crucial for a population’s role in ecosystem
functioning. The compilation of CO2 and temperature impacts on animal performance at different
trophic levels, e.g. from herbivores to top predators will facilitate the development of scenarios of
future marine ecosystem functioning. Such integrated assessments of sensitivities to ocean
acidification would also become available for future ecosystem modelling.
Project 2.1: Effects on grazers and filtrators (Thomas Brey)
Project 2.2: Long-term physiological effects on different life stages of benthic crustaceans
(Felix Mark)
Project 2.3: Effects on top predators (fishes, cephalopods) (Catriona Clemmesen)
18
References
Fabry VJ, Seibel BA, Feely RA, Orr JC (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICES J Mar Sci 65:
414–432.
Feely RA, Sabine CL, Hernandez-Ayon JM, Ianson D, Hales B (2008) Evidence for upwelling of corrosive "acidified" water onto the
continental shelf. Science 320: 1490 – 1492.
Hall-Spencer J.M., Rodolfo-Metalpa R, Martin S, Ransome E, Fine M, Turner SM, Rowley SJ, Tedesco D, Buia MC (2008) Volcanic carbon
dioxide vents show ecosystem effects of ocean acidification. Nature, doi:10.1038/nature07051.
Pörtner HO (2008) Ecosystem effects of ocean acidification in times of ocean warming: a physiologist’s view. Mar. Ecol. Progr. Ser. in review
Pörtner HO, Langenbuch M, Reipschläger A (2004) Biological impact of elevated ocean CO2 concentrations: lessons from animal physiology
and earth history. J. Oceanogr. 60: 705-718.
Pörtner HO, Langenbuch M and Michaelidis B (2005) Synergistic effects of temperature extremes, hypoxia, and increases in CO2 on marine
animals: From Earth history to global change, J. Geophys. Res. 110: C09S10
Pörtner HO, and Knust R (2007) Climate change affects marine fishes through the oxygen limitation of thermal tolerance. Science 315: 95 97.
Shirayama, Y., and H. Thornton (2005), Effect of increased atmospheric CO2 on shallow water marine benthos, J. Geophys. Res., 110,
C09S08, doi:10.1029/2004JC002618.
Wood HL, Spicer JI, Widdicombe S (2008) Ocean acidification may increase calcification rates, but at a cost. Proc R Soc B 275:1767-1773.
5.3. Theme 3 Calcification: Sensitivities across phyla and ecosystems
5.3.1.
Theme summary
1. What are the cellular mechanisms of calcification and decalcification in different marine
organisms, particularly with regard to ion transport to and from calcification sites?
2. How will OA and pH stress affect calcification at the level of organisms, communities
and ecosystems? Will concurrent temperature change modulate these effects?
3. What changes will occur in the ultra-structure, trace element partitioning, and isotopic
signature of the shells and skeletons of calcifyers in response to pH stress?
4. How does the changing water column chemistry influence carbonate dissolution and
deposition in sedimentary systems?
5. Are past OA events (e.g. in the Cenozoic) useful analogues for projected future OA in
their effects on calcifying organisms? Is the performance and sensitivity of past and
present calcifyers comparable, and does this information help to assess the future?
Calcification, the precipitation of calcium with carbonate, is influenced by the current increase in
atmospheric CO2 levels and its concurrent gradual decrease in ocean pH. Since acidification leads
to a decrease of the carbonate ion concentration, ocean acidification causes a decrease of the
carbonate saturation sate. Hence we can expect a future reduction in calcification rates, or even
net decalcification. Indeed reduction of calcification rates have been observed upon acidification
in several marine calcifying groups, but not in all. This is indicative for a variety of calcification
mechanisms. Calcification is a proton-generating process, and decalcification is proton
consuming. Thus, these processes are typically coupled to either proton consuming (calcification)
or proton producing (decalcification) processes. Conversely, calcification and decalcification can
buffer other pH changing processes, and will thus to some extend buffer the oceanic pH.
Biological calcification always occurs in more or less isolated microenvironments, in which the
carbonate and/or calcium concentrations are changed by biological activity. These changes can
be driven by ion pumps in specialized transport tissue, e.g. for Ca2+ or H+, which is typical for the
19
highly controlled calcification in corals, foraminifera, bivalves and coccolithophores.
Alternatively, calcification can be a side effect of metabolic activities such as photosynthesis,
occurring in a matrix with mass transfer limitation (a sediment or microbial mat). We will
investigate if some calcification mechanisms are more sensitive than others, and the extent to
which decalcification can buffer the decrease in oceanic pH. We will further investigate if
decalcification can contribute to pH buffering of the oceans. We will finally investigate if the
oceanic pH may leave signatures in the biogenic carbonates, and learn from acidic events in the
past what the effect is on biodiversity on calcifying nano-plankton.
Differences in calcification mechanisms
may be responsible for observed
differences in CO2/pH sensitivities of
marine calcifiers. Left is a possible
calcification
scheme
by
actively
calcifying organisms, which depend on
transmembrane pumping of ions to and
from a calcification site, such as shells
and skeletons. Right is a scheme of
calcification by diffusion-controlled
calcification. A calcifying microenvironment is created by the combined
effect of diffusion resistance and
metabolic activity, like photosynthesis.
Fig. 6: Groups of calcifying organisms and processes studied in Theme 3
5.3.2.
Progress expected
Within this theme we will try to elucidate how pH affects calcification rates of two principally
different mechanisms. We will study (1) calcification driven by membrane transport processes,
i.e. fully organism-controlled, and (2) calcification as a side reaction of photosynthesis, i.e.
environmentally controlled. In both cases the calcification occurs in a more or less shielded
space, but in the first case it is shielded by membranes and tissue within the calcifying organisms,
in the second case calcification is rather an encrustation outside the organism, and the transport
limitation is by diffusion.
20
Hypotheses: As the first mechanism involves considerable energy input from the actively
calcifying organisms, which will increase upon acidification, we expect significant effects on the
ecological fitness of calcifiers and on the biodiversity of ecosystems with calcifying
communities. In the second case calcification does not require metabolic energy, and reduced
calcification does not present an ecological disadvantage for the organisms that drive
calcification, and will not have an effect on their biodiversity. We expect to have a better
understanding on this issue through the projects 3.1 and 3.2. Secondly, we will investigate if pH
changes leave a signature in the biogenic carbonates, and whether the isotope and trace metal
signatures can be used to better understand the calcification process. More specifically, it will be
tested in project 3.3 whether calcification in corals and foraminifera is indeed a result of
enveloping seawater parcels in vacuoles in which subsequently the chemistry is shifted towards
an environment where carbonates precipitate. This will enhance understanding of the chemical
signatures of biogenic carbonates. We will investigate in project 3.4 in how far calcification in
sediments and other matrices is resilient against ocean acidification, and construct a diffusionconversion model describing calcification and decalcification. This model will include the
carbonate system, pH, photosynthesis and respiration. In project 3.5 we will investigate if several
acidic events in the past have left traces in the carbonate skeletons. The calcification driven by
photosynthesis occurs in sediments and mats where transport is diffusionally limited. This to
some extend uncouples the microenvironment from the seawater.
5.3.3.
Projects under this Theme
Project 3.1:
Cellular mechanisms of calcification (Frank Melzner)
Project 3.2:
Calcification under pH-stress: Impacts on ecosystem and organismal levels
(Ralph Tollrian)
Project 3.3:
Ultra-structural changes and trace element / isotope partitioning in calcifying
organisms (foraminifera, corals) (Jelle Bijma)
Project 3.4:
Micro-environmentally controlled (de-)calcification mechanisms
(Michael Böttcher)
Project 3.5:
Impact of present and past ocean acidification on metabolism,
biomineralization and biodiversity of pelagic and neritic calcifiers
(Adrian Immenhauser)
21
5.4.
Theme 4
Species interactions and community structure in a
changing ocean
1. What is the role of differential sensitivities to OA at the community, species and intraspecific (ontogenetic stages, genotypes) level? Are there emerging properties resulting
from organism interactions that are not visible from single-species investigations
alone?
2. Does the community structure change as a consequence of OA and how do shifts in
competitive abilities of benthic and pelagic organisms affect community structure?
3. What is the role of OA induced changes in food quality and quantity in primary
producers to higher trophic levels?
4. To which extent will energy transfer between lower and higher trophic levels change as
a result of changing competitive interactions and/or changes in the feeding
environment?
5. Do different types of communities (benthic – pelagic, microbial – macrobial) react
differently to acidification/warming stress?
Observed effects of low pH and/or high CO2 conditions are mostly based on experiments with
single species. Hence, not much is known about how these factors affect interactions between
species and, through that, communities. Indeed, the projected shifts in pCO2 and pH in many
species often only slightly impact performance and fitness of a given species, and many studies
find reactions in single species experiments only with very high CO2 concentrations or at
unrealistically low pH (e.g. Mayor et al. 2007). However, as shown for other stressors
(Christensen et al. 2006) ensuing modifications of species interactions may substantially amplify
or buffer the original stress (Wahl 2008). How environmental stress spreads through a
community via shifts in composition and interaction is still very much an open question.
Theme 4 will therefore address the pivotal role of interaction modulation in close cooperation
with a number of projects from other themes which, in contrast, focus on single species reactions
to ocean acidification. Thus, Theme 4 is the logical extension of these projects, placing the
responses of individual organisms to OA into a community and ecosystem context. We expect to
find effects of OA on community structure and interaction that are not visible when studying
single organism reactions. Hence, we will focus on shifts in competitive and trophic interactions
as well as on community structure. Organisms will vary considerably in their reaction to ocean
acidification. As a consequence, formerly superior competitors may be weakened, as is the case
in interactions between calcifying and non-calcifying species (e.g. Kuffner et al. 2007), or
relative susceptibility to predation may change (Swanson & Fox 2007). Furthermore, there may
be strong selective pressures within species, if susceptibility to stress differs between genotypes
or between ontogenetic stages. At the unicellular level, less sensitive clones or strains may
become more dominant under ocean acidification, and sensitive rare populations may disappear.
In multicellular species, the most sensitive ontogenetic phase will determine survival, and genetic
diversity may determine the fate of a population under OA.
22
The changes in species composition on one trophic level will obviously affect the transfer of
energy and matter to higher trophic levels. Shifting species composition at the base of the food
chain may represent different quality feeds for predators, which may result in a complete
restructuring of the trophic web. Also, direct effects of increased CO2 availability may change the
quality of organisms as food for higher trophic levels. Higher carbon availability will typically
result in higher carbon to nutrient ratios in primary producers. This has on the one hand been
linked to decreased toxicity of some dinoflagellates (Parkhill & Cembella 1999), with the
potential of higher palatability of previously noxious algae, but on the other hand to a decrease in
the quality as food for higher trophic levels (Malzahn et al. 2007).
So, if we are to understand the effects of OA on ecosystem structure and function, it is essential
that we understand the shifts of interaction mode or strength (Tortell et al. 2002), because the
typical non-linearity of the ensuing effects has the potential to cause regime shifts in marine
ecosystems. To date, regime shifts have mainly been linked to climate forcing, but we expect
ocean acidification – amplified by interaction modulation - to have the same potential.
Approach:
Fig. 7: Compartments and interactions studied in Theme 4
In Theme 4, we will investigate how interactions between and within species in benthic [4.1.1.4.1.4] and pelagic [4.2.1-4.2.2] communities shift under the influence of OA. We investigate
changes in trophic grazer- alga interactions in the benthos [4.1.1], and competitive interactions
between sessile organisms [animals 4.1.2 and plants 4.1.3]. Moreover, we will investigate the
effects of ocean acidification on bacterial communities, both directly, as well as a result of altered
excretion products of algae and herbivores faced with resources of different quality [4.1.4 for the
benthos, 4.2.1 for the pelagic zone]. Competition between pelagic microalgae will be studied in
4.2.2, and the resulting prey community will be fed to pelagic herbivorous grazers in 4.2.1
23
Theme 4 will further our understanding of between- and within-species interactions under OA
stress. We expect that after a successful completion of the project we will be able to define the
expected impacts of ocean acidification that go beyond single species reactions but are a result of
changing interactions between and within species. As stated above, only by taking these
interactions into account we will be able to truly appreciate the potential impact of OA. More
specifically we expect progress in the following areas:
1. Assessment of the role of differential sensitivities to OA at the intra-species (ontogenetic
stages, genotypes), and species level. Populations and communities can only re-structure and,
thus, adapt if their respective components differ in sensitivity towards the stressor(s). In close
cooperation with appropriate projects in the other themes we will quantify how the responses
to stress varies within species between life stages (not applicable for unicellular organisms),
genotypes/strains, and between species in the focal communities.
2. Shifts in competitive abilities of model organisms. Co-culturing of organisms exhibiting
differing stress sensitivity (at the ontogenetic, genetic, species level) allows the quantification
of the stress-induced shifts in competitiveness as compared to a low-stress situation. Strong
shifts will lead to fast changes in relative abundance of the components in the focal
communities: benthic and pelagic microbes, microalgae, macroalgae, macroalgae/sea grass,
sessile invertebrates. This issue will highlight the role of genetic and functional diversity for
the adaptability of populations and communities. Further, prolonged co-culturing of simple
communities in micro- and mesocosms will produce re-structured communities adapted to
the OA scenario applied. Since such a re-organization depends on the gene and species pool
available, and on the number of generations allowed, and both these conditions can not be
simulated realistically in experiments we do not pretend to simulate the ecosystem changes
which will happen in the future. However, the kind of shifts among functional groups – e.g.
macroalgae vs. seagrass or calcifiers vs. non-calcifiers – will improve our capacity for
plausible projections.
3. Quantification of the changes in community structure and community services. The structural
shifts driven by OA will lead to changes in the functions within communities. Thus, an
altered prey community will differ with regard to food quality and quantity. How this affects
structure and dynamics of primary or secondary consumers, and storage in or flow of
energy/matter between different strata of the system, will be a further outcome of Theme 4.
Ultimately, we aim to make predictions of how OA is likely to affect issues such as surface
water quality or the yield of economically relevant species.
4. We will strive to incorporate the findings on direct and indirect OA impacts at the different
organisational into descriptive and/or predictive models.
Project 4.1: OA impacts on interactions in and structure of benthic communities
(Martin Wahl)
Project 4.2: OA effects on food webs and competitive interactions in pelagic ecosystems
(Maarten Boersma)
24
References
Christensen MR, Graham MD, Vinebrooke RD, Findlay DL, Paterson MJ, Turner MA (2006) Multiple anthropogenic stressors cause
ecological surprises in boreal lakes. Glob Chan Biol 12:2316-2322
Kuffner IB, Andersson AJ, Jokiel PL, Rodgers KS, Mackenzie FT (2007) Decreased abundance of crustose coralline algae due to ocean
acidification. Nature Geosci 1:114-117
Malzahn AM, Aberle N, Clemmesen C, Boersma M (2007) Nutrient limitation of primary producers affects planktivorous fish condition.
Limnol Oceanogr 52:2062-2071
Mayor DJ, Matthews C, Cook K, Zuur AF, Hay S (2007) CO2-induced acidification affects hatching success in Calanus finmarchicus. Mar
Ecol Prog Ser 350:91-97
Parkhill J, Cembella A (1999) Effects of salinity, light and inorganic nitrogen on growth and toxigenicity of the marine dinoflagellate
Alexandrium tamarense from northeastern Canada. J Plankton Res 21:939-955
Swanson AK, Fox CH (2007) Altered kelp (Laminariales) phlorotannins and growth under elevated carbon dioxide and ultraviolet-B
treatments can influence associated intertidal food webs. Glob Chan Biol 13:1696-1709
Tortell PD, DiTullio GR, Sigman DM, Morel FMM (2002) CO2 effects on taxonomic composition and nutrient utilization in an Equatorial
Pacific phytoplankton assemblage. Mar Ecol Prog Ser 236:37-43
Wahl M (2008) Ecological modulation of environmental stress: interactions between UV radiation, epibiotic snail embryos, plants and
herbivores. J Anim Ecol 77:549-557
5.5.
Theme 5
Integrated assessment: Sensitivities and uncertainties
Objectives and overarching questions
1. Synthesize information obtained in Themes 1 to 4 in order to achieve an integrated
understanding of biological responses to ocean change.
2. Develop a framework for integrating ocean acidification sensitivities at the organism
level into ecosystem models.
3. What are the integrated effects of ocean acidification and warming on ecosystem to
global ocean scales?
4. What are the critical threshold levels (‘tipping points’) of ocean acidification for
irreversible ecosystem changes?
5. What is the most suitable definition of dangerous ocean acidification in terms of the
goods and ecosystem services lost due to OA?
German Advisory Council on Global Change (WBGU, Berlin 2006) recommends a guard rail for
future ocean pH decrease of 0.2 units as a margin of safety according to the precautionary
principle. This suggestion is motivated by the intention of avoiding an aragonite undersaturation
in the ocean surface layer. As stated in the report, the tolerable window for ocean acidification
defined by WBGU presently relies on an extremely small data base. In fact, rather than using the
limited data on observed biological consequences of ocean acidification, the WBGU reaches its
recommendation on the basis of projected changes in water chemistry (aragonite saturation state).
While this is an appropriate approach in view of the scarcity of biological information, there is a
clear need to establish a reliable data base on tolerance levels for ocean acidification in key
groups of ocean-acidification sensitive marine organisms in order to reach a more informed
recommendation.
25
Fig. 8: Schematic diagram of Theme 5. Proposed projects are indicated (5.1, 5.2, 5.3).
Theme 5 of BIOACID will take the challenge of integrating the information gained under
Themes 1 to 4 in order to identify the potential thresholds associated with ocean acidification.
Uncertainties, probabilities and risks to the marine environment have to be assessed as well as
their feedback to climate system. This will be achieved through a meta-analysis of process
studies and process parameterisations, and by combining models and data in a data-assimilative
framework. In return, feedback from the modelling work will inform the experimental work in
BIOACID about uncertainties in models and the relevant process parameterisations.
During the first 3-year phase of BIOACID, our main aim is to develop and establish the tools that
will allow us to fulfil the BIOACID synthesis needs. For the three subprojects proposed here, the
synthesis tools to be established within BIOACID range from meta-analysis techniques over
regional and global numerical ecosystem models to economic methods of integrated assessment.
These tools will help to better understand ongoing changes in chemical and biological state of the
North Sea from alkalinity fluxes originating from the Wadden Sea over a synthesis model that
integrates OA sensitivities at organism level into a North Sea ecosystem model (5.1) to an
economical impact assessment. (5.3). Newly developed assessment tools will also be used to
improve parameterisations of calcium carbonate production in global biogeochemical climate
models (5.2). By investigating the combined effects of variations in temperature and ocean
acidity, such parameterisations will allow to put better constraints on possible threshold levels on
ocean acidification in a warming world.
We subdivide expected progress into three categories, development of synthesis tools, synthesis
of research results, and large scale modelling ocean acidification impacts and biogeochemical
26
feedbacks to carbon cycling and climate. Categories which reflect the major research aspects
covered by projects of Theme 5 as well as collaborative research carried out together with
projects from Themes 1-4 (see table 0.1, Section 11.1 for details).
Development of synthesis tools
This theme will
•
carry out supplementary model development and evaluation of a shelf sea ecosystem
model (ECOHAM) for studying ocean acidification effects on the ecosystem level as
well as alkalinity exchange of the North Sea with Wadden Sea and open ocean,
•
implement proposed parameterization of open ocean calcium carbonate production,
export and dissolution into a tracer transport matrix rapid spin-up model system,
•
initiate a Bayesian meta-analysis of BIOACID experimental findings
•
further develop the ecological-economic viability-method towards a general approach for
integrated assessment of human actions influencing ocean acidification and the
consequences for human well-being that takes uncertainties about future development
into account
Synthesis of research results
This theme will
•
critically review existing parameterisations of calcification currently used in large-scale
biogeochemical climate models,
•
provide a review of experimental findings on temperature and pCO2 sensitivity of
calcium carbonate production, its export and dissolution to be put in perspective with
model parameterisations.
Large scale modelling ocean acidification impacts and biogeochemical feedbacks to carbon
cycling and climate
This theme will
•
provide an evaluation and quantification of alkalinity fluxes from the Wadden Sea to the
North Sea and their role in buffering ocean acidification from anthropogenic CO2
invasion. Response of calcifying and non-calcifying primary producers to pH changes
will be addressed by extrapolating results from mesocosm studies and future scenario
model simulations (pCO2=1000µatm) studying critical ecological and biogeochemical
aspects,
•
quantitatively assess the ability of current parameterisations of calcification in global
biogeochemical models to reproduce observed alkalinity fields, and to suggest improved
parameterisations that can reliably predict the response of pelagic calcium carbonate
production to variations in both temperature and ocean carbonate chemistry
•
by means of the viability-method, assess the impacts of different human actions on ocean
acidification, taking into account the uncertainties about the future development of
acidification and the exact impact on cod recruitment. This modelling approach will take
into account measures to mitigate acidification or to adapt the management of the North
Sea cod fishery.
27
Project 5.1: Impact of Alkalinity fluxes from the Wadden Sea on the carbon cycle and the
primary production in the North Sea, (Johannes Pätsch)
Project 5.2: Evaluating and optimising parameterisations of pelagic calcium carbonate
production in global biogeochemical ocean models, (Andreas Oschlies)
Project 5.3: Viability-method for the impact assessment of ocean acidification under
uncertainty (Martin Quaas)
6.
Management structure and procedures
A good organizational structure and an effective labour division based on clearly identified tasks
and accompanied with a well-defined decision-making system will be put in place in order to
fulfil the scientific objectives of BIOACID, to ensure an efficient work-flow and to reduce risks
of failure. An overall organizational chart depicting the communication links is depicted in
Figure 9 and is described in further detail below. The management procedures clearly define the
consortium tasks, responsibilities and lines of command in a transparent decision-making
structure.
Fig. 9: Management structure and executive and advisory bodies.
28
6.1.
Project Coordinator and Project Office
The Coordinator of BIOACID will be Ulf Riebesell at the Leibniz Institute of Marine Sciences,
Kiel. Hans-Otto Pörtner at the Alfred-Wegener Institute, Bremerhaven will act as Deputy
Coordinator. The coordinator (or in the case of his absence the Deputy Coordinator) will be
responsible for the day-to-day management of BIOACID and contact with the Ministry of
Education and Research (BMBF) and the Projektträger Jülich (PTJ). He will ensure that the
project runs in compliance with the requirements of the contract (administrative, operational and
scientific aspects). The Project Management Office will be located at IFM-GEOMAR. It will
act upon decisions taken by the Executive Board (EB), the Scientific Steering Committee
(SSC) and the Member’s General Assembly (MGA).
6.2.
Executive Board (EB)
The Executive Board (Table 2), chaired by the Project Coordinator, will prepare decisions to be
approved by the SSC and MGA and implement decisions on executive management. It will meet
annually at the consortium meeting and will communicate regularly by phone and/or e-mail.
Major decisions and recommendations will be made at the meetings with one vote per member of
the EB. In addition to the meetings, the Executive Board will communicate via the project
website. All communication with other Committees will be through the Project Office.
Table 2: Composition of the executive board.
Executive board Members
Coordinator:
Ulf Riebesell (IFM-GEOMAR)
Theme leaders: Maren Voss (IOW), Hans-Otto Pörtner (AWI), Dirk de Beer (MPI Bremen),
Maarten Boersma (AWI), Andreas Oschlies (IFM-GEOMAR)
The responsibility of the EB will include:
• ensuring that sufficient management is in place for monitoring the scientific work in
project and perform quality control
• enforcing decisions of the SSC and MGA
• ensuring the preparation of reports and work plans
• informing the SSC about project progress, any problems and risks encountered and any
change in strategy
6.3.
Scientific Steering Committee (SSC)
The SSC (Table 3) will advise on the overall scientific policy, direction and management of the
project to be decided by the MGA. It will meet annually at Consortium meetings and be chaired
by the Coordinator. All communication with the MGA will normally be through the Project
Office.
29
Table 3: Composition of the scientific steering committee (SSC).
SSC
Members
Coordinator:
Ulf Riebesell (IFM-GEOMAR)
Deputy coordinator: Hans-Otto Pörtner (AWI)
Theme leaders:
Maren Voss (IOW), Hans-Otto Pörtner (AWI), Dirk de Beer (MPI
Bremen), Maarten Boersma (AWI), Andreas Oschlies (IFM-GEOMAR)
Project leaders:
Michael Diepenbroek (U Bremen), Magnus Lucassen (AWI), Thorsten
Reusch (IFM-GEOMAR), Anja Engel (AWI), Birgit Schneider (CAU
Kiel), Thomas Brey (AWI), Felix Mark (AWI), Catriona Clemmesen
(IFM-GEOMAR), Frank Melzner (IFM-GEOMAR), Ralph Tollrian (U
Bochum), Jelle Bijma (AWI), Martin Wahl (IFM-GEOMAR), Maarten
Boersma (AWI), Johannes Pätsch (U Hamburg), Andreas Oschlies (IFMGEOMAR), Martin Quaas (CAU Kiel)
Additional member: Antje Boetius (MPI Bremen), as member of the original planning group
The responsibilities of the SSC will include:
•
acting on the initiative of the Executive Board (EB) on issues or problems relating to the
progress towards the fulfilment of the scientific objectives
•
assessing scientific progress against the objectives and, where necessary, make
recommendations to the EB
•
providing advice on any call for participants or partners that might be needed to finalize
the project’s objectives
•
giving recommendations to the Executive Board on any scientific aspects it foresees as
requiring ethical considerations
6.4.
Members’ General Assembly (MGA).
The MGA consists of all project and sub-project PIs and will be the overall decisive body of the
Consortium. At annual meetings, it will give scientific advice to the Steering Committee on the
most important issues of the project. If needed, extraordinary meetings will be held. The MGA
will be responsible for all major formal decisions regarding project strategy and any change to
the consortium. Decisions in the MGA need two thirds of the votes present at the meeting. The
MGA will act upon proposals from the Executive Board and decide on the following issues:
30
•
any major change in the scientific plans
•
any major budget reallocation between partners
•
any alteration of the management structure and management procedures
•
any change in the advisory bodies
7.
Data management and dissemination
7.1.
Data management
The data management in BIOACID will be carried out by the World Data Center for Marine
Environmental Sciences WDC-MARE (www.wdc-mare.org) at the University of Bremen. WDCMARE uses the information system PANGAEA (Publishing Network for Geoscientific &
Environmental Data – www.pangaea.de), which is a system for acquisition, processing, long-term
storage, and publication of geo-referenced data related to all earth science fields. Specifically,
WDC-MARE will be responsible for
1. Coordination of data capture, integration and quality control activities for the five
BIOACID Themes.
2. Archiving and publishing data sets and data collections online and as offline products
(DVD) using persistent Digital Objects Identifiers (DOI).
3. Implementation of the BIOACID data infrastructure
-
enabling a distributed storage of observational and model simulation data within a
common networked structure, and
-
establishing a robust and long lasting data network which can be extended by or
integrated into ongoing projects and programmes.
4. Maintain the website and data portal for BIOACID.
(For details on data managing see Theme 0.2 below). A half-time data manager position will be
established by WDC-MARE at the University of Bremen.
7.2
Dissemination
Dissemination will include a multifunctional web service comprising a publicly accessible area to
present the activities, techniques, and results of the project and a secure (password protected) area
for internal project communication. This website will be maintained and administrated by the
data manager (see project 0.2).
Jointly with the administration of the Coordinator Project Office, all BIOACID partners
contribute to
• preparing a series of fact/information sheets and high level presentation packages aimed
at policy makers’ needs
• generate TV-quality video material on BIOACID research activities for distribution to
interested TV channels and to produce a 8-10 minutes video clip on the BIOACID
project and the emerging problem of ocean acidification (see project 0.1)
• communicate with society through on-line debates, web site, media interviews, press
releases, events and articles in popular magazines, and consortium consensus responses
to frequently asked questions
• scientific publications in international refereed journals and presentation at conferences,
short reports and flyers to authorities, public bodies, agencies and organisations
31
Flyers and posters will be available on the BIOACID web site for members to download and
distribute or display. Synergies with other national and EU projects (see 9 National and
international Cooperations) will be formed to ensure effective exchange of information.
8.
Infrastructure development, training and transfer of know how
8.1.
Infrastructure development
BIOACID will support the development of instrumentation and infrastructure urgently needed in
ocean acidification research which will become of direct and general use to all BIOACID
partners. In particular, two activities are proposed in this regard facilitating measurements of CO2
partial pressure and improving CO2 perturbation-based experimentation.
1. Development and maintenance of aquarium culture facilities and research infrastructure
needed for long-term incubations in the form of a re-circulating system. The proposed
system, to be developed and installed at the AWI in Bremerhaven, is expected to
establish and maintain baseline conditions in larger experimental set-ups at pre-industrial
levels and various CO2 scenarios (for details see project 0.3.1.). Time permitting the
system will be open access to all BIOACID partners.
2. Development of new chemical optical sensors and instrumentation for the determination
of carbon dioxide partial pressure (pCO2) in fluids of marine organisms and in marine
environments based on the dual lifetime referencing (DLR) method patented by PreSens.
Emphasis will be put on the development of pCO2 sensors with appropriate resolution in
the trace concentration range (380-1900 ppm in seawater, 380-3000 ppm in body fluids).
The performance of the newly developed instrumentation will be optimised according to
the experimental applications provided by project partners with prototype pCO2 sensors
(for details see project 0.3.2.).
8.2.
Training and transfer of know how
BIOACID will embrace a wide range of scientific disciplines, many with different approaches to
monitoring the carbonate system and designing perturbation experiments. It is of vital importance
for a large integrating project such as BIOACID to have common understanding on
methodological and reporting procedures to facilitate comparison between investigators.
Moreover, several BIOACID partners are international leaders for specific techniques which are
of direct relevance to ocean acidification research. To this end, a series of training activities will
be designed to ensure high data quality and inter-comparability and open up state-of-the-art
technology to BIOACID partners. All workshops are intended to also facilitate the exchange and
collaboration of young researchers employed in BIOACID and – space permitting – from outside
the consortium. In this respect, close collaboration with interdisciplinary graduate schools such as
the Integrated School for Ocean Sciences (ISOS) of the Excellence Cluster “The Future Ocean”
and the Excellence Cluster “GLOMAR” (Global Change in the Ocean Realm) is envisioned.
With over 30 Ph.D. positions and several Post-Doc positions offered in BIOACID, the project
will also make a major contribution to the education of young scientists in a wide range of marine
disciplines, from molecular biology, physiology, ecology, to biogeochemistry and
palaeoceanography.
32
Table 4: Training workshops
Organizer
Institution
Titel
Duration
Dirk de Beer
MPI Bremen
Microsensor applications
10 days
Kai Schulz
IFM-GEOMAR
Seawater carbonate chemistry
2 days
Anton
Eisenhauer
IFM-GEOMAR
Isotope geochemistry and laser-ablationtechniques
4 days
Martin Wahl
IFM-GEOMAR
Experimental design
3 days
Franz Sartoris
AWI
Physiological approaches to body fluid
5 days
physicochemistry and acid-base regulation
9.
International and National Cooperation
German scientists, including members of the BIOACID planning group, have played a key role
in recent reports or working groups related to ocean acidification such as: The Royal Society
Working Group on Ocean Acidification "Ocean acidification due to increasing atmospheric
carbon dioxide" (London 2005), the OSPAR intercessional working group on ocean acidification
(2006), IPCC's fourth Assessment Report, Special IPCC report on Carbon Dioxide Capture and
Storage (2006), Special Report of the German Advisory Council on Global Change "The Future
Oceans – Warming Up, Rising High, Turning Sour" (2006), NSF-NOAA-USGS Report "Impacts
of Ocean Acidification on Coral Reefs and Other Marine Calcifiers" (2006), the IGBP-SCOR
Fast Track Initiative on “Ocean acidification, atmospheric CO2 and ocean biogeochemistry:
modern observations and past experiences“(Lamont, 2006), and related to ocean biodiversity
such as the DIVERSITAS Marine Biodiversity Cross Cutting Network and Census of Marine
Life programmes.
Members of the BIOACID planning group have also organized or co-organized meetings,
workshops, special sessions or edited special issues of journals on ocean acidification. For
example: The Ocean in a high CO2 World I and II, (I: Paris, May 2004, II: Monaco, October
2008), ocean acidification sessions at the Ocean Sciences Meeting of the American Society of
Limnology and Oceanography (ASLO) and at the annual conference of the European
Geophysical Union (EGU) (Vienna, April 2006), a press conference on ocean acidification at the
33
EGU conference (Vienna, April 2006), a special issue of the Journal of Geophysical Research
"The Ocean in a High-CO2 World" (2005) and a special issue of the journal Biogeosciences
“PeECE: Pelagic ecosystem CO2 Enrichment Studies” (2007). Members of the BIOACID
planning group were invited to attend and contribute to the planning of U.S. and UK national
programmes on ocean acidification, and will remain to be involved in further developments of
OA programmes in these countries.
9.1.
International Cooperation
Close coordination of BIOACID activities with European research on ocean acidification will
be ensured through high level participation of members of the BIOACID planning group in
relevant EU projects, e.g. as co-coordinator, theme and work package leaders of the European
Project on Ocean Acidification (EPOCA) and as leading scientists of the EU coordinated project
Marine Carbon Sources and Sinks (CARBOOCEAN). As outlined by the coordinator of EPOCA,
Dr. Jean-Pierre Gattuso (see p. 36): “The contribution from the European Commission (6.5 M€)
is relatively limited for a 4-year project comprising 29 laboratories and more than 100 permanent
researchers. Hence, its partners must complement the EU contribution with national funding in
order to fulffill the objectives of the project and maintain the leadership that the EU currently has
in the field of ocean acidification.”
UK’s Natural Environment Research Council (NERC) has just launched its new five-year
strategy which includes seven science themes and within one of these (Earth System Science) is a
specific challenge “to understand changes in ocean ecosystems in response to ocean
acidification”. During a preparation workshop of NERC’s five-year strategy, initial steps have
been taken to aim for a UK-German partnership in developing a joint programme on ocean
acidification (see letter by NERC Theme leader Prof. Tim Jickells, p. 37). The planned
cooperation with UK NERC is consistent with BMBF policy as stated at the German-British
conference on “Climate Change: Meeting the Challenge together” held in November 2004 in
Berlin ending with a commitment to strengthening German-British cooperation in building up
joint activities in climate research.
9.2.
National Cooperation
BIOACID will be closely coordinated with National research programmes focussing on related
aspect, in particular the DFG Priority Programme AQUASHIFT (The Response of Aquatic
Ecosystems to Climate Change) and the BMBF Verbundprojekt SOPRAN (Surface Ocean
processes in the Anthropocene).
AQUASHIFT (coordinator: U. Sommer, IFM-GEOMAR, Kiel) assesses the effect of global
warming on aquatic ecosystems. The anthropogenically enhanced greenhouse effect and the CO2induced ocean acidification are two sides of the same coin. Their impacts on marine ecosystems
will go hand in hand, with a high probability of synergistic effects. Collaboration between
AQUASHIFT and BIOACID opens the opportunity to address such synergistic effects, with the
aim to allow a more realistic representation of future ocean conditions and the corresponding
ecosystem responses.
34
SOPRAN (coordinator: D. Wallace, IFM-GEOMAR, Kiel) focuses on biogeochemical processes
operating within and close to the surface ocean as drivers of ocean-atmosphere material
exchanges. Particular attention is given to the impacts of climate induced changes in surface
ocean processes (upwelling, mixing, light, nutrient supply) and of changes in atmospheric
composition (e.g. increased CO2, dust) on surface ocean biogeochemistry and air-sea exchange of
climatically relevant gases. Due to its focus on the organismal and community level, BIOACID
will provide valuable input to SOPRAN. With its focus on a wide spectrum of global change
forcings, SOPRAN will allow BIOACID to upscale its results to the ocean-wide level.
Specifically, Theme 1 Primary production, microbial processes and biogeochemical feedbacks
and Theme 5 Integrated assessment: Sensitivities and uncertainties of BIOACID will serve as
cross-over with SOPRAN, facilitating cross fertilization between the two programmes and
opening opportunities for joint investigations, particularly at the level of field observations and
model simulations.
BIOACID includes project leaders of the two marine Excellence Clusters “The Future Ocean”
(Kiel) and “The Ocean in the Earth System” (Bremen/Bremerhaven) which host a variety of
projects with relevance to ocean change, and will contribute to the cooperation between these and
other national marine institutions.
35
36
37
10.
38
Summary Budget
Koordination
Themen
1.3
(Birgit Schneider)
2.3
2.2
(Catriona Clemmesen)
Ausw irkungen auf TopPrädatoren: Fische und
Cephalopoden
(Felix Mark)
Abbau und Umsetzung v on
organischen Substanzen
(Anja Engel)
Physiologische
Langzeiteffekte auf
Lebensstadien
benthischer Krebse
Modellierung biogeochemischer
Rückkopplungsmechanismen
der organischen
Kohlenstoffpumpe
1.2
(Thomas Brey )
(Jelle Bijma)
Ultrastruktur und
elementarer Aufbau v on
Kalkskeletten
(Foraminiferen, Korallen)
(Ralph Tollrian)
3.5
(Adrian Immenhauser)
Der Einfluss rezenter und
fossiler Ozean Versauerung
auf den Stoffw echsel, die
Biomineralisation und die
Biodiv ersität pelagischer und
neritischer Kalkschaler
(Michael Böttcher)
Durch Mikroumgebungen
kontrollierte (De)Kalzifizierungsmechanismen
3.4
3.3
3.2
Kalzifizierung unter pHStress: Wirkungen auf
organismischer und
ökosystemarer Ebene
(Frank Melzner)
Zelluläre Mechanismen
der Kalzifizierung
3.1
Ausw irkungen auf
2.1 Weidegänger und Filtrierer
(Dirk de Beer)
(Hans Pörtner)
(Thorsten Reusch)
Anpassung in
1.1 autotrophen
Organismen
Akklimatisierung v ersus
(Maren Voß)
4.1
4.2
(Maarten Boersma)
Effekte der
Ozeanv ersauerung auf
Nahrungsnetze und
kompetitiv e Interaktionen in
pelagischen Ökosystemen
(Martin Wahl)
Effekte der Ozeanv ersauerung auf
artspezifische Performance
sow ie Konkurrenz und
Struktur in mikrobiellen und
makrobiellen
Gemeinschaften
(Maarten Boersma)
5.1
(Andreas Oschlies)
(Martin Quaas)
Bew ertung unsicherer
ökologisch-ökonomischer
Folgen der Ozeanv ersauerung mit der
Viabilitäts-Methode
5.3
5.2
Ev aluierung und
Parameteroptimierung der
pelagischen
Kalziumkarbonat
Produktion in globalen
biogeochemischen
Modellen
(Johannes Pätsch)
Einfluss der
Alkalinitätsflüsse vom
Wattenmeer auf den
Kohlenstoffkreislauf und
die Primärproduktion der
Nordsee
(Andreas Oschlies)
5
Integrierte
Abschätzung:
Sensitivitäten und
Unsicherheiten
4
2
Leistungsmerkmale bei
Tieren: Reproduktion,
Wachstum und
Verhaltensweisen
1
Primärproduktion,
mikrobielle Umsätze
und biogeochemische
Rückkopplungsmechanismen
Interaktionen zwischen
Arten und die
Zusammensetzung der
Gemeinschaften in
einem sich ändernden
Ozean
3
0.4
Kalzifizierung:
Empfindlichkeiten
von Phyla bis zu
Ökosystemen
Infrastruktur-Ent wicklung (Hans Pörtner)
Training und Wissenstransfer (Michael Meyerhöfer)
0.3
0
Projektkoordination (Ulf Riebesell)
Daten-Management (Michael Diepenbroek)
0.2
11.
Projekte
0.1
Detailed descriptions of Themes and Projects
Fig. 0.1: Consortium structure: overarching activities, themes and projects. Responsible PIs in
parentheses.
39
Table 0.1: Links between projects. Coloured boxes indicate projects with direct exchange (e.g. joint experiments, joint
0.3.1
0.3.2
0.4
1.1.1
1.1.2
1.1.3
1.1.4
1.1.5
1.2.1
1.2.2
1.2.3
1.2.4
1.2.5
1.3
2.1.1
2.1.2
2.1.3
2.2.1
2.2.2
2.3.1
2.3.2
3.1.1
3.1.2
3.1.3
3.1.4
3.2.1
3.2.2
3.2.3
3.2.4
3.3.1
3.3.2
3.4.1
3.4.2
3.4.3
3.5.1
3.5.2
3.5.3
4.1.1
4.1.2
4.1.3
4.1.4
4.2.1
4.2.2
5.1
5.2
5.3
40
5.3
5.2
5.1
4.2.2
4.2.1
4.1.4
4.1.3
4.1.2
4.1.1
3.5.3
3.5.2
3.5.1
3.4.3
3.4.2
3.4.1
3.3.2
3.3.1
3.2.4
3.2.3
3.2.2
3.2.1
3.1.4
3.1.3
3.1.2
3.1.1
2.3.2
2.3.1
2.2.2
2.2.1
2.1.3
2.1.2
1.3
2.1.1
1.2.5
1.2.4
1.2.3
1.2.2
1.2.1
1.1.5
1.1.4
1.1.3
1.1.2
0.4
1.1.1
0.3.2
0.3.1
use of equipment and measurement capacity, exchange of samples, etc.)
Theme O: Overarching activities
Project 0.1: Project coordination
Coordinator: Ulf Riebesell
Project Coordination
BIOACID will be coordinated at the Leibniz-Institute of Marine Sciences (IFM-GEOMAR) in
Kiel by Prof. Ulf Riebesell. The coordinator will be responsible for the day-to-day management
of BIOACID. He will ensure that the project runs in compliance with the requirements of the
contract (administrative, operational and scientific aspects). The Coordinator Project Office will
be organized at IFM-GEOMAR. Prof. Hans-Otto Pörtner at the Alfred-Wegener-Institute for
Polar und Marine Sciences in Bremerhaven will act as Deputy Coordinator. He will fill in for the
Coordinator position under circumstances when the Coordinator is unavailable.
To uphold an efficient project management, the project coordinator will employ a full time
project manager (PM) and a half time secretary. The assistance of financial officers and
personnel at central administrative departments will be provided at no cost.
The PM will regularly report to the Coordinator. His scientific and administrative tasks will be to:
• assist the Coordinator as the overall contact person on scientific matters and reporting,
and serve as the intermediary for communication between the partners on administrative
and financial matters
• organise meetings and workshops, as well as any action for training and dissemination
purposes
• prepare agendas for all meetings
• communicate all decisions regarding the implementation of the scientific work to the
theme leaders and project PIs
• work in close collaboration with the theme leaders and project PIs, collecting all formal
research and training plans and progress reports
• coordinate and run the day-to-day administrative and financial tasks
• control that financial contributions are transferred to contractors on time
• assist the project communication.
Budget and Budget Justification
First Year
Second Year
Third Year
Total
Personnel costs
Project manager E13-5
Secretary E6 1/2
Student helpers (8 x 75hs)
41
First Year
Second Year
Third Year
Total
Overtime technicians
Dissemination: Camera
personnel for TV –
production
Subtotal
Consumables
Printing costs management
Meetings, workshops,
catering
Consumables office
Dissemination: printing
costs (poster, flyer etc.)
Dissemination: material
TV production
Subtotal
Travel
Coordinator
Project manager
Dissemination: camera
personnel
Subtotal
Investments
Computer project manager
Computer secretary
Subtotal
Other costs
Dissemination: cutting TV
production
Subtotal
TOTAL
Budget justification
Personnel: The scientist position is for a full time project manager (PM). This position will be
offered to Dr. Michael Meyerhöfer, who has assisted during the development of the BIOACID
programme and has contributed immensely to the preparation of the BIOACID pre- and full
proposals. The PM will handle logistical organisation and workshop coordination as well as
reporting tasks for the project and dissemination activities. Due to the administrative complexity
of BIOACID the PM will be supported by a half time secretary and student helpers.
One important dissemination activity of BIOACID will be the production of a TV-quality video
for distribution to different media, decision makers and other stakeholders interested in ocean
acidification. This includes the engagement of professional film and production personnel.
42
Consumables: A yearly budget is requested to cover costs of printing, copying and publication
charges for BIOACID relevant material for administrative (e.g. reports) and dissemination
purposes. Further costs arise for the organisation of the BIOACID annual meetings, preparatory
meetings for joint research and dissemination activities, meetings of BIOACID’s Executive
Board individually and jointly with the coordination team of the UK-NERC programme on ocean
acidification, as well as meetings with governmental and NGO stakeholders. All meetings of the
project will be held at locations where meeting rooms are available at low or no charge so that
costs can be kept at a minimum level.
Travel: To maintain close contact to partners within the project and with international
collaborators and to guarantee an effective exchange of information, travel funds are requested
for the coordinator and the PM.
Investment: A desktop computer including licenses for standard software is requested for the
project manager and the secretary.
43
Project 0.2
Data management
PI: Michael Diepenbroek, WDC-MARE / PANGAEA, University of Bremen
i.
Objectives:
•
To coordinate data capture within BIOACID themes 1-5 and promote dataflow between
these themes and their working groups.
•
To store, manage and distribute output from the modelling exercises.
•
Perform technical quality control on all data and check the validity and completeness of
metadata.
•
Provide open access to all BIOACID data and information in a timely and efficient
manner via interfaces that are scalable for different partners and other stakeholders.
•
To ensure the long-term archiving, publication, and distribution of data according to
international standards and protocols.
•
To harmonize data management and facilitate data exchange between BIOACID and
related projects, such as EPOCA and SOPRAN, which both use PANGAEA for data
management
The sharing of data and information among partners of large coordinated projects, such as
BIOACID, strengthens the collaboration between different disciplines and research groups and
ultimately leads to an increased scientific output through synergistic effects. Experience from
previous projects has shown that both individual scientists as well as the project as a whole
benefit from data sharing and collaboration. The benefit of BIOACID to the wider scientific
community also requires data generated within the project to be freely available and easily
accessible. To encourage and promote data and information exchange in a timely and efficient
manner, BIOACID will establish a data policy with binding rules for all project partners. While
calling for openness and free flow of data between partners, these rules must also protect the
intellectual property rights of the data producers.
A major requirement of the data policy is that all data must be lodged in the BIOACID database
within 3 months after the time of measurement (data provided at zero cost to the project may by
lodged in other databases as long as they are publicly-accessible). Access restrictions may be set
up during the proprietary rights period (2 years after measurement) on request from the data
originator. During the proprietary rights period, data will only be passed to project partners with
the data originators agreement. BIOACID will encourage all PIs to share data during the
proprietary rights period and should ensure that the data originators contribution is respected
appropriately through co-authorship or acknowledgement.
ii.
Background and Previous Work
The proponents of this subproject have a broad and substantiated background in the fields of data
management and related IT subjects (databases, data infrastructures, protocols, system design,
and automatization). During the past 15 years the group has built up an information system
(PANGAEA – Publishing Network for Geoscientific & Environmental Data – www.pangaea.de),
44
which is a system for acquisition, processing, long-term storage, and publication of georeferenced data related to all earth science fields. In 2001 the PANGAEA group founded the
ICSU World Data Center for Marine Environmental Sciences (WDC-MARE – www.wdcmare.org), which uses PANGAEA as operating platform. The organization of data management
includes quality check and publication of data and the dissemination of metadata according to
international standards.
The challenge of managing the heterogeneous and dynamic data of environmental and
geosciences was met in the PANGAEA system through a flexible data model which reflects the
information processing steps in the earth science fields and can handle any related analytical data.
The basic technical structure corresponds to a three-tiered client/server architecture with a
number of clients and middleware components controlling the information flow and quality. On
the server side a relational database management system (RDBMS) is used for information
storage. To ensure fast data access the data are mirrored in a data warehouse which is also used
as interface to the German GRID community. RDBMS and warehouse currently hold about
450.000 fully documented data sets, comprising more than 2 billion analytical values
(observational data). Each data set can be referenced by a DOI (Digital Object Identifier). All
interfaces to the information system are based on web services including a simple map supported
(WFS, UMN) search engine (PangaVista).
For a number of international projects, PANGAEA is providing data portals for community
specific data access to a heterogenic data center infrastructure [Schindler U et al., 2006].
Currently portals for CARBOOCEAN and EUR-OCEANS are running.
The PANGAEA and the WDC-MARE are operating on a long-term basis. The institutional frame
is supplied by RCOM in cooperation with the Alfred Wegener Institute (AWI), Bremerhaven.
PANGAEA is a partner in over 50 national and international projects covering all fields of
environmental sciences (http://www.pangaea.de/Projects), in particular the EU Integrated Project
EPOCA, EU Coordinated Project EPOCA and the German BMBF Coordinated Project
SOPRAN, which all have close links with BIOACID.
References
Diepenbroek, M; Grobe, H; Reinke, M; Schindler, U; Schlitzer, R; Sieger, R & Wefer, G (2002): PANGAEA - an Information
System for Environmental Sciences. Computer & Geosciences, 28, 1201-1210, doi:10.1016/S0098-3004(02)00039-0
International DOI Foundation (2003): DOI Handbook. doi:10.1000/182
Schindler, U; Brase, J; Diepenbroek, M (2005): Webservices Infrastructure for the Registration of Scientific Primary Data. In:
Rauber, A et al. (eds.): ECDL 2005, LNCS 3652, pp. 128-138, 2005, doi:10.1007/11551362_12
Schindler, U; Diepenbroek, M (2006): Generic Toolbox for Metadata Portals. In prep.
iii.
Detailed Description of the Work Plan
The BIOACID DIS will be developed and implemented as early as possible so that data and
information flow is initiated in parallel with the first research activities. The purpose of the
BIOACID data management plan is to create a semi-distributed, scalable, and flexible
international data system into which research, observation, and modelling activities can submit
data and which will provide services that will increase the value of the data. Data capture and
data flow will be managed by an experienced data manager employed half-time and located at
WDC-MARE at the University of Bremen. The data manager will be responsible for:
•
Maintenance of the BIOACID web-page
45
•
Planning of data reporting procedures with the PIs prior to data collection
•
Keeping track of project data and the associated metadata
•
Web-based data tracking, including provision of common access to data by project PIs
•
Archival of data at the end of the project
According to the rules of good scientific practice (ESF 2001) all data sets will be archived longterm, and published using the partners’ data centres, in accordance with the wishes of the
responsible PIs. The data manager will watch that intellectual property rights are obeyed.
To provide access to the project-wide information BIOACID will be embedded in a common data
and information infrastructure implemented and maintained by the data manager and technical
staff from WDC-MARE. Contents and services of the website include:
•
General project information (documents, workshops, cruises, news etc.)
•
Data portal
•
Inventory of site and sampling locations (dynamic map)
•
Bibliography of BIOACID specific publications
•
“Yellow pages” for BIOACID related scientists
The data retrieval and access system will be based on distributed metadata catalogues located at
the cooperating data centres. Catalogue contents and access protocols are based on international
standards and protocols for geospatial data and metadata (e.g. ISO19115, DIF, DC, OGC-WFS,
OAI-PMH). Use and maintenance of these standards will ensure a high level of data consistency
and quality and will also facilitate later incorporation or union with information systems from
other communities. User interfaces will range from a map-based full text search engine to
hierarchical access through dynamic web pages. The implementation of the data infrastructure
will be carried out with regard to international SDI initiatives, in particular EU initiative
INSPIRE and GSDI.
Dissemination will be a continuous process throughout the project. All dissemination actions
and products will be negotiated with the BIOACID Management Team. Dissemination of project
results and deliverables will be carried out to the scientific community, political decision makers,
and the general public. Data management activities will be evaluated and critically reviewed
during workshops and the annual BIOACID meetings.
iv.
Specific Tasks and Deliverables
1. Coordination of data capture, integration and quality control activities for the five BIOACID
Themes.
As outlined in the Overview paper, original data sets have to be compiled from a number of
different sources, including data from BIOACID related cruises, experiments and from existing
data centres and working databases. Furthermore, the quality and state of these data sets might
46
vary enormously, as data are stored in a variety of formats with varying levels of metadata.
Therefore, in the initial phase an implementation plan for data and information management will
be set up. This plan will outline a structure and schedule for overall data flow between working
groups, quality management schemes, and data processing schemes. In addition it will regulate
the property rights of the data originators and the appropriate time intervals for public release
from creation of the data, as well as confidentiality issues.
An initial metadata catalogue of available data sets will be drawn up and used to define a priority
list for data acquisition and compilation. The catalogue will be continuously updated to enable all
BIOACID participants to locate and download target data sets. The metadata catalogue will be
complemented by a BIOACID specific bibliography.
2. Archiving and publishing data sets and data collections online and as offline products (DVD)
using persistent identifiers as DOIs.
Data sets will be described consistently according to the common metadata schema used. Data
still subject to access restrictions will be password secured and for internal use only. Published
data sets will be registered and made citable through usage of Digital Objects Identifiers (DOI) as
persistent identifiers (http://www.std-doi.de/). By way of the DOI registry data sets can also be
retrieved from library catalogues.
3. Implementation of the BIOACID data infrastructure
The data infrastructure (SDI) developed in this task has two objectives:
•
Enabling a distributed storage of observational and model simulation data within a
common networked structure, and
•
Establishing a robust and long lasting data network which can be extended by or
integrated into ongoing projects and programs.
The first is directly necessary for BIOACID to handle the potentially large mass of data from
modelling as well as the highly heterogeneous observational data. The second objective
corresponds to the worldwide efforts (as with INSPIRE) in developing a global spatial data
infrastructure (GSDI). A first setup will be based on the Open Archives Initiative protocol (OAIPMH) with Dublin Core as content standard. OAI-PMH is a widely used “de facto” standard,
supported by ad hoc usable open source software packages. One potential difficulty arises from
the currently rapid pace of SDI developments. Therefore, from the start of the project, all
implementations will follow a generic and flexible design allowing for later modifications and
extensions.
4. Maintain the website and data portal for BIOACID
The existing BIOACID website will be converted to WDC-MARE standards. The website will
include the data portal which enables efficient access to the individual and integrated data sets
(‘one stop shopping’). Key components will be a BIOACID specific search engine allowing for
retrieving and downloading of data sets relevant to the project. The search engine will form the
47
front-end of the common metadata catalogue. For best performance a harvesting approach will be
chosen for assembly and maintenance of the catalogue. The index for the search engine will be
complemented by a thesaurus. In order to minimise any friction losses during requests and
delivery of data sets a first version of a general SOPRAN data portal will be set-up in the first
phase of the project.
v. Budget and Budget Justification
First Year
Second Year
Third Year
Total
Personnel costs
Data manager E13 1/2
Subtotal
Consumables
Office consumables
Subtotal
Travel
Domestic travel
Subtotal
Investments
Computer
Supplies
Subtotal
Other costs
Subtotal
TOTAL
0.1
Personnel costs: A half-time data manager position will be established by WDC-MARE at the
University of Bremen. The data manager will be integrated in the staff of WDC-MARE while at
the same time keeping close contact with the coordinator and the project manager at all times
during the project.
Consumables: This will cover costs related to day-to-day data managing, including copying
charges for management relevant material (e.g. reports) and dissemination purposes.
Travel: To maintain close contact to the different partners within the project, to participate in the
BIOACID annual meetings and meetings relevant to data management and to guarantee an
48
effective exchange of information, travel funds are requested for the data domestic travel oif the
data manager.
Investment: Funding is requested for a workstation for the data manager and related supplies.
Project 0.3
Infrastructure development
PI: Hans O. Pörtner
i. Objectives
This project intends to support and develop the aquarium facilities and research infrastructure
needed for long term incubations (0.3.1.) and analyses of CO2 partial pressures in body fluids of
marine animals and ambient waters (0.3.2.). Technique development described under this project
is relevant for many projects of the network relying on animal maintenance and experimentation.
Firstly, the use of maintenance and culture facilities in recirculated aquaria like the one at AWI
Bremerhaven brings with it the inherent shortcoming that water physicochemistry is fluctuating
over time (largely due to net proton equivalent ion exchange of the organisms) and needs to be
monitored and stabilized on long time scales. Such shortcomings are not experienced by marine
institutes at rocky shores where flow through systems can be used for maintenance of water
physicochemistry (However, such flow through systems will strongly be influenced by
environmental variability). Such aspects also have to be considered for experimental mesocosms
and larval cultures under recirculated versus flow through conditions. The techniques proposed in
subproject 0.3.1. are intended to close this gap and should support establishment of stable
baseline conditions in recirculated systems. The solutions to be developed are also suitable to
establish baseline conditions in larger experimental systems at pre-industrial levels and various
levels of CO2 induced ocean acidification and thus appear as a relevant contribution which
widens the scope of experimental research in the field of ocean acidification and may also
become relevant in aquaculture systems.
Secondly, the online monitoring of carbon dioxide as outlined in subproject 0.3.2. can support the
elaboration of new knowledge of the effects of increased carbon dioxide content on marine
organisms and environment. The aim of this subproject is the development of new chemical
optical sensors and instrumentation for the determination of carbon dioxide partial pressure
(pCO2) in fluids of marine organisms and in marine environments based on the dual lifetime
referencing (DLR) method patented by PreSens. Emphasis will be put on the development of
pCO2 sensors with appropriate dynamic range in order to meet the conditions present in marine
systems and on the development of suitable instrumentation. The performance of the new
equipment will be optimised according to the experimental applications provided by project
partners with the prototype pCO2 sensors and instruments.
ii. State of the Art
Aquarium maintenance and mesocosms
Stability of sea water physicochemical parameters needs to be secured in animal maintenance
systems. This is a matter of significant concern in all laboratories which do not have access to
49
open ocean flow through seawater and rely on recirculated aquaria with a water treatment
section. This includes the large recirculated aquarium system at AWI Bremerhaven, funded by
BMBF upon the unification of AWI and BAH.
Oxidative catabolism of heterotrophs is characterized by net proton production (Pörtner 1995)
which is compensated for by ion exchange mechanisms and leads to a net release of protons into
the water with the result of disturbed physicochemical parameters at constant or falling DIC (the
latter occurs once excess CO2 generated by titration of (bi)carbonate is released into the
atmosphere). Furthermore, any delay in the equilibration of air or gas mixtures with the aquarium
water will prevent precise analysis of relevant water physicochemistry and thus sabotage any
efforts to maintain constant settings of relevant parameters. The goal of this development must
therefore be to monitor the effects of non-respiratory proton loads to the system water under
equilibrium conditions and to compensate for any pH disturbance resulting from net proton
equivalent ion exchange by aquarium maintained organisms.
Rearing of larvae of crustaceans, cephalopods and fishes (esp. cod) also needs to be carried out in
recirculated aquarium systems under largely undisturbed conditions. These activities will make
the putative most sensitive life stages available for studies of effects of ocean acidification.
Rearing would ideally occur under control vs. elevated CO2 conditions. The proposed system is a
miniaturized AquaInno Pond-in-Pond system for hatchery developed by AWI and IMARE (R.
Fisch, B.H. Buck). It offers a “system within a system” approach by providing process water with
additional purification and water exchange on demand, without disturbing sensitive life stages.
Modification and change of modules is possible and support a wide range of experiments. The
system can be isolated from air and ambient water for equilibration with various CO2 levels.
During long term incubations repeated water exchange is feasible in the pH stabilized
recirculated large scale aquarium systems (see above).
Fig. 0.2 (1): Draft of the floating AquaInno nearshore technology illustrating its components for water treatment. The
waste water flows from the fish sector (A) to a first waste collector (B), where particles will sediment in the tubes
system. A following bioreactor (C) eliminates dissolved nitrogen and phosphate. The re-use of water is controlled by
sensors (G) and sediment disposal (F) takes place. A pneumatic system simplifies handling and harvesting (E) of
cultured organisms. The platform (D) around the culture module is improving the overall handling of the system.
Surrounding algal cultures (I) eliminate the final product nitrate. D and I are not included for indoor use.
Fig. 0.3 (2): First freshwater prototype of a floating in pond raceway used for cultivation of ornamental fish in the year
2004. Currently, the system is developed for hatchery and grow-out in seawater and has been modified as a
recirculation system with different modules.
For CO2 equilibration according to IPCC emission scenarios large temperature controlled
experimental chambers have to be maintained at constant CO2 partial pressures according to gas
mixtures provided by commercial suppliers. A prototype developed by AWI (B. Klein) needs to
50
be further modified to reduce gas quantities used and improve stability of physicochemical
parameters.
Fig. 0.4: Prototype of larger scale CO2 incubation system developed at AWI (B.Klein) (total volume 2.5 m3, aerated
and filtered, temperature controlled).
CO2 sensor development
Standard methods for the determination of gaseous carbon dioxide include the direct detection
via infrared spectroscopy (Werle et al. 1998). Most sensors for the determination of dissolved
CO2 are modified pH sensors exploiting the acidic nature of carbon dioxide in contact with
aqueous media. In 1958 Severinghaus and Bradley introduced an electrochemical pCO2 sensor
comprising a pH glass electrode immersed into a small volume of a weak bicarbonate buffer
solution which is separated from the sample by a gas-permeable but ion-impermeable membrane,
such as Teflon. This type of sensors has found widespread application in industrial (e.g.
biotechnology or beverage) analyses (www.mt.com).
Various efforts were made to develop optical pCO2 sensors since Lübbers and Opitz (1975)
reported an optical pCO2 sensor based on the pCO2-dependent fluorescence intensity change of 4methylumbelliferone in a PTFE-covered bicarbonate buffer. These efforts resulted in optical
pCO2 sensors for biotechnology applications (www.ysilifesciences.com, www.fluorometrix.com)
based on fluorescence measurements. DeGrandpre et al. (1995) developed a submersible
autonomous moored instrument for CO2 which is based on a colorimetric pH measurement and
commercialised by Sunburst Sensors, LLC (www.sunburstsensors.com). Yet, this system is bulky
and not suitable for laboratory measurements where only small amounts of sample volume (e.g.
body fluids of marine organisms) are available.
51
Carbon dioxide dissolves in aqueous solutions both physically and chemically. Carbonic acid
thereby produced causes a pH drop which can be visualised with pH-sensitive indicators. Thus
principally all chemical optical carbon dioxide sensors are based on the detection of the pH
change in a CO2-sensitive matrix - whether they are based on colorimetric or luminescence
detection methods. Luminescence detection-based methods are superior to colorimetric
approaches due to their higher sensitivity. Here, the luminescence lifetime detection technique
offers an advantage compared to intensity-based methods because intensity fluctuations - caused
by e.g., fluctuations of excitation light source or background light - can be easily referenced. The
decay time is insensitive to such fluctuation whereas the luminescence intensity is affected. The
principle of luminescence quenching by carbon dioxide is illustrated in Fig. 0.5.
Excitation
Emission
Fig. 0.5: Principle of luminescence quenching by
carbon dioxide. The deprotonated form of the pH-
I-
sensitive indicator I- displays fluorescence when excited
I-
at a distinct wavelength. In the presence of carbon
dioxide a portion of the pH-sensitive dye in the sensor is
protonated. The emission intensity and thus the average
CO2
I
CO2
-
HI
HI
Protonation
lifetime of the indicator system decrease at increasing
carbon dioxide concentration since the protonated form
CO2
No Emission
cannot be excited by the excitation light used.
The measurement principle of chemical optical pCO2 sensors to be developed in this project is
based on the determination of the luminescence decay time of pH-sensitive dyes incorporated in a
gas-permeable – but ion-impermeable – membrane. With the DLR method patented by PreSens it
is possible to internally reference the luminescence decay time detected and to transfer it into the
µs time regime. This enables the use of easier and economically affordable instrumentation based
on the phase modulation technique.
Together with the luminescent dye sensitive to carbon dioxide an inert luminescent dye is
incorporated into the sensing membrane. Both dyes are excited by sinus-modulated light. Thus,
the luminescent response – a superposition of the CO2-sensitive dye and the inert dye – is also
modulated, but with a phase shift proportional to the mean decay time of the overall system,
which is again a function of the carbon dioxide concentration. The principle of the luminescence
decay time detection by phase modulation technique is visualised in Fig. 0.6. This technique is
used for all devices from PreSens. It proved to be a very reliable and robust method.
52
Fig. 0.6: Principle of luminescence
decay time detection. At low pCO2 the
Φ1
Φ2
Excitation
high CO2
low CO2
contribution of the CO2-sensitive dye
(short decay time) to the total signal is
high resulting in a low phase shift (Φ1)
Signal
compared to excitation. With increasing
pCO2 the weight of the reference dye
(long decay time) increases resulting in
an increase of phase shift (Φ2).
time / µs
iii. Previous Work of the Proponents
0.3.1
Franz Josef Sartoris has a strong background in the physiology of marine animals, including
acid-base regulation, ion- and osmoregulation, energy metabolism and respiration (Sartoris and
Pörtner, 1997; Pörtner and Sartoris, 1999; Sartoris et al., 2003; Metzger et al., 2007). He has a
broad experience with the implementation of physiological and biochemical methods in smallsized animals like amphipods, copepods and decapod larvae. He is the Scientific Manager of the
state of the art recirculating seawater aquarium at the AWI with a capacity of 150 m3 for the
rearing of temperate and polar animals.
Bela Hieronymus Buck is a marine biologist experienced in various aspects concerning the
cultivation of marine organisms (e.g. Buck et al., 2002, 2005 a,b). Research topics addressed
were the physiology and symbiotic interaction of giant clams, parasite infestation of bivalve,
cultivation strategies of various species including fish, and the development of new system
design featuring its realization by accepting socio-economic aspects and co-management. He also
works on the modification of modern aquaculture recirculation systems (RAS) for fish
cultivation. Bela H. Buck is the head of the unit “Marine Aquaculture, Maritime Technologies
and ICZM” in AWI, and of the unit “Marine Aquaculture for responsible fisheries” of IMARE
(Institute for Marine Resources)..
Ralf Fisch is a biologist experienced in bionics and aquaculture (Fisch and Buck 2006, 2008).
During his early work he was studying the biological surfaces of arthropods. To explore the
multifunctional nano-structures he used oSEM, AFM and TEM technologies. Today he is
working in aquaculture research topics including the modelling of nutrient budgets and ecological
aspects. Also educated as an engineer, he is experienced in the construction and design of
aquaculture systems and co-inventor of a floating recirculation aquaculture system (FRAS). His
prime interest is the development and implementation of a sustainable aquaculture.
Guido Krieten is the technical manager of the seawater aquarium system at AWI. He is a
qualified mechanic and builder of central heating and ventilation systems. He has a degree in
maritime engineering. He is responsible for the technical aspects of aquarium maintenance, the
53
maintenance of seawater quality and the operation in compliance with safety regulations.
Hans O. Pörtner has a long standing history in studying acid-base regulation of marine
ectotherms and its metabolic impacts in relation to ambient water conditions (e.g. Pörtner et al.
1991, 1998). His experience in analyses of proton equivalent ion exchange will be an asset in the
monitoring and establishment of stable water physicochemistry. Current interest covers the use of
these systems in studies of interaction between CO2 levels and other climatic factors, the
mechanisms shaping cellular and whole-animal energy budgets under various thermal and
carbon-dioxide regimes, and the molecular mechanisms of environmental adaptation and
limitation. He has published more than 190 publications in peer-reviewed journals.
0.3.2.
PreSens has been developing and selling optical sensors since 1997 (Klimant, 1997, Apostolidis
et al, 2004, Schröder and Klimant, 2005). The product portfolio includes the production and
marketing of fiber-optical chemical sensors and scientific instrumentation for the determination
of oxygen, pH and temperature. Oxygen and pH sensors were successfully applied by AWI and
IFM-GEOMAR in various experimental setups, among many other scientific and industrial
customers. pCO2 sensors and instrumentation based on the dual lifetime referencing method
patented by PreSens are currently under development for biotechnology and medical
applications. It is reasonable that this concept can be transferred also to the development of pCO2
sensors with dynamic range in the trace concentration range (380 ppm – 1900, in body fluids 380
– 3000 ppm). The scientific team has gained expertise on the development of optical sensors for
more than 10 years.
Athanas Apostolidis is a research engineer at PreSens GmbH, Regensburg. He received his
diploma in chemistry on development of a multi-channel optical protein detector for continuous
annular chromatography. He received his Ph.D. funded by the BMBF in 2004 on a combinatorial
approach for development of optical gas sensors at the Institute of Analytical Chemistry, Chemoand Biosensors (Prof. Wolfbeis) at the University of Regensburg. During his Ph.D. he was
working on the development of optical carbon dioxide sensors. In 2005 he was working at
Mühlbauer AG, Roding, Germany, as process engineer and materials specialist in the
development of production processes for Smart Card and passport solutions. His present interest
is in the development of optical chemical sensors for carbon dioxide determination in biological,
biotechnology and medical applications
Christian Huber is a research engineer at PreSens GmbH, Regensburg. He received his Ph.D. in
2000 on design and characterization of novel anion-selective optical sensors at the Institute of
Analytical Chemistry, Chemo- and Biosensors (Prof. Wolfbeis) at the University of Regensburg.
During his Ph.D. he was working on the development on anion-selective optical chemical sensors
based on the DLR method. At PreSens he is a product specialist for application of chemical
optical oxygen sensors in food and beverage, pharmaceutical, biotechnology and medical
industry.
54
iv. Work Programme, Schedules, and Milestones
Subproject 0.3.1.
Recirculated mesocosms and larval cultures (full strength sea water)
PI: Franz J. Sartoris, Bela H. Buck, Ralf Fisch, Guido Krieten, Hans O. Pörtner
Work Programme
a. Development of a large recirculated aquarium system into a stable system maintaining baseline
physicochemical parameters.
The adaptation of proton equivalent ion exchange monitoring systems according to Heisler
(1989, cf. Pörtner et al. 1991, 1998) will support monitoring of physicochemical parameters in
equilibrium. Furthermore, carbonate buffers need to be used for compensation of non-respiratory
proton loads to the water. Fine-tuning of pH should be possible through pH-stat controlled
addition of bicarbonate solutions. Developments thus include:
•
Instrumentation for the monitoring of pH and DIC content under well controlled
conditions of PCO2 and temperature.
•
Establishment of adequate calcite buffering in large scale animal maintenance systems
(simulating the CO2 dependent buffering action of ocean sediments).
•
Complementary pH-stat controlled addition of bicarbonate solutions.
b. Development of a rearing and incubation system for larvae of various groups
Available prototypes will firstly be tested for use with established cephalopod cultures (Sepia
officinalis, project 3.1, 2.3) and crustacean larvae (project 2.2.). It will be investigated whether
the rearing of fish larvae can be carried out under those conditions, thereby complementing or
deviating to some extent from established aquaculture procedures. One of us (B. Klein) has
carried out the rearing of cod larvae in aquaculture facilities in Tromso, Norway.
c. CO2 incubation system for mesocosms (pH stat)
Combining the methods established under a. with the large experimental CO2 incubation system
will support long term stability and provide the basis for pH stat control of the water through
controlled addition of high concentration bicarbonate solutions (available inhouse funding
20.000,- €).
Internal cooperation: Projects 2.1.1. / 2.1.2. / 2.2.1. / 2.2.2. / 2.3.1. / 2.3.2. etc.
55
Schedule
0.3.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
Establishment of pH bicarbonate
monitoring system
Establishment of stabilized aquarium
physicochemistry
Establishment of prototype for larval
culture
Successful rearing of cephalopod
larvae under various water
physicochemistry settings
Successful rearing of crustacean larvae
under various water physicochemistry
settings
Successful rearing of fish larvae under
various water physicochemistry
settings
Analysis of system quality, success
evaluation, data interpretation
Manuscript preparation, presentation of
results at conferences
Milestones
- Implementation of pH bicarbonate analysis system
month 6
- Establishment of stabilized aquarium physicochemistry
month 12
- Complementation of aquarium equipment
month 20
- Establishment of prototype for larval culture
month 12
- Successful rearing of cephalopod larvae under various water
- physicochemistry settings
month 18
- Successful rearing of crustacean larvae under various water
month 24
- Successful rearing of fish larvae under various water
56
month 30
- Evaluation of combined data sets, system properties and uncertainties
month 33
IV
Subproject 0.3.2.
Development of Chemical Optical Sensor Technology for the Determination of pCO2
Partial Pressure in Fluids of Marine Organisms and Marine Environment
PI: Athanas Apostolidis, Christian Huber
PreSens Precision Sensing GmbH, Josef-Engert-Str. 11, 93053 Regensburg
phone: +49 941 942 72 150 / 114
fax: +49 941 942 72 111
e-mail: [email protected] , [email protected]
Work Programme
WP 1: Sensor Chemistry Development
This work package is divided into two main tasks. First, materials feasible for the production of
pCO2 sensors for seawater application will be selected according to the environmental conditions
present for the application of a pCO2 sensor. This selection includes decision for feasible pH
indicator dyes, buffer systems and matrix polymers. Second, from these components sensor
membranes will be produced and tested with respect to their dynamic range and operational
performance in a calibration setup to verify best performing membrane compositions.
Duration:
12 months
Milestone M1: Sensor chemistry feasible for the determination of pCO2 in the concentration
range present in marine environment.
WP 2: Sensor Housing Development
The sensor membranes developed in WP 1 will be integrated into mini sensor housings (e.g. flow
through cell, dipping probe) that need to be designed based on the 2mm polymer optical fibers
(POF) used by PreSens. For applications that cannot be served with the mini sensor setup the
sensor chemistry will be adapted to micro sensor housings based on 140µm optical glass fibers.
The different sensor / housing will be subject to storage stability tests to evaluate storage
conditions required for retaining sensor performance during transport and storage of sensors at
the end user.
Duration:
18 months
Milestone M2.1:
Housing for mini pCO2 sensors and packaging of sensors for storage.
Milestone M2.2:
Housing for micro pCO2 sensors and packaging of sensors for storage.
WP 3: Instrument Development
Optical and electronic setup of the instrumentation will be developed to fit to sensor chemistry
and to provide reliable measurements based on sensing technology from PreSens. Instrument
housing will be developed for applications under laboratory and field conditions. Optical setups
will be developed to fit a) mini sensor setups and b) micro sensor setups.
Duration:
18 months
Milestone M3.1:
Laboratory demonstrator for pCO2 measurement in marine application
for use with mini pCO2 sensors.
Milestone M3.2:
Demonstrator for measurement of pCO2 with micro pCO2 sensors.
WP 4: Optimisation of Sensor Performance
57
According to the performance evaluated for the sensor chemistry developed in WP 1 and the
feedback of field testers the sensors will be improved with respect to their accuracy, sensitivity
and stability. The stability improvement will include both storage and operational stability.
Duration:
24 months
Milestone M4: pCO2 sensor with improved performance according to sensitivity and, both,
operational and storage stability.
WP 5: Optimisation of Device Performance
The laboratory demonstrators for pCO2 measurement with mini or micro sensors will be
improved with respect to environmental conditions of pCO2 determination in field. This can
include battery operated systems, PC-independent setup, etc.
Duration:
18 months
Milestone M5: Instrument for field application pCO2 measurement with optimised performance.
WP 6: Provision of Instruments, Sensors and Support to Project Partners
PreSens will provide some instruments and sensors developed to field testers to verify
performance of the calibration results in our labs to real experimental conditions. Presens will
implement the feedback of the testing partners to the sensor chemistry and the instrument
development and improvement.
Duration:
30 months
Result: Valuable feedback information for the validation of setup performance and the
improvement of the pCO2 measurement system.
Schedule
0.3.2
First Year
I
WP 1 Sensor Chemistry Development
WP 2 Sensor Housing Development
WP 3 Instrument Development
WP 4 Optimisation of Sensor
Performance
WP 5 Optimisation of Instrument
Performance
WP 6 Provision of Instruments, Sensors
and Support to Project Partners
58
II
III
Second Year
IV
I
II
III
IV
Third Year
I
II
III
IV
Milestones (see work packages above)
- Sensor chemistry feasible for determination of pCO2 in the concentration
range present in marine environment (M1)
- Housing for pCO2 mini sensors and packaging of sensors for storage
(M2.1)
- Housing for pCO2 micro sensors and packaging of sensors for storage
(M2.2)
- Laboratory demonstrator for pCO2 measurement in marine application for
use with pCO2 mini sensors (M3.1)
- Laboratory demonstrator for pCO2 measurement in marine application for
use with pCO2 micro sensors (M3.2)
- Instrument for field application pCO2 measurement with optimised
performance (M5)
- Instrument for field application pCO2 measurement with optimised
performance (M5)
month 12
month 12
month 21
month 15
month 24
month 33
month 36
Internal cooperation:
The sensors and devices that will be developed in this project will provide instrumentation
infrastructure for online monitoring of carbon dioxide partial pressure in organism body fluids
and marine environment for laboratory experiments of the project partners [link to 2.1.2, 2.1.3,
2.3.1, 2.3.2, 3.1.3, 3.1.4, 3.2.2, 3.4.2]. Their experience with prototype instrumentation and
sensors will be implemented in optimisation of performance.
Cooperation with other projects outside the Verbundproject
Collaborations beyond the project consortium that are relevant to the subproject project are not
planned. A request for funding for this subproject has not been submitted to any other addressee.
In case a request will be submitted, we will inform the Federal ministry of Education and
Research (BMBF) immediately.
59
First Year
Second Year
Third Year
Total
Personnel costs
0.3.1 Technician
0.3.2 Chemical Engineer, PhD *
7PM
0.3.2 Electronic Engineer, PhD *
3PM
0.3.2 Electronic Engineer, M.Sc *
3PM
0.3.2Technician *
6PM
Subtotal
Consumables
0.3.1.
0.3.2 Chemicals and Gases *
0.3.2 Polymer Optical Fibers *
0.3.2 Consumables Sensor Production *
0.3.2 Electronic and Optical
Components *
Subtotal
Travel
0.3.1
0.3.2 National *
0.3.2 International *
Subtotal
Investments
0.3.1
Subtotal
Other costs
0.3.1.
0.3.2 *
Subtotal
TOTAL
*:For subproject 0.3.2. numbers represent 50% of the total budget. The other 50% will be contributed by the company
(Presens) itself.
Budget justification 0.3.1
Personnel costs:
The success and feasibility of this project is dependent on the approval of the technical assistant,
who will be in permanent charge of the very time-consuming process of larval rearing (e.g.
60
crustaceans, cephalopods, fishes) and associated assessments of water quality. The success of the
project is dependent on a uniform manner of larval rearing to avoid errors and to minimize
variability in conditions or ways of handling larvae. This excludes the repeated hiring of
fluctuating personal, like students before or during their Masters periods. It is thus inevitable to
have technical support available for continuous larval rearing, to provide sufficient good quality
material for experimental work and to not distract scientific personal from the objectives of the
science projects.
After a period of vocational training and gaining daily routine, a full time position is needed to
establish the rearing of larvae of the various groups under both control and experimental
conditions.
Consumables:
Chemicals, Artemia eggs, algal and other (e.g. mysid) cultures for feeding of larvae
Travel:
The collection of animal specimens (ovigerous females) and eggs from various locations in
Northern Europe demands travel funds for the technician and other personnel to support
collection and safe arrival of fresh material at AWI. Exchange visits may also be needed with
laboratories experienced in the rearing of e.g. cod larvae.
Investments:
Control and maintenance of water physicochemistry
In accordance with the reasoning the aquarium systems mentioned above need complementation
by flow through tanks containing buffer material of various compositions. They are also
equipped by components of the pH bicarbonate system for online monitoring of pH under
equilibrium conditions as well as a computer controlled pumping system for the addition of
concentrated base.
Larval rearing system
In accordance with the reasoning above finalized development and construction of the prototype
will demand a lump sum of per 3 years.
CO2-Incubation mesocosm
In accordance with the reasoning above long term incubation at various CO2 tensions in larger
volume systems needs reproduction of the existing prototype, one for each CO2 tension. Together
with available inhouse funding an amount of is requested to install 4 further CO2 systems to
cover preindustrial, 2 x preindustrial, year 2100 and higher CO2 levels.
61
Budget and Budget Justification 0.3.2
Personnel:
Chemical Engineer, PhD: Responsible for planning, execution and the development of pCO2
sensors and measurement instruments;
Electronic Engineer, PhD: Development and production of measurement instruments for pCO2
sensors, especially circuit design and optical design;
Electronic Engineer, M.Sc.: Development and production of measurement instruments for pCO2
sensors, especially software development;
Technician: Execution of sensor characterisation with laboratory calibration setups
Consumables: Chemicals and gases are required for the production and the characterisation of
the pCO2 sensors. Polymer Optical Fibers will be coated with the pCO2 sensor materials to be
developed and will be supplied to project partners for evaluation.
Consumables Sensor Production: For the production of pCO2 sensors supplies are required that
are prone to deterioration and need regular replacement (e.g. membrane support materials,
pipetting tips or needles);
Electronic and Optical Components: Presens will build up to 5 demonstrators of pCO2
measurement instruments for marine applications and supply to project partners.
Travel: National: Travels to project partners and annual meetings; International: 1 international
congress p.a.
vi. References
Apostolidis A, Klimant I, Andrzejewski D, Wolfbeis OS (2004). Combinatorial Approach for Development of Materials for Optical Sensing of
Gases. J Comb Chem 6: 325 – 331
Buck BH, Buchholz CM (2005b) Response of offshore cultivated Laminaria saccharina to hydrodynamic forcing in the North Sea,
Aquaculture, 250: 674-691.
Buck BH, Rosenthal H, Saint-Paul U (2002) Effect of increased irradiance and thermal stress on the symbiosis of Symbiodinium
microadriaticum and Tridacna gigas, Aquatic Living Resources 15: 107-117.
Buck BH., Thieltges DW, Walter U, Nehls G, Rosenthal H (2005a) Inshore-offshore comparison of parasite infestation in Mytilus edulis:
Implications for open ocean aquaculture. J Appl Ichthyol 21: 107-113.
DeGrandpre, M.D., Hammar, T.R., Smith, S.P., and F.L. Sayles. (1995)In situ measurements of seawater pCO2, Limnol Oceanogr, 40: 969 –
975
Fisch R, Buck BH (2006) Neues Aquakultursystem für das Meer made in Germany, Fischerblatt 12: 13-16.
Fisch R, Buck BH (2008) Waste reduction in finfish aquaculture within suspended semi closed and closed systems. J Appl Ichtyol, in press.
Heisler N (1989) Parameters and methods in acid-base physiology. In Techniques in Comparative Respiratory Physiology: an experimental
approach (ed. C.R. Bridges and P.J. Butler). pp. 305-332, Society for experimental Biology Seminar Series. Cambridge, Cambridge
University Press.
Klimant I (1997) Verfahren und Vorrichtung zur Referenzierung von Fluoreszenzintensitätssignalen, Ger. Pat. Appl. DE 198.29.657
Lübbers DW, Opitz N (1975) The pCO2/pO2 optrode: A new probe for measuring pCO2 and pO2 of gases and liquids. Z Naturforschung 30C:
532-533.
Metzger R, Sartoris FJ, Langenbuch M, Pörtner HO (2007) Influence of elevated CO2 concentrations on thermal tolerance of the edible crab
Cancer pagurus. J Thermal Biol 32: 144-151
Pörtner HO, Sartoris FJ (1999) Invasive studies of intracellular acid-base parameters: quantitative analyses during environmental and
functional stress. In: regulation of Tissue pH in Plants and Animals, edited by EW Taylor, S Egington & JA Raven. S.E.B. Seminar
Series, Cambridge University Press. 68-98
Pörtner, HO (1995) pH homeostasis in terrestrial vertebrates: a comparison of traditional and new concepts. Adv Comp Env Physiol 22: 51-62.
Pörtner, HO, Reipschläger A, Heisler N(1998) Metabolism and acid-base regulation in Sipunculus nudus as a function of ambient carbon
dioxide. J exp Biol 201: 43-55.
Pörtner, HO, Andersen NA, Heisler N (1991) Proton equivalent ion transfer in Sipunculus nudus as a function of ambient oxygen tension:
relationships with energy metabolism. J exp Biol 156: 21-39.
Sartoris FJ, Bock C, Serendero I, Lannig G, Pörtner HO (2003) Temperature dependent changes in energy metabolism, intracellular pH and
blood oxygen tension in the Atlantic cod, Gadus morhua. J fish biol 62: 1239-1253
Sartoris FJ, Pörtner HO (1997) Temperature dependence of ionic and acid-base regulation in boreal and arctic Crangon crangon and Pandalus
borealis. J Exp Mar Biol Ecol 211: 69-83
62
Schroeder CR, Klimant I. (2005) The influence of the lipophilic base in solid state optical pCO2 sensors: a comparative study. Sens. Actuators
B 107: 572 – 579.
Severinghaus JW, Bradley AF (1998) Electrodes for blood pO2 and pCO2 determination. J Appl Physiol 13: 515 – 520.
Werle P, Mücke R, Amato F D, Lancia T (1998). Near-Infrared Trace-Gas Sensors based on Room-Temperature Diode Lasers, Appl. Phys. B
67: 307 – 315.
Project 0.4: Training and transfer of know-how
PI: Michael Meyerhöfer
Training and transfer of know how between the BIOACID participants will be carried out by the
following workshops, offered by the different institutions.
1.: Workshop on microsensors (Dirk de Beer, MPI Bremen)
Each of the two workshops should last ten days. The first half will be sensor making, the second
sensor use. Participants can bring there own samples for measurements.
2.: Workshop on isotope geochemistry and laser-ablationtechniques (Anton Eisenhauer,
IFM-GEOMAR, Kiel)
We intend to perform during four days a training workshop on the use of isotope geochemistry
and on its use for biomineralization studies. This workshop will also give an introduction to the
use of laser-ablation techniques as a new micro-analytical tool.
3.: Physiological approaches to body fluid physicochemistry and acid-base regulation
(Franz Sartoris, Christian Bock, Hans Pörtner and colleagues, AWI, Bremerhaven)
We intend to host a three day workshop on the concepts of quantitative acid-base physiology and
the use of various techniques associated with the quantification of changes in tissue and body
fluid acid-base and associated ion and metabolic status in animals. The workshop will enable
PhD students to interpret their findings in the light of changes in water physicochemistry and
their effect on acid-base, ionic and metabolic regulation.
4.: Workshop on seawater carbonate chemistry (Kai Schulz and Ulf Riebesell, IFMGEOMAR, Kiel)
All experimental projects proposed in BIOACID share a common aspect, i.e. the manipulation of
the seawater carbonate chemistry for different carbon dioxide (CO2) scenarios. Adjusting,
maintaining, and controlling the seawater carbonate system will be crucial for data interpretation
and quality control.
A joint two-day workshop on seawater carbonate chemistry will be held at IFM-GEOMAR at the
beginning of BIOACID to ensure high data quality and inter-comparability. The workshop will
cover the following topics:1) Principles of the seawater carbonate system: concepts, parameters
and speciation2) Calculations: techniques to calculate the carbonate system from measured
parameters 3) Measurements: dissolved inorganic carbon (DIC), total alkalinity (TA), pCO2 and
pH) Experimental manipulations: acid / base additions, CO2 aeration, DIC additions at constant
TA The workshop will enable all participants to choose the most appropriate CO2 manipulation
63
for their specific experimental approaches, determine the parameters necessary to calculate
carbonate chemistry speciation and evaluate the data obtained for quality.
5.: Workshop "Experimental design" (Martin Wahl, IFM-GEOMAR, Kiel)
We plan a five day workshop on setting up experiments in consideration of:
• Exact questions asked
• Carefully chosen response variables
• Temporal, spatial, methodological and biological scope of the investigation
• Sources of variance
• Desired data quality
• Available resources (time, funds, infrastructure)
• Synergetic combination with other investigation
• Suitable statistical approach
Schedule
First Year
I
Workshop on microsensors
Workshop on isotope geochemistry
and laser-ablationtechniques
Physiological approaches to body
fluid physicochemistry and acid-base
regulation
Workshop on seawater carbonate
chemistry
Workshop "Experimental design"
64
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
First Year
Second Year
Third Year
Total
Personnel costs
Subtotal
Consumables
Workshop on microsensors
Workshop on isotope
geochemistry and laserablation- techniques
Physiological approaches
to body fluid
physicochemistry and acidbase regulation
Workshop on seawater
carbonate chemistry
Workshop "Experimental
design"
Subtotal
Travel
Subtotal
Investments
Subtotal
Other costs
Subtotal
TOTAL
The costs for the course will largely be carried by BIOACID. The participants need to take care of accommodation, food, and travel.
The requested funds will only cover costs directly related to organizing and carrying out the
training workshops at the various partner institutions. Costs related to travel and accommodation
of the participants will be covered from travel funds of the individual sub-projects.
65
66
11.2: Theme 1: Primary Production, microbial processes and
biogeochemical feedbacks
i. Common Background
The partial pressure of CO2 (pCO2) in the atmosphere and the oceans has increased by 30% over
the past century and model scenarios predict a further increase to twice the current concentrations
by the end of the century, which is app. 700ppm by the year 2100 (Houghton et al. 2001). This
increase is far more rapid than any change over the last 400,000 years and expected to
significantly impact marine life (Fabry et al. 2008). Understanding the response of marine
autotrophs and of heterotrophic bacteria to rising CO2 concentrations and decreasing pH is of
special importance since these organisms provide the basis of marine food webs (link to theme
4). All aquatic plants/primary producers are adapted to pCO2 concentrations that varied only
between 200 and 300 ppm over the past 420 million years as ice core data suggested (Falkowski
et al. 2000), while they are facing novel pCO2 conditions that are several times higher on a very
rapid time scale within the next decades. To what extent species may currently be in the process
of adaptation to higher pCO2 levels by evolutionary adaptation is totally unresolved.
Fig. 1.1: Representatives of phytoplankton guilds that will be studied in theme 1 A Coccolithophore: Calcidiscus
leptoporus B diatom:Skeletonema costatum C. diatom:Thallsiosiara angulata D Cyanobacteria:Nodularia spumigena.
While increased pCO2 may enhance photosynthesis by alleviating pCO2 limitation, the extent and circumstances under
CO2 is limiting for photosynthesis and growth of primary producers in the field are still
unclear. Preliminary data reveal that species with effective carbon concentration mechanisms
(CCMs) are less sensitive to increased CO2 levels than those lacking efficient CCMs, analogous
to findings in terrestrial vegetation (Collins et al. 2006). Currently, our ignorance of the
metabolic diversity of oceanic autotrophy decreases the possibility of any realistic projection of
marine primary production in response to increased carbonation. Calcifying organisms like
coccolithophores and corals may suffer under decreasing calcification potentially leading to a
negative feedback to atmospheric CO2 (Gattuso et al. 1998, Wolf-Gladrow et al. 1999, Riebesell
et al., 2000, Orr et al 2005).
which
Species specific differences in inorganic carbon acquisition moreover indicate changing
competitive relationships between phytoplankton guilds (Fig. 1.1) at future CO2 concentration
(Rost et al., 2003). Experimental work on diazotrophic cyanobacteria suggests an increase in
productivity with increasing pCO2 (Hutchins et al., 2007; Ramos et al., 2007, Levitan et al.
2007). Recent data from seagrasses and several groups of micro- and macroalgae suggest an
enhancement of photosynthetic rates (Collins and Bell, 2004). Positively correlated CO2
concentrations and uptake rates potentially constitute an important negative feedback to
anthropogenic CO2 emissions, if enhancing the so-called biological carbon pump (Riebesell et al.
67
2007). A systematic assessment of autotrophic responses considering possible co-limiting factors
for photosynthesis such as nutrients, light and mixing depth, however, is still missing.
Evolutionary adaptation to enhanced pCO2 levels as response of species and populations
to environmental change is neglected in studies until now. In marine unicellular algae and
bacteria, in particular, rapid evolution to increasing pCO2 levels is expected due to short
generation times of <1 day. In the only experiment addressing the above hypothesis, populations
of the green algae Chlamydomonas rheinhardtii evolved high photosynthesis rates, higher
chlorophyll content and reduced cell size in comparison to untreated controls (Collins and Bell,
2004).
1,2
rel. TEP production
1,0
0,8
Present CO2
0,6
IPCC year 2100
0,4
0,2
Fig. 1.2: TEP increase under
high CO2 concentrations
(from Engel et al. 2002)
0,0
0
500
1000
1500
2000
2500
3000
pCO 2 (ppm)
Highly relevant to this proposal is the observed loss of the CCM in response to increased pCO2
which is in line with evolutionary theory, predicting that costly carbon concentration is no longer
selected for in a CO2 rich environment. Whether these results can be extrapolated to other species
is unknown. The observed genetic and physiological variation found among algal strains suggests
that there is ample genetic variation for rapid evolution to occur (Collins et al. 2006).
As a consequence of primary effects on photosynthesis and calcification biogeochemical fluxes
may become affected by increasing pCO2. One example is the sensitivity of carbohydrate
exudation and transparent exopolymer particle (TEP) production to changing CO2 concentration
(Engel, 2002); (Fig. 1.2). This may affect particle aggregation and export (Engel et al., 2004), and
hence represent a negative feedback to the atmospheric CO2 concentration. The release of
carbohydrates together with the excretion of other dissolved organic substances, will also serve as
a food source for heterotrophic bacteria and protozoa (Bronk et al., 2007).
The quantification and characterization of the production, exudation and microbial processing of
organic matter (OM) and how these processes respond to changes in seawater pCO2 and pH
requires further investigation to adequately assess the potential impact on future biogeochemical
cycling. Also, if ballasting substances like calcite decreases with decreasing pH, we may find
lower sedimentation rates as suggested by (Armstrong et al., 2002) resulting in a positive
feedback to atmospheric CO2 concentrations. Again, an improved understanding on how the
68
export of particulate matter from the upper water column is affected by high CO2 is essential to
improve the predictive capacity of global biogeochemical models.
An effect on the elemental stoichiometry of particles was found in large mesocosms where
increasing DIC was translated to carbon rich organic matter (Engel et al. 2005, Riebesell et al.,
2007). The combined influence of light, nutrients, and CO2 concentration, on elemental
stoichiometry and the relative importance of these factors individually is largely unknown
(Burkhardt and Riebesell, 1997, Burkhardt et al., 1999).
In the face of a not acceptable lack of information, the major objectives of theme 1 are to gain a
better understanding of the response of autotrophic communities and heterotrophic bacteria to
ocean acidification, and the related consequences for organic matter cycling, and the turnover of
key elements.
ii. Collaborative research
In theme 1 various aspects of responses to ocean acidification and carbonation are studied
including the export of primary production into the food-web and the formation of organic matter
by aggregation of particles. Currently, data on the responses of the diverse plankton groups to
Fig. 1.3: Set up with 5 CO2 controlled
chemostats in the lab of A. Engel at
the Alfred Wegener Institute in
Bremerhaven
where
joint
collaborations are planned within
project 1.2. (Photo: A. Engel)
ocean acidification are limited to a few species, a research gap that this collaborative effort is
intended to close through studies of key species and whole plankton communities in the field
(projects 1.2).
Broadening the taxonomic repertoire would also allow geographic predictions of sensitivities and
compositional shifts in major plankton groups to a global scale pCO2 increases. Several
subprojects under 1.1 address longer-lasting adaptations of phytoplankton and bacteria. Projects
under 1.2 focus on the interaction between autotrophs and heterotrophs and especially the role of
extracellular organic substances and aggregation. Finally project 1.3 models the impact of ocean
acidification on the soft tissue pump (POC + DOC) as a potential removal process of carbon from
the surface waters.
Additionally to the pCO2 induced changes the combined effect of the temperature increase will
be simulated under theme 1. Although most studies will be carried out in controlled labexperiments (e.g. chemostats from the AWI, Fig. 1.3) some studies will include open sea
plankton to test the response of communities on OA. Within the BIOACID Project we have
agreed upon several levels of CO2 which are: 280ppm (preindustrial) 380ppm (today), 560ppm
(twice preindustrial), 700ppm (2.5 preindustrial and the level of the IPCC "business as usual"
prediction for 2100), 980 – 1000ppm (3.5 preindustrial). Our experiments under Theme 1 will
69
follow these agreements and we plan joint experiments in several subgroups of Theme 1.
Furthermore it is planned to perform experiments by carbonation and not acidification only for
the pH reduction. This approach avoids conflicts from diverging culture setups. The effects of
changes in the temperature which are predicted for the same time span as the OA are tested for in
all the subprojects of theme 1.
Overall the results of theme 1 will supply a basis for modelling and future prognoses on the
ecological consequences of ocean acidification in biogeochemical model under theme 1.3. All
subprojects will feed their results and findings into a better assessment of acidification and global
warming on the so called “soft tissue pump”, especially the production and particle formation by
means of TEP. A more reliable estimate of the biogeochemical feedbacks of changing
productivity and export will be reached through a more realistic formulation of carbon production
and export in global marine biogeochemical models.
Project 1.1: Acclimation versus adaptation in autotrophs
T. Reusch (University Münster/ IFM-GEOMAR Kiel), M. Hippler/J. LaRoche (University
Münster/ IFM-GEOMAR Kiel), G. Jost/K. Jürgens (Leibniz Institute Warnemünde), M. Müller
(IFM-GEOMAR Kiel), U. Karsten/T. Hübener (University Rostock)
i. Objectives
The five subprojects within Project 1.1 address the effects of ocean acidification (OA) and
carbonation (rising seawater CO2 concentrations) on marine primary producers of diverse
taxonomic affiliation, including chemoautotrophic bacteria, benthic and planktonic microalgae.
Project 1.1 mainly studies effects of rising pCO2 on primary productivity and is mutually
complementary to project 3.1 (and here particularly 3.1.1) that focuses on calcifying primary
producers. The overall objectives are to:
• Identify guilds and taxonomic groups of autotrophic organisms that are sensitive to
acidification and carbon enrichment, compare treatments based on direct pCO2
enhancement versus changes in seawater carbon balance
• identify processes carried out by autotrophs that are affected by ocean acidification or
increased pCO2
• analyse short- and long-term physiological acclimation mechanisms that modulate
carbon acquisition in response to carbonation
• assess interactions between OA, nutrient status (in particular iron limitation) and ocean
warming for photosynthesis and photosystem proteomics of important phytoplankton
groups
• establish selection lines in order to assess evolutionary changes in physiology and carbon
acquisition under increased pCO2
• analyze genetic and proteomic changes as a response to long-term exposure under
increased pCO2
• quantify shifts in taxonomic composition among important autotrophic groups as a result
of acidification and carbonation, and possible second order effects on ecosystem
performance
Whereas effects of increased pCO2 in terrestrial systems have been intensely studied in the past
15 years (summarized in Ainsworth and Long 2005), work on marine primary producers is in its
70
infancy. Because planktonic and benthic microalgae form the basis of marine food chains (with
the exception of deep sea vent ecosystems), changes at the level of primary production may result
in cascading effects throughout the food web (link to Theme 4).
Increasing atmospheric pCO2 not only leads to drastic increases in seawater CO2 but also causes a
reduction in pH (i.e. acidification), and comparatively minor increases in HCO3- concentrations
(i.e. carbonation). Recent data suggest that the primary short-term response of micro- and
macroalgae is an enhancement of photosynthetic rates with rising pCO2. This may constitute an
important negative feedback to anthropogenic CO2 emissions, potentially enhancing the so-called
biological carbon pump (Riebesell et al. 2007). However, it is presently unclear to what extent
and under what conditions carbon availability limits photosynthesis and growth of primary
producers in the field. On the other hand, growth rates of calcifying phytoplankton species such
as coccolithophores may be impaired by rising pCO2 due to lower carbonate availability
(Riebesell et al. 2000) (link to theme 3), but recent data challenge this notion (Iglesias-Rodriguez
et al. 2008). On objective of this project will be to perform critical experiments that compare
different isolates with respect to their different (heritable) physiological performance. Moreover,
we will compare different pCO2 enrichment protocols to evaluate whether conflicting results may
arise from experimental procedures.
Because the sensitivity to carbon enrichment differs widely among taxa, rising CO2 levels will
alter competitive relationships and result in shifts of plankton species composition (Rost et al.
2003). Currently, data on the responses of the diverse plankton groups to ocean acidification is
limited to a few species, a research gap that this collaborative effort is intended to close.
Broadening the taxonomic repertoire is also a prerequisite for geographic predictions of
sensitivities and compositional shifts in major plankton groups to ocean acidification and
increasing pCO2. The project will study the response of several important photoautotrophic
groups such as non-calcifying planktonic diatoms and calcifying coccolithophores, benthic
diatoms to increases in pCO2 within a common analysis framework and largely congruent
experimental conditions.
In contrast to photoautotrophs, consequences of increased CO2 concentration on
chemoautotrophic organisms are probably quite different than on the surface phototrophic
primary production. Pelagic oxic-anoxic transition zones (redoxclines) found in several coastal
environments and marginal seas (e.g., Baltic Sea, Black Sea) are sites of significant
chemoautotrophic production (dark CO2 fixation) (Taylor et al., 2001; Jost et al. 2008). By
modifying the overall nutrient cycles, these redox-related transformations have an impact far
beyond the spatial scale of the redoxcline (best known for N losses). It is not known how changes
in CO2 concentration and pH affect the biogeochemistry of these areas. Because dark CO2
fixation is based on chemical energy from reactions including mainly pH-sensitive substances
(e.g. H2S, Mn, Fe), subproject 1.1.1 (Jost /Jürgens) hypothesizes significant changes in
biogeochemical process rates related to chemolithotrophy, microbial key organisms and
microbially based food webs. The thermodynamics and kinetics of some of the respiratory
processes, for instance Fe(III) and Mn(IV) reduction by microbes, are highly pH dependent
(Canfield et al. 2005). Changes in pH can induce changes in energy gain for microorganisms, in
the delicate balance of biogeochemical redox sensitive processes, and even the element fluxes of
the benthic-pelagic coupling in shallow euxinic systems. Subproject 1.1.1 will examine model
microorganisms that have been shown to constitute key players for distinct biogeochemical
transformations in pelagic redoxclines, and which had been successfully cultivated.
Ocean acidification accompanied with several concomitant changes in abiotic parameters that can
affect photosynthetic rates, one of which is the availability of dissolved iron. Despite its
71
abundance on earth, iron-deficiency is the most common nutritional deficiency in the world. Iron
deficiency affects at least 10% of oceanic photosynthetic productivity and is most prevalent in
vast high nutrient low chlorophyll (HNLC) regions of today’s oceans (Martin and Fitzwater
1988, Behrenfeld and Kolber 1999). As consequence, photosynthesis and carbon assimilation are
restricted by the low iron-bioavailability in these areas.
The decrease in pH will reduce the precipitation of dissolved iron as Fe oxides while also
reducing the binding of Fe to organic ligands, both factors being important in controlling Fe
bioavailability. The effects of ocean acidification on the bioavailability of iron and hence, on
phytoplankton productivity is currently not well understood. One hypothesis addressed by
subproject 1.1.2 (Hippler/LaRoche) is that OA minimizes the pressure of iron limitation on
primary productivity. Iron is essential for virtually all forms of life because it participates in
electron transfer reactions as a crucial cofactor in enzymes that catalyze redox reactions. Since
iron can also react with oxygen to generate cytotoxic agents, its accessibility within the cell has
to be under tight homeostatic control, which requires complex regulatory mechanisms (Finney
and O'Halloran 2003).
At the molecular level, photosystem I (PSI) is a prime target of iron-deficiency, probably because
of its high iron content. In the short-term, iron-deficiency leads not only to a pronounced
degradation of PSI, but also to a remodeling of the PSI associated light-harvesting antenna
(LHCI) (Moseley et al. 2002). Long-term adaptation to iron-deficiency even causes constitutive
differences in the photosynthetic architecture as demonstrated for coastal and oceanic diatoms
(Strzepek and Harrison 2004). The oceanic diatom strain contained up to fivefold lower PSI and
up to sevenfold lower cytochrome b6f complex concentrations as compared to a coastal diatom.
Surprisingly, the changes to the photosynthetic apparatus while decreasing the cellular iron
requirements of the oceanic diatom maintain its photosynthetic performance relative to the
coastal diatom. However, the lower iron-requirements of the oceanic diatom result in a reduce
ability to acclimate to changing light conditions. Additionally, iron-limitation appears to directly
interfere with carbon uptake and assimilation in phytoplankton (Schulz et al. 2007, Allen et al.
2008) although the mechanisms by which this occurs are poorly studied.
Insights into the interplay between carbon-and iron-availability and its impact on photosynthesis
of primary producers in the ocean subjected to increasing dissolved CO2 are thus highly
warranted. Subproject 1.1.2 will also provide a basis for better understanding the impact of open
ocean iron-fertilization, a controversial practice which is pursued by some companies (Buesseler
et al. 2008). In a high CO2 ocean the proposed study will bring additional knowledge essential in
assessing the effectiveness of open ocean Fe fertilization as a strategy to mitigate global
atmospheric CO2 increase. A recent transcriptomic study identified genes involved in silicon
bioprocesses in T. pseudonana (Mock et al. 2008) proving the suitability of this technique in the
analysis of the biology of diatoms. In Phaedactylum tricornutum, a species that shows a high
tolerance to iron limitation, several genes involved in Fe acquisition and in the remodeling of the
photosynthetic apparatus have been identified through a whole-cell system’s approach (Allen et
al. in press). Several of these genes are absent in the T. pseudonana genome, pointing to a genetic
basis for the phenotypic difference in their acclimation to iron limitation. An additional genome
project for the oceanic diatom T. oceanica, has been initiated jointly at the IFM-GEOMAR and
University of Kiel CAU. The outcome of this genome project together with the genome sequence
of T. pseudonana and P. tricornutum will provide a solid basis for comparative transcriptomic
and proteomics analyses (see also subproject 1.1.4).
Most experiments done so far examining phytoplankton are restricted in time and are observing
only a few generations of these single cell organisms. Regarding the fast generation time of most
72
phytoplankton groups (~one division per day) a possible acclimatisation and/or adaptation has to
be considered for prediction of future phytoplankton response (Collins and Bell 2004). Current
primary producers have co-evolved with a partial pressure of CO2 that never exceeded 300 ppm
in the past 20 Mio years. Conversely, evolutionary changes are to be expected as a response to
ongoing increases in pCO2. Such evolution in action is expected to be particularly rapid in
microalgal species having short generation times, such as unicellular plankton and bacteria.
Strain-to-strain differences of Emiliania huxleyi in their sensitivity to CO2 enrichment indicate
standing genetic variation for traits related to carbon acquisition under altered CO2 chemistry
(Delille et al. 2005). Interestingly, the only long-term experiment addressing evolutionary
changes in a microalgal species over approximately 1000 generations revealed that the green
unicellular algae Chlamydomonas rheinhardtii lost its carbon concentrating mechanism (CCM)
in response to increased pCO2. These findings are in line with evolutionary theory, predicting that
costly carbon concentration is no longer selected for in a CO2 rich environment. If long-term
exposure to increases in pCO2 leads to degeneration of CCMs, then short-term responses of
phytoplankton, for example as higher export production, may only be transient. On the other
hand, under sexual recombination, positive selection of novel physiological function may also
take place that produce physiological trait values outside the boundaries of original trait
distributions (Reusch and Wood 2007)
Because long-term effects are crucial but understudied in the marine plankton and
chemoautotrophic bacteria, three subprojects will address long-term physiological responses
(subproject 1.1.1 Jost/Jürgens; 1.1.3 Müller; 1.1.4 Reusch/Riebesell) and compare the
reversible plastic acclimation response with longer-lasting adaptive changes. Changing responses
of important target species to carbon enrichment have already been identified in pilot
experiments (cf. Fig. 1.1.1a,b). Cultures of coccolithophores showed a higher sensitivity for high
pCO2 regarding their division rates in comparison to short-term experiments (Riebesell et al.
2000, Langer et al. 2006). Responses of phytoplankton to single parameters (e.g. temperature,
pCO2 and calcite saturation state) are still in the progress of investigation. However, in nature all
physical parameters are predicted to change more or less in the next centuries. Therefore,
subproject 1.1.3 will also investigate any interaction effects of the two most rapid changing
parameters (temperature and pCO2) on different phytoplankton key groups (diatoms,
coccolithophores and calcareous dinoflagellates), which are the main primary producers
(diatoms), the main pelagic producers of biogenic calcite (coccolithophores and calcareous
dinoflagellates) and jointly contribute to the basis of the oceanic food web. A strong link will be
forged to subproject 3.1.1 that focuses on changes in calcification rates of coccolithophores.
In order to verify whether or not physiological changes are due to genetic adaptation, we aim at
identifying the genetic basis of observed changes using state-of-the art genomic and
transcriptomic techniques in two species where genomic and transcriptomic resources are readily
available, Thalassiosira pseudonana and Emiliania huxleyi (subproject 1.1.4 Reusch
/Riebesell). In these fast generation species, rapid evolutionary change may take place through
the combination of any of the following three mechanisms (i) clonal selection, whereby preadapted multi-locus genotypes outcompete others by mitotic division and population increases
(ii) de novo mutation in particular genotypic lines (iii) genetic recombination which may bring
trait values beyond the range of values seen in the original population. Recombination is
expected to bring together favourable mutations, while breaking up negative gene associations.
For E. huxleyi, culture conditions are established that allow us to introduce intermittent rounds of
sexual recombination during the course of the experiment (Laguna et al. 2001), permitting a test
for the role of sexual reproduction in promoting evolutionary adaptation. Genomic approaches
will be coordinated with subproject 1.1.2 for the diatom transcriptomic and proteomic analysis.
73
As additional response variables, information on the elementary composition of primary
producers will provide important information for subproject 4.2.1. that addresses the effects of
food quality on higher trophic levels.
Fig. 1.1.1a: Division rates of three coccolithophore species
under high pCO2 (filled red) and ambient pCO2 (green)
over time. Open red circles indicate the transition from
ambient to high pCO2. The percental decrease in division
rate is shown by the red numbers. Müller et al unpublished
Fig. 1.1.1.b: Change of the optimal division curve of
Emiliania huxleyi from ambient pCO2 (green) to high
pCO2 (red) in relation to different temperatures. Müller
et al unpublished
Benthic diatoms are another key primary producer guild with important ecological role on
intertidal and shallow subtidal marine sediments along the German coastline of the North and
Baltic Sea, being responsible for particularly high rates of elemental cycling and high primary
productivity. These communities provide a major food source (e.g. fatty acids) for benthic
suspension- or deposit-feeders, and act as control barrier for oxygen fluxes at the sediment/water
interface, and as stabilizer of sediment surfaces against erosion by the excretion of extracellular
polymeric substances (EPS) (Cahoon 1999). EPS are mainly composed of polysaccharides and
proteins, and besides their role in biostabilisation of sediments they are involved in gliding
motility and adhesion.
Despite their importance for production and elemental cycling almost nothing is known on the
ecophysiology and production biology per se, and even less on the interactive effects of rising
CO2 concentrations and temperatures on microphytobenthic primary production (Forster et al.
2006), a research gap that subproject 1.1.5. (Karsten/Hübener) intends to close. Building upon
such ecophysiological data, the relationship between biodiversity of benthic diatom communities
and their productivity will be assessed. Subproject 1.1.5. will also explore whether or not diatom
taxa are valuable indicator species for environmental changes associated with OA, as has been
shown for eutrophication before (EDDI: Battarbee et al. 2000, MOLTEN: Clarke et al. 2002,
2003, Juggins 2004). Notably, these organisms possess small tolerance values for important
control factors of water-quality (pH, nutrients, salinity). As a second step, it will explore how the
diatom community changes as a response to predicted ocean acidification, in combination with
temperature increases.
Subproject 1.1.1: G. Jost has experience in the work with autotrophic prokaryotes and the
analysis of their role in the biogeochemistry of pelagic redoxclines. K. Jürgens has extensive
74
experience in the analysis of prokaryotic and eukaryotic microorganisms, microbial food webs
and trophic interactions, including model systems and field studies in diverse aquatic systems. In
the last years a newly established group at the IOW in molecular and microbial ecology has made
significant progress in the understanding of redoxcline microbial communities (Labrenz et al.
2007, Jost et al. 2008), the identification of bacterial key players (Grote et al. 2007) and major
transformation processes (Hannig et al. 2007).
Subproject 1.1.2: Prof. J. LaRoche is an expert in marine phytoplankton ecology and
ecophysiology. She has performed and participated in seminal experiments on the role of iron on
phytoplankton productivity and physiology (LaRoche et al. 1996, McKay et al. 1997, Boyd et al.
2000, Jickells et al. 2005, Allen et al. 2008). She is also an expert in diatom transcriptomics. M.
Hippler is an expert in algal proteomics and physiology and particularly in adaptive remodelling
of the photosynthetic apparatus to iron-deprivation (Moseley et al. 2002, Naumann et al. 2005,
Allmer et al. 2006, Naumann et al. 2007). This combined expertise in physiology, transcriptomics
and proteomics will ensure the successful complementation of the outlined working program.
Subproject 1.1.3: M. Müller is currently working in the ESF project ’Casiopeia’ as a PhD,
finishing his doctoral degree in autumn 2008. He previously worked on coccolithophores (Müller
et al. 2008b) and gained over the last years a solid expertise in culturing phytoplankton in dilute
batch cultures and semi-continuous cultures under a manipulated carbonate system (Müller et al.
2008a). At the moment a variety of coccolithophorid species are cultured since several years and
will be available for experimental work in this project.
Subproject 1.1.4: The expertise and research interest of both applicants are mutually overlapping.
U. Riebesell is an expert in marine phytoplankton ecology and ecophysiology (Riebesell et al.
2000, Riebesell 2004, Riebesell et al. 2007). T. Reusch has broad experience in design and
execution of selection experiments (Reusch and Wood 2007, Wegner et al. 2007), and in
characterizing the genetic basis of phenotypic change (Wegner et al. 2003), including
transcriptomic work (Reusch et al. 2008)
Subproject 1.1.5: U. Karsten has worked with benthic diatoms from brackish and marine waters
and successfully established a culture collection of polar diatoms at the University of Rostock.
The research interest of Prof. Karsten is mainly related to the ecological and physiological
performance of these microalgae under different environmental conditions, as well as to the
development of new methodological approaches (e.g. Karsten et al. 2006; Wölfel et al. 2007;
Gustavs et al. 2008). T. Hübener has a long record in studies on diatom ecology and taxonomy
(Dreßler and Hübener 2006, as well as on diatoms as indicators for environmental change
(palaeolimnology) (e.g. Adler and Hübener 2007, Hübener et al. 2007). The expertise of U.
Karsten and T. Hübener is well reflected in numerous grants funded and refereed publications.
iv. Work Programme, Schedules and Deliverables
Work Programme – general
The experimental levels and the rate of increase in pCO2 as well as other important experimental
manipulations will be coordinated as much as possible especially with the experiments of
Jost/Jürgens, LaRoche/Hippler, Müller, and Reusch/Riebesell. We also plan to share some of the
experimental set-ups, in particular the chemostats, among the subprojects. All projects have
agreed upon the specific levels of pCO2 for their experimental manipulation, namely the five core
treatment levels 280ppm (pre-industrial), 380 (present day), 560ppm (2x pre-industrial), 700ppm
75
(IPCC business as usual for year 2100), 1000ppm (3x pre-industrial). These treatment levels
should be established by a slow change from the present day level.
Subproject 1.1.1 Impact of changing pCO2 and temperature on a chemolithoautotrophic
Epsilonproteobacterium from a pelagic redoxcline
G. Jost & K. Jürgens
Work Programme
The response of microbial populations at pelagic redoxclines to changes in single parameters
(pCO2, pH, and temperature) will be studied in controlled experimental laboratory systems
(diluted batch and chemostat cultures, including a comparison between both systems). The effects
of ambient pCO2 and slowly increasing pCO2 on the performance of a representative
chemoautotrophic prokaryote will be compared. Additionally, it will be investigated whether
increasing pCO2 in combination with decreasing pH or only changes in the pH are the important
factors. The selected model organism epsilonproteobacterium strain GD1, which is abundant in
the redoxclines of the central Baltic Sea (Grote et al. 2007) will be used for selected process
studies (e.g. chemolithotrophic denitrification). The changing performance of this strain in
response to the investigated factors will be compared to relative changes in the performance of
other single organisms (1.1.3, 1.1.4) as well as whole (bacterial) communities (1.1.2, 1.1.5,
1.2.4). The performance of the microorganisms will be assessed as specific growth rate,
elemental composition (CNP) and related chemical transformations (e.g., sulphur oxidation,
denitrification). To differentiate between phenotypic and genotypic long-term adaptations
molecular biological methods will be applied (e.g. differential gene expression of functional
genes) in cooperation with other projects.
In order to assess the acclimation potential, different rates of increase pCO2 attaining the final
treatment levels will be compared. Finally, in line with sub-projects 1.1.3 and 1.1.5, the
combined versus single effects of temperature and pCO2 will be assessed in factorial experiments.
The results will be used for biogeochemical modelling, including the evaluation of the influence
of proton activity on biogeochemical processes. Additional, a comparison of the bacterial
population at the end of the long-term experiments grown under different conditions will be done
not only by investigating phenotypic performances but also genotypic differences (e.g.
differential gene expression of selected functional genes). A comparison of the results of the
reaction a single bacterium will show on changing conditions related to climate change with
observations of carbon dioxide fixation in CO2-enriched sediments and the appearances of
different phylogenetic groups of bacteria including epsilonproteobacteria (subproject 4.1.4.1)
may give a hint on possible extrapolations of the results even to sediments.
Work Schedule
1.1.1
First Year
I
Set-up of culturing facility, instrument
calibration
76
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
1.1.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Diluted batch experiments
Long-term chemostat experiment
Sample processing and measurements
Combined CO2/tempertaure
perturbation experiments
Molecular investigations
Data analysis, statistical evaluation,
data interpretation
results at conferences, PhD paper
Milestones (1.1.1)
- Implementation of experimental facility for controlled continuous cultures
- Experimental data set on CO2/pH sensitivity of epsilonproteobacterium GD1
- Data set on synergistic effects of CO2 and temperature on strain GD1
- Evaluation of combined data sets, sensitivities and uncertainties
month 9
month 18
month 24
month 33
Subproject 1.1.2 The interplay between carbon-and iron-availability and its impact on
photosynthesis of primary producers in the ocean
J. LaRoche & M. Hippler
Work programme
We will study the effects of ocean acidification on photosynthesis and carbon assimilation in
diatoms, exposing them to iron-limited and replete conditions. Physiological approaches will be
combined with systems biology approaches where transcriptomics and proteomics will be
applied. (i) Physiological measurements of growth and photosynthetic performance for a oceanic
and costal diatom strain will be conducted at distinct CO2 partial pressures of 380, 700 and 1000
ppm combined with iron-sufficient and –deficient conditions. (ii) Comparative transcriptomic
profiling of the two diatom strains T. pseudonana and T. oceanica will be performed under the
outlined physiological conditions taking advantage of 454 DNA sequencing (Eveland et al. 2008)
to elucidate dynamics in gene expression in regard to different carbon and iron resources. (iii)
Comparative and quantitative proteomics will be performed to unravel alterations in the proteome
with a focus on plasma and thylakoid membranes in response to iron and carbon availability. In
particularly we will concentrate on changes of the photosynthetic apparatus. Here we will take
advantage of high precision and resolution mass spectrometry using an LTQ-Orbitap mass
spectrometer (ThermoFisher) and proteotypic peptide profiling.
The project is complementary to 1.1.1 and 3.3 which also study the effect of OA and pCO2
increase on the speciation of trace metals and their effect on chemoautotrophs and calcifyiers.
The result of our study on short term effects of high CO2 on the photosynthetic apparatus of
77
diatoms can be used to help predict the long term genetic effects of elevated CO2 studied in
1.1.4. The results will also be used in biogeochemical models (1.3).
Work Schedule
1.1.2
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Establish trace-metal buffered cultures
Experiments with T. oceanica
Experiments with T. pseudonana
Establishment of proteomic techniques
with T. oceanica
Transcriptomic analysis T. oceanica
Data analysis & synthesis
Establishment of proteomic techniques
with T. pseudonana
Transcriptomics T. pseudonana
Data analysis & synthesis
PhD. Thesis preparation
Milestones (1.1.2)
- Establishment of semi-continuous trace metal-buffered culture system and CO2 levels
- Experimental data set with T. oceanica grown at varying CO2 and iron levels
- Experimental data set with T. pseudonana grown at varying CO2 and iron levels
- Evaluation of combined data sets
Subproject
1.1.3
Long-term
response
of
phytoplankton
on
climate
month 6
month 18
month 27
month 36
change:
a
multidimensional approach
M. Müller
Work programme
For each phytoplankton species (Thalassiosira pseudonana, Emiliania huxleyi and
Thoracosphaera heimii) incubations will be run in duplicate under lab-controlled conditions
(light, temperature, nutrients and carbonate system) over 1.5 year what corresponds to 500 to
1000 generations depending on the species. For this purpose 24 chemostat units will be combined
in a computer-controlled incubation system where the pCO2 and the temperature can be slowly
increased to a certain level. The newly constructed CO2 aeration system at the IFM-GEOMAR is
a ideal tool to set up a chemostat system where the carbonate system can be controlled in a
78
appropriate manner. The system will be monitored by dissolved inorganic carbon, alkalinity and
nutrient analysis carried out in the lab of U. Riebesell. Primary response variables that are
coordinated throughout projects 1.1.2 – 1.1.5 are division rates, photosynthesis, calcification rates
and chlorophyll production. Photosynthesis and calcification rates will be measured at the isotope
lab using radioactive 14C accompanied by carbon acquisition determination with the Inlet-MassSpectrometry (MIMS) in cooperation with K. Schulz (3.1.1). Division rates and chlorophyll
autofluorescence will be determined by flow cytometry. Genetic analysis at the end of the
experimental exposures will be realized in cooperation with J. LaRoche, T. Reusch and M. Bleich
(1.1.2; 1.1.4 and 3.1.4), additionally we will identify stress proteins in the named species in
regard to pCO2 and temperature (LaRoche, 1999). Calcite analytics (δ13C, δ18O, δ11B) can be
performed in the geochemistry lab at the Ruhr-University Bochum (3.5.2) and in collaboration
with subproject 3.5.3 (S. Meier). During the acclimation phase we plan to analyze the expression
of stress proteins such as heat shock proteins (hsps) in order to identify surrogates of the
acclimation response.
Work Schedule
1.1.3
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Establishing chemostats
Long-term CO2 enrichment exps
Physiological measurements
Identify stress proteins in dilute batch
cultures
Quantification of stress proteins in
chemostats
Synthesis of results & analysis
Milestones (1.1.3)
- Completion of chemostat system and data set on facility-accuracy
- Identification of stress proteins
- Data set on synergistic effects of CO2 and temperature under long-term conditions
- Evaluation of combined data sets (with all collaborators), sensitivities and
month 9
month 15
month 30
month 32
79
Subproject 1.1.4 Rapid evolution of key phytoplankton species to a high pCO2 ocean
T. Reusch & U. Riebesell
Work programme
We propose to establish replicated selection lines with large population sizes (>106 cells) in two
important members of the marine phytoplankton, a diatom (Thalassiosira spp.) and a
coccolithophorid (Emiliania huxleyi) under ambient (= present day; 380 ppm) and increased
pCO2 (~700 ppm/1000 ppm). Ideally, pCO2 will be increased by bubbling enriched atmosphere
into culture water. Experiments will start from novel isolates obtained from natural
phytoplankton assemblages (Rynearson and Armbrust 2000). Control populations /lines in all
diversity levels will also be established and subjected only to laboratory selection under ambient
pCO2. Cultures will be run for at least 500 generations (~6-12 months), with some replicates
being shared among the subprojects (i.e. 1.1.3 Müller).
Using diagnostic microsatellite markers in combination with quantitative real-time PCR, we will
closely follow the genotypic composition of all multi-clonal populations to detect genotypic
selection, i.e. the dominance of certain pre-adapted genotypes over others. Results from this part
of the project will be compared to results from project 4.1.2 (M. Wahl) that addresses the role of
genetic diversity for juvenile survival in invertebrates. Because recombination may enhance
selection responses, in E. huxleyi, half of the experimental lines will undergo sexual reproduction
every 10 – 50 mitotic cells cycles (Laguna et al. 2001). The published methods may have to be
modified in order to allow several (~10-20) intermittent sexual generations. Relative fitness of
selected and ambient replicates will be assessed singly, and as direct competition experiments
under a reciprocal combination of selection regimes. Response variables to be monitored every
50 mitotic generations will be coordinated throughout 1.1.2 – 1.1.4 and include photosynthetic
rates, pigment content/composition, division rates, chemical stoichiometry, calcification rates and
stable isotope composition. The latter data will be interconnected with project 4.2.1 (M.
Boersma) that studies the effects of changing chemical composition of phytoplankton for
zooplankton. As in subproject 1.1.3, calcification rates will be measured at the isotope lab using
radioactive 14C accompanied by carbon acquisition determination with the Inlet-MassSpectrometry (MIMS) in cooperation with K. Schulz (3.1.1). In order to identify the genetic basis
of adaptive evolution in a high pCO2 ocean, we will assess the transcriptomic response of both
selection lines (ambient / high pCO2) in full combination with the actual test regime (ambient /
high pCO2). In E. huxleyi, transcription profiling will be done using tagged 3’-expressed
sequence tags in combination with direct 454 DNA sequencing (Eveland et al. 2008). In
Thalassiosira, we use the whole genome tiling array developed by (Mock et al. 2008). Changes
in transcriptomic response in Thalassiosira will be compared to responses obtained in project
1.1.2 (J. LaRoche) where the response to iron deficiency is studied. Interpretation of data on
evolutionary responses will be closely discussed with the long-term data on the fossil record
obtained through project 3.5.2 (J. Mutterlose).
80
Work Schedule
1.1.4
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Collect field isolates, establish
Selection lines
Induction of sexual reproduction
Establish Q-PCR to discriminate
genotypes
Run evolution experiment
Test for adaptation in reciprocal
experiment
Perform transcriptomic analyses
data interpretation
results at conferences, PhD thesis
preparation
Milestones (1.1.4.)
- Selection lines are running
- Sexual Induction possible
- Experimental evolution lines finished (~1000 generations)
- Transcriptomic data obtained
- Bioinformatic analysis of evolutionary changes
month 9
month 12
month 24
month 26
month 33
Subproject 1.1.5 Interactive effects of CO2 concentration and temperature on
microphytobenthic biodiversity and ecosystem function
U. Karsten, T. Hübener
Work Programme
The main goal is to evaluate the interactive effects of rising pCO2 and temperature on
microphytobenthic ecophysiology, biodiversity and primary production under controlled
laboratory conditions and at representative shallow water stations in the Baltic Sea. In order to
test species specific responses of benthic diatoms to a range of CO2 concentrations and
temperatures we expose unialgal cultures of dominant species from experimental habitats in
laboratory cultures and measure photosynthesis, division rates and primary production under
different light, CO2 and temperature conditions using fluorimetric techniques and O2 optodes. For
the improvement of the methodological approach strong collaboration with project 0.3.2.
(Apostolidis/Huber, Presens) is intended. The response parameters will be compared to relative
changes in the performance of other aquatic organisms and communities within theme 1.1.,
81
particularly project 1.1.1 (Jost/Jürgens) as well as to other macrobenthic primary producers in
project 4.1.1 (Asmus). As a next step, in order to study the community level response, we will
artificially mix communities with up to 10 diatom taxa. Species selection will be guided by
abundance estimates from natural sediment communities of the shallow Baltic Sea. Community
experiments will allow us to test the hypothesis that there is a predictable shift in taxonomic
distribution upon OA, possibly interacting with temperature, and driven by different
ecophysiological requirements obtained from previous experiments. Microphytobenthic primary
production will be assessed via area-based biomass determination (standing stock, chlorophyll a
content in sediment cores). We also apply in situ primary production measurements using new
benthic chambers with planar O2 optodes and puls amplitude modulation (PAM) fluorometry.
There are many scientific overlaps to projects 3.4.1 and 3.4.2 concerning the metabolic activity of
benthic microorganisms and their effects on the water column, sediment and porewater
chemistry.
Work Schedule
1.1.5
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
Isolation, identification,cultivation of
diatom species
Assessment of natural diversity
Ecophysiological measurements
Mesocosm experiments on primary
productivity versus diversity
Data analysis & completion of PhD
thesis
Milestones (1.1.5.)
- Field data on natural biodiversity
- Unialgal cultures for experimental studies
- Ecophysiological response profiles for CO2 x temperature conditions
- Experiments on primary production as function of pCO2 and temperature
- Mesocosm data on diversity versus primary productivity
- Statistical evaluation of data, Interpretation, conclusion
month 6
month 9
month 18
month 24
month 30
month 34
vi. Budget and Budget Justification
First Year
Personnel costs
1.1.1 (PhD)
1.1.1 (HiWi)
1.1.2 (PhD)
1.1.3 (Post Doc)
1.1.3 (HiWi)
82
Second Year
Third Year
Total
IV
First Year
Second Year
Third Year
Total
1.1.4 PhD (Reusch)
1.1.4 Tech (Riebesell)
1.1.5 (PhD, tech supp.)
Subtotal
Consumables
1.1.1
1.1.2 Kiel
1.1.2 Münster
1.1.3
1.1.4
1.1.5
Subtotal
Travel
1.1.1
1.1.2
1.1.3
1.1.4
1.1.5
Subtotal
Investment
1.1.1
1.1.2
1.1.3
1.1.4
1.1.5
Subtotal
Other costs
1.1.1
1.1.2
1.1.3 pub charges
1.1.4 (Münster)
1.1.5
Subtotal
Total
Budget justification 1.1.1.
Personnel costs: One PhD-student, will establish an anaerobically run chemostat system,
allowing the application of changing pCO2 for long term cultivation of the isolated
epsilonproteobacterium strain GD1. During the period of establishing the chemostat system
already diluted batch experiments will be done to investigate the effect of changing pCO2 versus
only pH changes on the growth of the selected strain. The proposed student assistant will help in
the preparation of media for cultures and during sampling of the chemostat experiments as well
as in preparation of samples for elemental and flow cytometric measurements.
83
Consumables: Will cover the preparation of media for the chemostat and batch cultures,
measurements of bacterial concentrations by flow cytometry, comparisons with epifluorescence
microscopy, estimation of elemental composition of the bacterial biomass, estimations of the
concentrations of the chemical reactants, radioactive substrates and chemicals for rate
measurements, chemicals for molecular biological methods to estimate genotypic differences.
Travel: Project collaborators will take part in workshops ensuring comparative measurements of
the pCO2 and the establishing the proposed concentrations in the different experimental systems
as well as in workshops concerning the construction and running of chemostats with constant and
slowly increasing pCO2. Especially, regular exchanges will take part during the establishing of
the chemostat system with the WP (M. Müller). Participation within regular annual project
meetings as well as in international conferences, exchanging ideas and presenting the results of
the project, is also necessary.
Budget justification 1.1.2
Personnel costs: The Ph.D. student position will be split between the two institutions as follows:
2 years for J. LaRoche in Kiel and 1 year for M. Hippler in Münster. The Ph.D. student from the
LaRoche lab will be responsible for setting up the trace metal buffered cultures at the various
CO2 and Fe levels and for making the basic physiological measurements (photosynthesis, growth
rate, iron uptake, flavodoxin accumulation, PAM fluorometry). The student will also collect the
samples for transcriptomics and proteomics. He will be responsible for the RNA extraction and
transcriptomics analysis.
The Ph.D. student (one year) in the Hippler lab will be responsible for the analysis of the
proteomics samples grown at the various CO2 and Fe levels.
Consumables: The consumables will be used to pay for the cost of sequencing with the 454
tagged pyrosequencing (estimated at
for 6 conditions run in duplicates) and for the
proteomics analysis.
Personnel costs: Post Doc is necessary to construct and maintain the required chemostat system.
Additionally, to culture the organisms over long time scales reliable skills are essential what is
not fulfilled by a PhD student and even a Post Doc will need additive help (student assistance).
Consumables: To sample and monitor the chemostat system a bunch of consumables are needed
and analysis is the base for publications.
Travel: Visiting international conferences is essential to present the latest results and to keep in
contact with the scientific community.
Investment: The construction of the chemostat system will provide a unique platform to
investigate the combined effect of CO2 and temperature on phytoplankton under completely
controlled lab conditions over a long time scale. Accessorily, it provides the possibility to
identify acclimatization and/or adaptation in marine organisms over time. This system has to be
controlled by a set of intertwined computer framework. Other costs: Flow cytometric
measurements are needed to monitor the cell abundance in the chemostat system.
84
Personnel costs: One technician (
) for maintaining the selection lines,
will be shared with project 1.1.3. Selection lines need to be replicated under different pCO2
conditions. For E.huxleyi, they will be combined with inducing intermittent sexual phases. For
both species we plan to establish 48 batch cultures of 1.5 L in total which will be very time
consuming. One PhD student will focus on the induction of sexual phases, on developing the QPCR assays, and on trasncriptomic analyses at the end of long-term experiments.
Consumables:
for general maintenance of cultures, continual physiological
measurements, genetic analyses for determining genotypic composition (clonal selection) using
Q-PCR and clone specific microsatellites
Travel: paid by institute. Investment: none required
Other costs: replicated transcription profiling of evolved and non-evolved strains under ambient
and novel conditions (complete reciprocal design;
for transcriptomic analyses using
454 tagged pyrosequencing (Emiliania),
for whole genome transcript profiling
(Thalassiosira) using whole genome tiling array designed by (Mock et al. 2008).
Personnel costs: One PhD-student (
) for evaluation of morphometric and molecular
diversity shifts in different microphytobentic habitats (field work) and artificial lab-communities
(mesocosms, incl. isolation, cultivation) under different pCO2 and temperature conditions.
Students technical support (
) for maintaining stock cultures, growth experiments under
different light, pCO2 and temperature conditions(growth fluorimeter for benthic microalgae) and
primary production measurements with O2 optodes with unialgal cultures and defined mixed
benthic diatom communities.
Consumables:
for chemicals, glass and plastic vessels for culturing, microscopy,
chemical analysis, molecular biology.
Travel:
for field work, participation in workshops ensuring comparative measurements
of pCO2, in annual project meetings and in international conferences presenting results of the
project.
Investments:
once; CO2 aeration system for the mesocosm, growth and primary
production measurements, multichannel O2 optode system.
85
Project 1.2 Turnover of Organic Matter
Anja Engel (AWI, Bremerhaven), Maren Voss (Leibniz Institute Warnemünde), M. Nausch/ G.
Nausch (Leibniz Institute Warnemünde), Hans-Peter Grossart (IGB, Berlin), Laurenz Thomsen/
Giselher Gust (Jacobs University Bremen)
i. Objectives
This projects aims to elucidate the effects of ocean acidification on the turnover of organic matter
in pelagic ecosystems. During field and laboratory studies, we will quantify and characterize how
the production, exudation and microbial processing of organic matter (OM) respond to changes in
seawater pCO2 and pH. Specifically, the project will investigate:
•
physiological and community structure responses of marine microbes. This will enable
us to differentiate between functional and structural changes of planktonic communities.
•
the turnover of biochemical key components, such as polysaccharides, proteins and
organic phosphorus compounds, and the resulting changes in the C:N:P stoichiometry of
organic matter.
•
how the remineralisation and final deposition of sinking OM, i.e. aggregates, will
become affected by potential changes in mineral ballast.
Our ultimate goal is to contribute to a better understanding of the direction and strength of
biogeochemical feedback processes in the future ocean. We aim to generate reliable data on
highly variable chemical and microbial processes in order to provide a better basis for modelling
and future prognoses on the ecological consequences of ocean acidification.
There is now increasing awareness that ocean acidification will affect marine algae and biogenic
production in the ocean (Orr et al. 2005, Arrigo 2007, Riebesell et al. 2007, Fabry et al. 2008).
Direct responses of the microbial food-web to changes in seawater pCO2 and pH, or in the supply
with organic resources are yet little exploited. In order to estimate the sensitivity of
biogeochemical cycles to ocean acidification, an improved understanding of potential changes in
the turnover of organic matter during production and decomposition processes is urgently
needed.
Recent studies indicated that the production of acidic polysaccharide particles, also known as
transparent exopolymer particles (TEP), is sensitive to changes in seawater CO2 concentration
(Engel, 2002; Engel et al., 2004a, Mari 2008). Acidic polysaccharides are exuded by
phytoplankton cells as a carbon-rich ‘overflow’ product of photosynthesis under nutrient
depletion (e.g. Obernosterer and Herndl, 1995; Biddanda and Benner, 1997; Søndergaard et al.,
2000), or derive from bacterial oxidation of dissolved carbohydrates (Giroldo et al., 2003).
Acidic polysaccharide can facilitate the bonding between organic components and therewith
affect particle stickiness, and the partitioning between the pools of dissolved and particulate
organic matter, and organic matter export (Logan et al., 1995; Engel 2000, Passow 2002). On the
other hand, varying concentrations and chemical signatures of released organic matter potentially
can affect the activities and community composition of heterotrophic bacteria (e.g. Biersmith and
Benner, 1998; Grossart et al., 2005, 2006a). Thus, besides exudation and aggregation, the
86
concentration of polysaccharides in seawater is determined by bacterial decomposition processes,
which may respond differently to ocean acidification (Piontek et al., 2007a, b).
In general, elemental cycles do not exist independently from each other in marine ecosystems.
The cross-linking of C, N and P-cycles is pronounced in heterotrophic bacteria, as the availability
of DOC has consequences for P utilization and can decide if DIP or DOP is the preferred
compound (Hoppe and Ullrich 1999, Nausch and Nausch 2004, 2007). Changes in carbon
exudation due to ocean acidification may therefore affect the phosphor demand of bacteria
(Tanaka et al. 2007). On the other hand, N2-fixation rates can also be enhanced under high CO2
(Hutchins et al. 2007, Ramos et al., 2007). DOM release can reach up to 40% of assimilated N
(Bronk et al., 1994), indicating that DON- and DOC- release are tightly coupled (Bronk. and
Ward, 1999). The regulation of release and uptake processes in the environment is poorly
understood (Bronk et al., 2007). Potentially, competitive interactions for resources may define the
community of autotrophs and small heterotrophs in the field. To reliably predict organic matter
turnover in the future, a better understanding on factors regulating DOM release and uptake and
their sensitivity towards changing pCO2/pH are necessary.
Export of organic carbon into the benthic boundary layer (BBL) is enhanced by acidic
polysaccharides and by mineral ballast (Passow 2002, Armstrong et al., 2002; Klaas and Archer
2002). Recent studies indicate that ballast through calcification can either be decreased or
increased due to CO2-induced changes in seawater chemistry (Fabry, 2008). Until final burial
and once on the sea floor, organic aggregates are more easily remobilised into the benthic
boundary layer than the bulk sediments beneath, and are frequently resuspended and hence
modified in the water column (Thomsen et al., 2002, Keil et al., 2004). After an extended
residence time of weeks to month, oxygen consumption within most organic aggregates is often
similar to that of the background sediment and produces a minimal impact on sediment
mineralization rates (Beaulieu and Smith 1998.). The part of this organic matter that seems too
refractory to be recycled is buried in ocean sediments, sequestering carbon for long time periods
and consequently influencing atmospheric carbon dioxide concentrations (Hedges et al., 2002).
Bioavailability of aggregates in the BBL may be more related to their organo-mineral content
than to the molecular composition of organic matter and consequently changes in mineral ballast
will affect future carbon sequestration.
Subproject 1.2.1: A. Engel (AWI): A. Engel has comprehensive experience on studying CO2
effects on marine ecosystems, mineral ballasting, and in particular on organic matter exudation
(i.e. Engel 2002, Engel et al. 2004a,b, Engel et al. 2005, Engel et al. 2008, Engel et al. in press).
Since 2005, she leads a Helmholtz Young Investigators Group to investigate global change
effects on marine biogeochemistry. The group has been involved in several national and
international programs dealing with potential effects of global change (PeECE, PEACE,
AQUASHIFT, SOPRAN, and EPOCA). A Ph.D. thesis has been performed dealing with the
effects of ocean acidification on organic matter enzymatic hydrolysis and bacterial
decomposition in calcifying algae.
Subproject 1.2.2: M. Voß (IOW): M.Voß co-leads the subproject in the SOPRAN project
“Ecosystem response to CO2 enrichment” where the role of nitrogen fixers under high CO2 is
studied in free drifting mesocosms. Within the WGL network, TRACES she supervises a PhD
thesis on the DON release of cyanobacteria and the transfer of fixed nitrogen into higher trophic
levels. She worked on the regulation of N-fixation in various regions (Wasmund et al., 2005;
Wasmund et al., 2001, Voss et al., 2006, Capone et al., 1998, Voss et al., 2004). Voss leads the
87
“N-cycle group” at the IOW consisting of 5 externally funded people with a special focus on
marine nitrogen cycling, nitrogen fixation and stable isotope ecology.
Subproject 1.2.3: M. and G. Nausch (IOW): Phosphorus dynamics in surface water was a major
topic of research during the last years and resulted in the formation of an interdisciplinary
“phosphorus” group (2 senior scientists, 1PhD, 3 diploma students, 1 technician). Substantial
investigations on alkaline phosphatase activity as an indicator for P-limitation were done (Nausch
1998; Nausch 2000; Nausch and Nausch 2000). A new method for the determination of
bioavailable DOP (BAP) was established (Nausch and Nausch 2004) allowing investigations of
BAP and its importance for phytoplankton and bacterial nutrition (Nausch and Nausch 2006,
2007). Currently, they participate in the SOPRAN-project “Ecosystem response to CO2
enrichment”.
Subproject 1.2.4: H.-P. Grossart (IGB): H.-P. Grossart has intensively worked on
phytoplankton-bacteria interactions, especially on effects of phytoplankton release, its subsequent
aggregation and on mineralization by specific bacterial communities (Grossart et al., 2005,
Grossart et al., 2006a+b, Grossart and Simon 2007, Grossart et al. 2007). He has participated in
the PeECE II and III studies, collaborating with national and international partners (Løvdal et
al.,2008, Tanaka et al. 2008, Allgaier et al. 2008, Riebesell et al. 2008). Grossart et al. (2006c)
showed effects of pCO2 on bacterial abundance and activities, which was mainly linked to algal
and particle dynamics. H.-P. Grossart is currently collaborating with M. Lunau, A. Engel, and M.
Voss in the SOPRAN mesocosm studies.
Subproject 1.2.5: L. Thomsen/ G. Gust (Jacobs University): L. Thomsen has long experience in
BBL studies at continental margins, (OMEX, HERMES, ESONET, CORAMM). G. Gust has
developed and utilized high pressure laboratories with decompression free access and benthic
chambers (LOTUS, OMEGA, GRAL, SEDYMO). Joint studies Thomsen/Gust indicate that with
increasing residence time within the benthic boundary layer, a carbon protection mechanism is
built up through further aggregation processes, which reduces their bioavailability; including a
pressure effect (Thomsen and McCave, 2000; Thomsen and Gust, 2000; Thomsen et al., 2002;
Thomsen, 2004; Tengberg et al., 2004; Mendes et al., 2007, Kleeberg et al., 2008; Bigalke et al.,
in press).
iv. Detailed Description of the Project Work Plan
Description of common experiments and field studies
To establish a close collaboration between partners within this project, IOW, IGB, JU, and AWI
will perform joint laboratory experiments, using natural and cultured key microorganisms
(diatoms, cyanobacteria, heterotrophic bacteria) under CO2 perturbation. The group of A. Engel
(AWI) provides a system of five CO2, T, nutrient and light controlled chemostats (10L, Fig. 1.3).
Chemostat experiments will be set up for five different core CO2 concentration with two groups
of primary producers (diatom and nitrogen fixer) in two separate experiments in the 1st and 3rd
year. Those experiments will run over 3 month periods with sampling of subprojects 1-4 for their
respective programs (see below). We will study changes in chemical composition as well as
microbial colonization and decomposition of sinking aggregates in pressure chambers established
by Thomsen/ Gust at JU. In the 2nd year two cruises into the central Baltic Sea are planned for the
investigation of the responses of natural plankton population to CO2 and pH perturbations during
onboard experiments. The cruises will be organized by IOW. Additional laboratory experiments
will be performed to investigate combined effects of CO2/ pH and temperature, pressure, light
and nutrient availability.
88
Subproject 1.2.1 Production and decomposition of exudates
A. Engel (AWI)
We propose to test the following hypotheses:
•
An increase in seawater CO2 concentration stimulates the production and
exudation of exopolymer substances, in particular polysaccharides, by autotrophic
organisms
•
CO2 induced ocean acidification affects the enzymatic hydrolysis of organic matter
and processes to follow, resulting in changes of the microbial decomposition of
exopolymer substances
•
The impact of ocean acidification (OA) on production and decomposition
processes is different for different organic matter components (carbohydrates,
proteins).
Ocean acidification may therefore affect the turnover organic matter in the water-column
differently, with adverse biogeochemical consequences and feedbacks. During the joint activities,
1.2.1 will determine the production and decomposition of exopolymers, i.e. transparent
exopolymer particles (TEP), coomassie stained particles (CSP), and bulk organic matter (POC,
PON, DOC, DON). To better describe processes relevant for biogeochemical feedbacks we will
measure the chemical composition of exudates, i.e. size fractionated neutral and acidic sugars,
and amino acids, together with rate measurements of processes governing their production, i.e.
14
C-exudation, DOM aggregation. Information on the chemical composition of algal exudates at
changing pCO2 will further be used to evaluate the food quality of OM for heterotrophic
organisms in collaboration with 4.2.1.
For comparison between different phytoplankton functional groups exudates production and
composition will be determined in selected samples provided by 4.2.2. To study pH effects on
organic matter decomposition, we will investigate the potential activities of major bacterial
exoenzymes (glucosidases, peptidases, phosphatases, lipases), as well as bacterial production
(thymidine/ leucin), and respiration. A comparison between pH effects on the activities of
bacterial exoenzymes and of those produced by grazers will be made in collaboration with project
4.1.1. Substrate uptake (carbohydrates) by bacteria will be investigated in collaboration with M.
Voss, M. Nausch and H.-P. Grossart.
Information on algal performance during chemostat experiment will be used by modelling
projects in theme 5, in particular in project 5.1, where results on the production of DOM and its
dependence on pH, temperature, and nutrient availability will be used for model refinement, and
by 5.2., where information on the production and decomposition of exudates will be included in a
Bayesian meta-analysis of experimental data
This subproject will also participate in the Svalbard mesocosm experiment planned within the
frame of the European project EPOCA to investigate effects on arctic pelagic communities in
2009.
89
Work Schedule 1.2.1
First Year
1.2.1
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
lab work
field work
Chemostat-Exp.
Synthesis and Publication
Milestones (1.2.1)
- Set-up of improved PCHO/AAS analysis (microwave hydrolysis)
- Dataset from Mesocosm-Experiment Svalbard
- Chemostat data set on CO2/pH sensitivity of diatoms
- Field data set from spring/ summer cruise to the Baltic Proper
- Chemostat data set on CO2/pH sensitivity of cyanobacteria
month 6
month 9-10
month 18
month 24
month 33
Subproject 1.2.2 Dissolved organic nitrogen release and uptake under stress
M. Voß (IOW)
To better understand the release and uptake of DON and uptake of DOC of primary producers
under different pCO2 concentrations and under nutrient stress situations several approaches will
be used. 1. In the IOW batch cultures of diatoms and cyanobacteria under variable pCO2 will be
established together with M. Nausch. The different nutrient stress situations will be limitation of
PO43- for N2-fixers and PO43- and NO3- limitation for diatoms. Additional to this stress 3-5 pCO2
levels will be established. A suite of variables will be then be analysed; nitrogen fixation and
primary production rates, POC/PON, DON/DOC concentrations, DON release and DON/DOC
uptake with 15N /13C labelled substrates. These experiments aim to separate between stresses from
nutrient limitations from the one by acidification 2. We will participate in two 3-4 month long
chemostat experiments at the AWI (see workplan 1.1.1) with again the same group of
phytoplankton organisms and make the same analysis as before. The chemostats will be
investigated by Engel, Nausch, and Grossart at the same time to generate a broad overview over
production and degradation processes. 3. In the field a spring bloom of diatoms and summer
bloom of cyanobacteria will be studied. Here we are faced with a whole plankton community
which will affect rates and processes. All groups will cooperate again during field sampling and
during large volume (0.1m³) mesocosm experiments on board where the plankton community
will be studied under enriched pCO2 conditions.
Relations to theme 4 are given through the study of C:N ratios in phytoplankton species which
are studied in 4.2.2. and in 4.2.1 in different phytoplankton species. We will produce similar
results on relationships between C:N ratios of primary producers and CO2 concentrations in
culture experiments as planned by Rost and Boersma and will compare the results. Furthermore
our data will be used by project 5.2 to improve the model parameterisation of biological
responses to OA.
90
Work Schedule 1.2.2
First Year
1.2.2
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
lab work
field work
Chemostat-Exp.
Milestones (1.2.2)
- Establishing the δ15N -DON measurements at the IOW
- Data from batch culture exp. with cyanobacteria or diatoms
- Field data set from spring and summer cruise to the Baltic Proper
- Data sets from chemostat experiments
- Evaluation of the regulation of DOM release under high pCO2
month 6
month 12
month 24
month 30
month 36
Subproject 1.2.3 DOM availability and phosphorus utilization
M. Nausch and G. Nausch (IOW)
To study the DOM (DOP) release by phytoplankton and the DOM effects on phosphor utilization
by heterotrophic bacteria different approaches will be used. Continuous culture experiments and
batch culture experiments under controlled pCO2 conditions with representative organisms of the
spring and summer bloom (diatoms and cyanobacteria) will be conducted (cooperation with
subprojects 1.1.1., 1.1.2, and 1.1.4). In our responsibility are measurements of DOP and its
constituents (ATP, DNA, RNA, Phospholipids) during the course of the experiments. Two joint
field experiments in spring and summer will show how the whole plankton community will affect
rates and processes compared to the individual organisms in batch and chemostat cultures.
Experiments for DOP utilization by heterotrophic bacteria will be done in batch cultures. Similar
experiments in chemostats are envisaged in cooperation with A. Engel. Supernatants of algal
cultures will be inoculated with bacteria (strain or mixed population) and changes in DOP and
bacterial P will be followed and compared with the turnover of radioactive compounds. The
response of the natural bacterial community to additionally supplied DOC compounds under
variable pCO2 will be investigated. Changes in DOP and bacterial P will be followed. The results
will be incorporated in the carbon cycle model of subproject 1.3. The taxonomic composition of
bacteria in the experiments and the contribution of the different groups to the P uptake will be
studied with molecular biological methods (cooperation with H.-P. Grossart, see subproject
1.2.4).
Our investigations on the effects of ocean acidification on the turnover of organic matter and the
expected changed C:N:P of DOM are urgently needed to explain the hypothesis in subprojects
4.1.1, 4.1.4, 4.2.1 and 4.2.2.
91
Work Schedule 1.2.3
First Year
1.2.3
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
lab work
Field work
Chemostat-Exp.
Milestones (1.2.3)
- Establishing batch cultures with different CO2 levels
- Data from batch and chemostat cultures for DOM production
- Data for DOM effects on phosphorus utilization by bacteria
- Field data from spring cruise to the central Baltic Sea
- Field data from the summer cruise to the central Baltic Sea
- Evaluation of both chemostat experiments
month 3
month 12
month 24
month 18
month 24
month 33
Subproject 1.2.4 Microbial response to DOM release and aggregation
H.-P. Grossart (IGB)
We aim to address the following major questions to achieve a better understanding of the
coupling between ocean acidification and biogeochemical processes mainly controlled by
activities of heterotrophic bacteria:
•
Do changes in pCO2/pH and subsequent changes in particle quality affect bacterial
colonization rate and do they select for specific phylogenetic groups?
•
Do increasing fractions of C-rich phytoplankton-derived matter affect microbial organic
matter remineralisation?
•
Does nutrient availability control microbial degradation of the presumably C-rich
phytoplankton-derived matter and, thus, control the efficiency of the biological pump?
•
Are changes in pH and/or substrate quality reflected by the microbial transcriptome, i.e.
expression of key genes?
For this purpose, batch and continuous cultures of diatoms and cyanobacteria in the lab as well as
mesocosms in the Baltic Sea will be used at different pCO2 concentrations. The experiments will
be jointly conducted with A. Engel, M. Voss, and M. and G. Nausch. We will perform
experiments both at atmospheric pressure (see above) as well as in pressurized incubation
chambers together with L. Thomsen and G. Gust (mimicking particle sinking) to get more
realistic data on pCO2-induced changes of microbial community composition and related
92
processes as well as their potential impact on carbon cycling. In all of our studies, we will
consistently distinguish between free-living and attached bacterial communities. Field
investigations with natural communities will be done in spring during the bloom of diatoms and
in summer during the bloom of cyanobacteria in the Baltic Sea. By simulating various pCO2 and
nutrient concentrations subsequent changes in microbial community structure and function of
specific key players will be studied using molecular (DGGE, RFLP, CARD- and MAR-FISH),
traditional microbial and biogeochemical methods. By combining molecular and biogeochemical
methods, such as Stable Isotope Probing (SIP) we will be able to better link physiological and
community structure response of heterotrophic bacteria to ocean acidification.
Besides the tight linkage between sub-projects of the present cluster this project is linked to the
following projects: 3.4.2 knowledge on the microsensor technique will be exchanged, 4.1.4 direct
effects of ocean acidification on bacteria will be simultaneously measured and knowledge on
physiological effects will be exchanged, 4.2.1 changes in plankton C:N:P ratios and effects on
bacterial populations will be studied together, for project 5.3 our data provide valuable
information for ocean acidification impact assessment.
Work Schedule 1.2.4
1.2.4
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
lab work
field work
Chemostat-Exp.
Milestones (1.2.4)
- Establishing SIP method
- Experimental data set on direct CO2/pH sensitivity (microbial processes)
- Field data for verification of lab data
- Data set on combined effects of CO2 and pressure
- Exchange and compilation of data (for modelling)
month 6
month 18
month 28
month 33
month 33
Subproject 1.2.5 Effect of changing calcareous/lithogenic ballast on aggregates in the
benthic boundary layer
L. Thomsen, G. Gust (Jacobs University)
In collaboration with AWI, Phytoplankton (diatoms, coccoliths) and cyanobacteria will be
cultivated in bioreactors (100 – 5000 l) under different pCO2, temperature, radiation and nutrient
inputs and then transferred into BBL laboratories. Detrital aggregates will be mixed with fine
sediments typical for continental margins and further formed on roller tables (differential settling)
and in shear tanks (turbulent shear). Then the aggregates will be transferred into in benthic flow
simulation chambers and flumes and exposed to different pCO2, temperature, flow and
hydrostatic pressures (0.1 – 20 MPa). The bioavailabity (via HPLC) and mineral composition
93
will be analysed and aggregates classified. One set of experiments will evaluate the importance
of increased CO2 levels and temperature on calcification and ballast formation on carbon
degradation within the BBL with special focus on sorptive preservation. Data will be discussed
with other partners to evaluate the pathway/transportation of aggregates from pelagic to benthic
environments. Phytodetrital aggregates formed under current CO2 levels are compared with those
formed in a future ocean. Another set of experiments will evaluate the importance of increased
pressure on carbon degradation to determine sensitive biogeochemical parameters at atmospheric
and in-situ pressures to understand and model the fate of organic matter in a future ocean. A third
test of experiments will be carried out during spring/summer 2010/2011 to take in situ samples
and expose them to varying pCO2, temperature and hydrodynamic conditions. Background of this
research, the results and discussion will be made available to outreach and training.
Our investigations are linked to the Themes 3 and 5. We will support subproject 3.4.3 to further
understand the importance of the BBL for “carbonate sediment dissolution”, and improve and
better understand the model results of 5.2 "Evaluating and optimising parameterisations of
pelagic calcium carbonate production in global biogeochemical ocean models".
Work Schedule 1.2.5
1.2.5
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
calibration
Production of organisms
CO2 BBL and ballast
CO2 BBL and pressure
Field campaign
Milestones (1.2.5)
- Implementation of experimental facility
- Exp. data set on CO2 & temp. on BBL ballast formation
- Exp. data set on CO2 & pressure on BBL carbon degradation
- In-situ data set on CO2 & pressure on BBL ballast & carbon degradation
month 6
month 18
month 24
month 30
month 33
vi. Budget and Budget Justification
First Year
Personnel costs
1.2.1
1.2.2
1.2.3
1.2.4
94
Second Year
Third Year
Total
First Year
Second Year
Third Year
Total
1.2.5
Subtotal
Consumables
1.2.1
1.2.2
1.2.3
1.2.4
1.2.5
Subtotal
Travel
1.2.1
1.2.2
1.2.3
1.2.4
1.2.5
Subtotal
Investment
1.2.1
1.2.2
1.2.3
1.2.4
1.2.5
Subtotal
Other costs
1.2.5
Total
Personnel costs: 1 Scientist (PhD-candidate; diploma or master required,
): Set-up and
sampling of chemostat experiments at the AWI, and participation in field studies. The Ph.D.
students will examine effects of changes in pCO2 and pH on the release and decomposition of
exopolymer substances, i.e. polysaccharides and amino acids, TEP and CSP, incl. measurements
of bacterial decomposition activities using enzyme assays, bacterial biomass production and
respiration. Analyses of the elemental composition of organic matter will be supported by a
technician at the AWI. Funds for student helpers are to support the set-up and sampling of
chemostat experiments, and to help with cruises preparations, routine lab work and maintenance
of cultures.
Consumables: Consumables are primarily requested for Nuclepore-, membrane- and GF/F
filters, Cytoclear slides for the microscopy of TEP and CSP, chemical reagents, glasware,
nutrient media, ion exchange columns, HPLC columns, isotopes (14C-bicarbonate, 14C Leucine,
3
H Thymidine), fluorogenic substrate analogues, and for Macrosept tubes (10, 100, 1000 kDa) for
ultrafiltration.
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Travel: Travel funds are for exchange visits between partners, and for national meetings and 1
international conference per year. In 2009, funds include travel costs to Svalbard (mesocosm
experiment).
Investments: Investments are needed for a START-1500 MLS microwave, which enables high
throughput sample pre-processing during the hydrolysis and drying steps of CHO and AAS
analyses under defined and reproducible conditions. Investments are also required for O2-optodes
to determine bacterial respiration during the CO2 perturbation experiments, and for a, Shimadzu
TNM-1 expansion of TOC analyzer.
Personnel costs: A PostDoc will work half time on the project to establish the stable isotope
measurements in DON in the IOW where all technical prerequisites are given. This work can be
accomplished by an experienced person within the first 6 months. Batch culture experiments
under 3-5 constant pCO2 levels will be carried out on cooperation with subproject 1.2.3. Cruises
to the Baltic Sea need to be prepared and equipped; furthermore the person will participate in the
common chemostat experiments at the AWI and carry out the analysis as described above. We
aim to find one PostDoc to work the other half time for the subproject 1.2.4. A student is
supposed to help with the preparation of the cruises and routine lab analysis (nutrients, CO2) for
app. 60 hours per week throughout the whole project time.
Consumables: Consumables include isotope tracer substances (15N2 gas, and Na-H13CO3-),
bottles, septa and syringes for the nitrogen fixation and carbon uptake experiments. The isotope
mass spectrometers need high purity compressed gases and for the combustion of filters a number
of reagents.
Travel: The travel funds are for exchange visits to the partner institutes in Bremerhaven, Berlin,
and Kiel and at least one international conference.
Personnel costs: A PhD student will be employed to perform the extensive and ambitious
experiments in the lab at IOW and AWI as well as during planned cruises, to interpret the data
and to summarizes them in PhD thesis. It is aimed to engage a students help throughout the whole
project to conduct routine pCO2, DOC, DON and DOP measurement. These basic parameters like
the stable CO2 conditions in the experiments have to be ensured for this subproject but also for
the cooperating subproject 1.2.2.
Consumables: Consumables include experimental equipment (microwave digestion tubes,
incubation bottles, CO2-gas etc.) and chemicals for the determination of DOC, DOP and their
components, radioactive phosphorus compounds, and CO2 standards and filters.
Travel: Travel funds are for stays in the partner institutes (Bremerhaven, Berlin) for joint
experiments, to meetings and workshops (Kiel), and for participation in international
conferences.
Personnel costs: A PostDoc will work half time on the project to establish the stable isotope
probing method at IGB where all molecular prerequisites are given and to measure stable
96
isotopes at IOW (in collaboration with M. Voss). This work can only be accomplished by an
experienced person within the first 6 months. Batch culture and pressure chamber experiments (in
cooperation with subproject 1.2.5) under 3-5 constant pCO2 levels will be carried out. Cruises to
the Baltic Sea need to be prepared and equipped and the person will be responsible for all
molecular work in the common chemostat experiments at the AWI. We aim to find one PostDoc
to work the other half time for the subproject 1.2.2. A student is supposed to help with the
preparation of the cruises and routine lab analysis (PCR, DGGE) mainly throughout the
chemostat experiment and the cruises.
Consumables: Consumables include molecular supply (such as Primers, Taq-polymerase, DNA
extraction and purification kits, acrylamide gels etc.) but also substrates labelled with either
stable (SIP method) or radioactive isotopes (bacterial production), fluorogenic substrates
(enzymes), fluorochroms (SYBR Gold for DNA labelling and cell counts) and optodes for
respiration measurements. Further it will include consumables for stable isotope analyses at IOW
(see 1.2.2), reaction tubes, pipette tips etc.
Travel: The travel funds are for exchange visits to the partner institutes in Bremerhaven, Berlin,
and Kiel and at least one international conference.
Personnel costs: One PhD student will create organic rich aggregates of different organic and
inorganic compositions and carry out the flume experiments and pressure tank studies under the
guidance of the supervisors and support of the OceanLab technician teams. The student will also
participate in the field campaigns. Student helpers will support these activities and prepare
specific web-modules (animations, class material) for outreach.
Consumables: Consumables are primarily requested for Nuclepore-, membrane- and GF/F
filters, chemical reagents, glassware, nutrients, HPLC columns, and chemicals for mineralogical
analyses.
Travel: Travel funds are for exchange visits with partners, annual meetings and one international
conference.
Investment: One investment during year 1 is needed for three identical benthic chambers with
control electronics for the specific experiments under varying pCO2 levels, fully integratable into
the pressure laboratory electronics system.
Other costs: Other costs for 1.2.5 include the purchase of different, highly concentrated
microalgae species from a commercial vendor (each year) and access (rent) to the mobile DL2
pressure lab of TUHH (0.1 - 50 MPa) with full thermodynamic/hydrodynamic controls in year 2
and year 3. The rental costs include transport, support by technician and access to specific
sampling gear for in-situ studies during the field campaign. As Giselher Gust is retired from TU
HH, these costs are not covered by the TU HH.
97
Project 1.3: Modelling biogeochemical feedbacks of the organic carbon pump
B. Schneider (Universität Kiel)
i. Objectives
The aim of the project is to assess the impact of ocean acidification and global warming on the
marine soft-tissue pump. In particular the cycling of dissolved organic carbon (DOC) will be
investigated, as its production and decay is sensitive to changes in pH and/or temperature and it
strongly interacts with particle flux dynamics. The processes involved are presently not very well
understood, consequently implementations of DOC in global marine carbon cycle models can be
considered as tentative. In collaboration with the experimental projects in Themes 1 and 3 this
modeling sub-project will review current knowledge about DOC and transparent exopolymer
particles (TEP) to be implemented in a global marine biogeochemical model (PISCES).
The vertical transport of particulate and dissolved organic carbon (POC, DOC) in the ocean, the
organic carbon (soft-tissue) pump, is largely responsible for the vertical gradient in the
concentrations of dissolved inorganic carbon (DIC) in the water column (Volk and Hoffert,
1985). Consequently, ocean carbon storage is more effective than it would be without this
mechanism. A change in the organic carbon pump might result in alterations of the partitioning of
carbon between the oceanic and atmospheric carbon reservoir. In current marine biogeochemical
models the organic carbon pump is generally insensitive to changes in atmospheric pCO2.
However, recent observations from mesocosm studies have shown a CO2-stimulated enhanced
carbon drawdown by marine phytoplankton (Riebesell et al., 2007), which is partly transferred
into DOC. Enhanced production of transparent exopolymer particles (TEP), a successor of DOC,
has been documented from earlier mesocosm and laboratory studies (Engel et al., 2004; Engel et
al., 2002). DOC plays a key role for the CO2-exchange between atmosphere and ocean as (1) its
oceanic reservoir is much larger than that of POC, (2) it is sensitive to ocean circulation changes
as its production depends on nutrient supply and export takes place via subduction, and (3) due to
its sticky nature when transformed into TEP it is favoring aggregate formation and may thus
affect particle flux dynamics to either enhanced (Engel et al., 2004) or reduced particle flux
(Mari, 2008). A model-sensitivity study has already shown that a change by one unit in the C:N
ratio of sinking particulate organic matter would have a considerable and persistent effect on the
atmospheric pCO2 (Schneider et al., 2004), whereas global warming is generally expected to
increase DOC degradation by microbial processes (Pomeroy and Wiebe, 2001). The combined
effect of CO2 and temperature on DOC and the resulting alterations in DOC subduction, particle
flux dynamics and air-sea CO2-exchange have not yet been studied.
iii. Previous Work of the Proponent
Birgit Schneider is the head of the Junior Research Group ‘Ocean Circulation and PaleoModeling’ in the framework of the Cluster of Excellence ‘The Future Ocean’ at the University of
Kiel, Germany. The aim of this group is to address the interactions of climate and marine
biogeochemical cycles, in particular with relevance to the global carbon cycle on time scales of
present (Schneider et al., 2008; Schneider et al., 2003), past (Schneider and Schmittner, 2006)
and future (Steinacher et al., in prep.). In collaboration with M. Gehlen and L. Bopp (LSCE,
France) she has already performed model simulations estimating the effect of ocean acidification
98
on calcite forming phytoplankton (Gehlen et al., 2007) and aragonite forming zooplankton
(Gangstø et al., 2008). Furthermore, she has worked on sensitivity studies estimating the
relevance of changes in the vertical flux of particulate organic and inorganic carbon (POC, PIC)
on the uptake and storage of CO2 by the ocean, which have demonstrated the potential for
considerable and persistent effects on the atmospheric CO2 concentration (Schneider et al. 2004;
Schneider et al., 2008b).
iv. Detailed Description of the Project Work Plan
In a first step (six months) an extensive review of what is currently known about the processes
contributing to the production and decomposition of DOC and TEP will be summarized.
Therefore, next to current literature the experience of the collaborators within Themes1 and 3 and
their hypotheses for the new experimental setups will be explored, in particular the dependence
of POC/DOC/TEP production and decay on pH and/or pCO2 (Schulz, 3.1.1; Engel, 1.2.1;
Grossart, 1.2.4; Müller, 1.1.3; Nausch, 1.2.3, and Voß, 1.2.2, temperature (Müller, 1.1.3, Voß,
1.2.2), light and nutrients (Hippler, 1.1.2; Nausch, 1.2.3, Voß, 1.2.2) as well as the role of DOC
and TEP in the formation and fate of marine aggregates (Engel, 1.2.1, Thomsen, 1.2.5; Riebesell,
3.2.1. Examples from paleo analogues will be considered in collaboration with two sub projects
from Theme 3 (Mutterlose, 3.5.2; Meier, 3.5.3). In a second step (one year) the current
implementation of DOC cycling in the marine biogeochemical model PISCES (Aumont and
Bopp, 2006) will be revised. Therefore, together with participants from Themes 1 (see above), 3
(Hoppema, 3.4.3) and 5 (Pätsch, 5.1; Oschlies, 5.2) and with colleagues at LSCE (France) new
parameterizations for DOC cycling will be developed and implemented into PISCES. If
available, these parameterizations should already include new experimental results obtained in
Themes 1 and 3. The major points to be addressed here will be the effects of changes in the
environmental conditions (pCO2, pH, temperature, light, nutrients) on (1) species specific DOC
exudation rates of the (four) plankton functional types represented by the model, (2) DOC
mineralization, and (3) the influence of DOC (TEP) on particle aggregation. For the latter a
higher (lower) abundance of DOC and thus TEP will automatically increase (decrease) the
frequency of collisions between DOC and particles and thereby favor (inhibit) aggregate
formation, however, an increase in the stickiness of organic material with lower pH as observed
by (Engel et al., 2004) is so far not considered by the model. In the following phase (1 year) the
complex interplay of physical and biological responses to anthropogenic CO2 perturbations on
the marine carbon cycle and potential climate feedbacks will be addressed by the application of a
state-of-the-art coupled climate carbon cycle model (KCM - Kiel Climate Model, PISCES). By
driving the biogeochemical model with pre-determined ocean circulation fields for either
constant or changing climate conditions the effects of ocean acidification and global warming
will be assessed separately and in combination. The final step (six months) will be the synthesis
of new DOC and TEP related findings from Theme 1 that will also be relevant for the
synthesizing projects in Theme 5 (Oschlies, 5.2; Quaas, 5.3).
Work Schedule
1.3
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
review sensitivity of DOC and TEP to
changes in environmental conditions
99
1.3
First Year
Second Year
Third Year
analyse implementation of DOC
cycling in the model
develop new parameterizations for
DOC cycling also including TEP
run CO2 scenarios
syntesize DOC and TEP related results
from Theme 1
Milestones (1.3)
- Review of the sensitivity DOC and TEP production and decay to changes in
environmental conditions
- Review of the sensitivity of POC, DOC and TEP turnover processes to hydrostatic
pressure effects
- Modelled climate feedbacks of the organic carbon pump under ocean acidification
and climate change
- Synthesis of new results on the sensitivity of DOC and TEP cycling to ocean
acidification and climate change
month 6
month 18
month 30
month 36
First Year
Second Year
Third Year
Total
Personnel costs
1.3
Subtotal
Consumables
1.3
Subtotal
Travel
1.3
Subtotal
Investments
Subtotal
Other costs
1.3
Total
Budget justification 1.3
Personnel costs: A PhD-student will work in Project 1.3 over the entire 3-year period of Bioacid
(first phase), whereby the initial synthesis work provides optimal conditions to get familiar with
the related science and to establish solid cooperations with the other project partners.
Consumables: As consumables mainly data storage media like DVDs and external hard disks
will be needed, e.g. one year of biogeochemical model (PISCES) output in monthly resolution on
the ORCA2-grid has about 1.5 GB. Running several climate change scenarios over 250 years
(preindustrial-2100) will afford disk space on the order of several terabytes.
100
Travel: Travel funds will be needed to visit national (Bioacid) and international collaborators (in
particular L. Bopp, LSCE, France) and to attend at least one international conference per year.
Other costs: Further costs will appear for a desktop PC including screen as well as for
publication fees. All necessary model simulations can be performed on the NEC-SX8 supercomputer at the University of Kiel without additional costs.
vi. References theme 1
Adler S, Hübener T (2007) Spatial variability of diatom associations in surface lake sediments and its implications for transfer functions. J
Paleolimnol. 37: 573-590.
Ainsworth EA, Long SP (2005) What have we learned from 15 years of free-air CO2 enrichment (FACE)? A meta-analytic review of the
responses of photosynthesis, canopy properties and plant production to rising CO2. New Phytologist 165:351-372
Allen AE, La Roche J, Maheswari U, Lommer M, Schauer N, Lopez PJ, Finazzi G, Fernie AR, Bowler C (2008) The whole-cell response of
the pennate diatom Phaeodactylum tricornutum to iron starvation. Proc Natl Acad Sci USA in press
Allgaier M, Vogt M, Thyrhaug R, Riebesell U, Grossart HP (2008) Coupling of heterotrophic bacteria to phytoplankton bloom development at
different pCO2 levels: a mesocosm study. Biogeosciences, Spec. Issue: Pelagic Ecosystem CO2 Enrichment Studies, 5: S317-359
Allmer J, Naumann B, Markert C, Zhang M, Hippler M (2006) Mass spectrometric genomic data mining: Novel insights into bioenergetic
pathways in Chlamydomonas reinhardtii. Proteomics 6:6207-6220
Armstrong RA, Lee C, Hedges JI, HonjoS, Wakeham SG, (2002) A new, mechanistic model for organic carbon fluxes in the ocean based on
the quantitative association of POC with ballast minerals, Deep Sea Res, 49: 219-236
Arrigo KR (2007) Marine manipulations. Nature 450, 491-492
Aumont O, Bopp L, (2006): Globalizing results from ocean in situ iron fertilization studies, Global Biogeochemical Cycles, 20, GB2017,
doi:10.1029/2005GB002591
Azam F, Hodson RE (1977) Size distribution and activity of marine microheterotrophs. Limnol Oceanogr 22:492–501
Battarbee RW, Juggins S, Gasse F, Anderson NJ, Bennion H, Cameron NG (2000) European Diatom Database (EDDI). An information system
for paleoenvironmental reconstruction. European Climate Science Conference, European Commission, Vienna, Austria 1998, 1-10
Beaulieu, SE, Smith KLJr (1998) Phytodetritus entering the benthic boundary layer and aggregated on the sea floor in the abyssal NE Pacific:
macro- and microscopic composition. Deep-Sea Res. II. 45, 781-815
Behrenfeld MJ, Kolber ZS (1999) Widespread Iron Limitation of Phytoplankton in the South Pacific Ocean. Science 283:840-843.
Biddanda B, Benner R (1997) Carbon, nitrogen, and carbohydrate fluxes during the production of particulate and dissolved organic matter by
marine phytoplankton, Limnol. Oceanogr., 42, 506–518
Biersmith, A, Benner R. (1998)Carbohydrates in phytoplankton and freshly produced dissolved organic matter, Mar. Chem., 63, 131–144
Bigalke N, Rehder G, Gust G Experimental investigation of the behaviour of CO2 droplets rising in seawater under hydrate-forming
conditions. Environmental Science and Technology, in press
Boyd PW, Watson AJ, Law CS, Abraham ER, Trull T, Murdoch R, Bakker DC, Bowie AR, Buesseler KO, Chang H, Charette M, Croot P,
Downing K, Frew R, Gall M, Hadfield M, Hall J, Harvey M, Jameson G, LaRoche J, Liddicoat M, Ling R, Maldonado MT,
McKay RM, Nodder S, Pickmere S, Pridmore R, Rintoul S, Safi K, Sutton P, Strzepek R, Tanneberger K, Turner S, Waite A,
Zeldis J (2000) A mesoscale phytoplankton bloom in the polar Southern Ocean stimulated by iron fertilization. Nature 407:695-702
Buesseler KO, Doney SC, Karl DM, Boyd PW, Caldeira K, Chai F, Coale KH, de Baar HJW, Falkowski PG, Johnson KS, Lampitt RS,
Michaels AF, Naqvi SWA, Smetacek V, Takeda S, Watson AJ (2008) Environment - Ocean iron fertilization - Moving forward in
a sea of uncertainty. Science 319:162-162
Bronk DA, Glibert PM,Ward BB. (1994) Nitrogen Uptake, Dissolved Organic Nitrogen Release, and New Production, Science, 265: 1843 –
1846
Bronk DA, See JH, Bradley P, Killberg L (2007) DON as a source of bioavailable nitrogen for phytoplankton, Biogeosciences: 283-296
Bronk DA,Ward BB (1999) Gross and net nitrogen uptake and DON release in the euphotic zone of Monterey Bay, California, Limnology and
Oceanography, 44(3): 573-585
Burkhardt S, Zondervan I, Riebesell U (1999) Effect of CO2 concentration on C:N:P ratio in marine phytoplankton: A species comparison,
Limnol. Oceanogr., 44,(3): 683-690
Burkhardt, S,Riebesell U, (1997) CO2 availability affects elemental composition (C:N:P) of the marine diatom Skeletonema costatum, Marine
Ecology Progress Series, 155: 67-76.
Cahoon LB (1999) The role of benthic microalgae in neritic ecosystems. Oceanogaphy and Marine Biology: an Annual Review 37:47-86
Canfield DE, Thamdrup B, Kristensen E (2005) Aquatic Geomicrobiology. Elsevier Academic Press, San Diego and London
Capone DG, Subramaniam A, Montoya JP, Voss M, Humborg C, Johansen A, Siefert RL., Carpenter EJ. (1998) An extensive bloom of the N2
fixing Cyanobacterium Trichodesmium erythraeum in the Central Arabian Sea during the spring intermonsoon, Marine Ecology
Progress Series, 172: 281-292
Clarke, A, (2003) Costs and consequences of evolutionary temperature adaptation, Trends Ecol Evol, 18: 573-581.
Clarke AL, Weckström K, Andrén E (2002) Diatom- based transfer functions as a tool to study coastal eutrophication: the MOLTEN project.
International Diatom Symposium Ottawa, Canada 25-31 August, 2002
Clarke AL, Juggins S, Conley DJ (2003) A 150-year reconstruction of the history of coastal eutrophication in Roskilde Fjord, Denmark.
Marine Pollution Bulletin 46: 1615-1618
Collins SL, Bell G (2004) Phenotypic consequences of 1,000 generations of selection at elevated CO2 in a green alga. Nature 431:566-569
Collins S, Sultemeyer D, Bell G, (2006) Changes in C uptake in populations of Chlamydomonas reinhardtii selected at high CO2, Plant, Cell &
Environment, 29,(9): 1812-1819.
101
Delille B, Jerome Harlay, Zondervan I, Jacquet S, Chou L, Wollast R, Bellerby RGJ, Frankignoulle M, Borges AV, Riebesell U, Gattuso J-P
(2005) Response of primary production and calcification to changes of pCO2 during experimental blooms of the coccolithophorid
Emiliania huxleyi. Global Biochemical Cycles 19:doi:10.1029/2004GB002318
Dreßler M, Hübener T (2006) Morphology and ecology of Cyclostephanos delicatus (Genkal) Casper and Scheffler (Bacillariophyceae) in
comparison with C. tholiformis Stoermer, Hakansson and Theriot. Nova Hedwigia 82, 3-4, 409-434
Duffy JE, Stachowicz JJ (2006) Why biodiversity is important to oceanography: potential roles of genetic, species, and trophic diversity in
pelagic ecosystem processes. Mar Ecol Progr Ser 311: 179-189
Engel, A, (2002) Direct relationship between CO2 uptake and transparent exopolymer particles production in natural phytoplankton, Journal of
Plankton Research, 24,(1): 49-53
Engel, A, Delille, B, Jacquet, S, Riebesell, U, Rochelle-Newall, E, Terbrüggen, A, Zondervan, I (2004a) Transparent exopolymer particles and
dissolved organic carbon production by Emiliania huxleyi exposed to different CO2 concentrations: a mesocosm experiment, AME,
34: 93-104
Engel, A, Thoms, S., Riebesell, U., Rochelle-Newall, E.,Zondervan, I. (2004b) Polysaccharide aggregation as a potential sink of marine
dissolved organic carbon, Nature, 428: 929-932
Engel, A, et al. (2005) Testing the direct effect of CO2 concentration on a bloom of the coccolithophorid Emiliania huxleyi in mesocosm
experiments, Limnology Oceanography, 50, 493-507
Engel, A, Schulz, K., Riebesell, U., Bellerby, R., Delille, B., Schartau, M. (2008) Effects of CO2 on particle size distribution and
phytoplankton abundance during a mesocosm bloom experiment (PeECE II), Biogeosciences, 5, 509-521
Engel, A, J. Szlosek, L. Abramson, Z. Liu , Lee C. (in press). Investigating the effect of ballasting by CaCO3 in Emiliania huxleyi: Formation,
settling velocities and physical properties of aggregates. Deep Sea Research II
Eveland AL, McCarty DR, Koch KE (2008) Transcript profiling by 3'-untranslated region sequencing resolves expression of gene families.
Plant Physiol 146:32-44
Fabry, VJ, (2008) Marine Calcifiers in a High-CO2 Ocean, Sciencexpress, science.1157130
Fabry, V. J., Seibel B. A., Richard A. F. , Orr J. C. (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICES
Journal of Marine Science: Journal du Conseil 2008 65(3):414-432; doi:10.1093/icesjms/fsn048
Falkowski, P, Scholes RJ, Boyle EA, Canadell J, Canfield D, Elser J, Gruber N, Hibbard K, Högberg P, Linder S, Mackenzie FT, Moore III B,
Pedersen T, Rosenthal Y, Seitzinger S, Smetacek V,Steffen W, (2000) The Global Carbon Cycle: A Test of Our Kowledge of
Earth as a System, Science, 290: 291-296.
Finney LA, O'Halloran TV (2003) Transition metal speciation in the cell: insights from the chemistry of metal ion receptors. Science 300:931936
Forster RM, Creach V, Sabbe K, Vyverman, Stal LJ (2006) Biodiversity-ecosystem function relationship in microphytobenthic diatoms of the
Westerschelde estuary. Mar Ecol Prog Ser 311: 191-201
Gangstø R. Gehlen M, Schneider B, Bopp L, Aumont O, and Joos F (2008) Towards understanding the role of aragonite in shallow water
CaCO3 dissolution and the response of aragonite production and dissolution to increasing atmospheric CO2. Biogeosciences
Discussions 5: 1655–1687
Gattuso JP, Frankignoulle M, Wollast R (1998) Carbon and carbonate metabolism in coastal aquatic ecosystems. Annual Review of Ecology
and Systematics 29: 405-434
Gehlen M, Gangstø R, Schneider B, Bopp L, Aumont O, Ethe C (2007) The Fate of pelagic CaCO3 production in a high CO2 ocean: A Model
Study. Biogeosciences 4: 505-519
Giroldo, D, Vieira, AAH, Paulsen, B (2003) Relative increase of dexoysugars during microbial degradation of an extracellular polysaccharide
released by a tropical freshwater Thalassiosira sp. (Bacillariophyceae). Journal of Phycology, 39, 1109-1115
Grossart, HP, Levold, F, Allgaier, M, Simon, M, Brinkhoff, T (2005) Marine diatom species harbour distinct bacterial communities, Environ.
Microbiol., 7, 860–873
Grossart HP, Kiørboe, T, Tang, KW, Allgaier, M, Yam, EM, Ploug, H. (2006a) Interactions between marine snow and heterotrophic bacteria:
Aggregate formation, bacterial activities and phylogenetic composition. Aquat. Microb. Ecol. 42:19-26.
Grossart HP, Czub, G, Simon, M (2006b) Specific interactions of planktonic algae and bacteria: Implications for aggregation and organic
matter cycling in the sea. Environ. Microbiol. 8:1074-1084
Grossart HP, Allgaier M, Passow U, Riebesell U, (2006c) Testing the effect of CO2 concentration on dynamics of marine heterotrophic
bacteriaoplankton. Limnology Oceanography. 51:1-11
Grossart HP Simon, M (2007) Interactions of planktonic algae and specific bacteria: Influences on dissolved and particulate organic matter
fluxes. Aquat. Microb. Ecol. 47(2):163-176
Grossart HP, Engel, A, Arnosti, C, DeLaRocha C, Murray A, Passow U (2007) Microbial dynamics in autotrophic and heterotrophic seawater
mesocosms: III Organic matter fluxes. Aquat. Microb. Ecol. 49: 143-156
Grote J, Labrenz M, Pfeiffer B, Jost G, Jürgens K (2007) Quantitative distributions of Epsilonproteobacteria and a Sulfurimonas subgroup in
pelagic redoxclines of the central Baltic Sea. Appl Environ Microbiol 73:7155-7161
Gustavs L, Schumann R, Eggert A, Karsten U (2008) In vivo growth fluorometry: a non-invasive and rapid method to measure growth of
microalgae and cyanobacteria. Aqua. Microb. Ecol. (revision submitted)
Hannig M, Lavik G, Kuypers MMM, Woebken D, Martens-Habbena W, Jürgens K (2007) Shift from denitrification to anammox after inflow
events in the central Baltic Sea. Limnol Oceanogr 52:1336-1345
Hedges, JI, Lee, C, Peterson, ML (2002) The biochemical and elemental compositions of marine plankton: A NMR perspective. Marine
Chemistry 78, 1, 47-63
Hoppe HG, Ullrich S (1999) Profiles of ectoenzymes in the Indian Ocean: Phenomena of phosphatase activity in the mesopelagic zone. Aquat
Microb Ecol 19: 139-148
Houghton JT, Ding Y, Griggs DJ, Noguer M, Linden PJvd, Xiaosu D, (2001) Climate change 2001: the scientific basis. Contribution of
Working Group I to the Third Assessment, Report of the Intergovernmental Panel on Climate Change., Cambridge University
Press, Cambridge. 944 pp.
Hübener T, Dreßler M, Schwarz A, Langner K, Adler S. (2007) Dynamic adjustment of training sets (`moving windows` reconstruction) by
using transfer functions in paleolimnology – a new approach. J. Paleolimnology DOI 10.2007/s10933-007-9145-7
Hutchins DA, Fu, FX, Zhang Y, Warner ME, Feng Y, Portune K, Bernhardt PW, Mulholland MR (2007) CO2 control of Trichodesmium N2
fixation, photosynthesis, growth rates, and elemental ratios: Implications for past, present, and future ocean biogeochemistry,
Limnology and Oceanography, 52,(4): 1293-1304
102
Iglesias-Rodriguez MD, Halloran PR, Rickaby REM, Hall IR, Colmenero-Hidalgo E, Gittins JR, Green DRH, Tyrrell T, Gibbs SJ, von Dassow
P, Rehm E, Armbrust EV, Boessenkool KP (2008) Phytoplankton Calcification in a High-CO2 World. Science 320:336-340
Jickells TD, An ZS, Andersen KK, Baker AR, Bergametti G, Brooks N, Cao JJ, Boyd PW, Duce RA, Hunter KA, Kawahata H, Kubilay N,
laRoche J, Liss PS, Mahowald N, Prospero JM, Ridgwell AJ, Tegen I, Torres R (2005) Global iron connections between desert
dust, ocean biogeochemistry, and climate. Science 308:67-71
Jost G, Zubkov MV, Yakushev E, Labrenz M, Jürgens K (2008) High abundance and dark CO2 fixation of chemolithoautotrophic prokaryotes
in anoxic waters of the Baltic Sea. Limnol Oceanogr 53:14-22
Juggins S (2004) The MOLTON diatom data base and transfer functions. User guide, version 1.0
Karsten U, Schumann R, Rothe S, Jung , Medlin L (2006) Temperature and light requirements for growth of two diatom species
(Bacillariophyceae) isolated from an Arctic macroalga. Polar Biol 29: 476-486
Keil RG, Dickens AF, Arnarson TS, Nunn BL, Devol AH (2004) What is the oxygen exposure time of laterally transported organic matter
along the Washington margin? Mar. Chem. 92, 157-165
Klaas C, Archer DE (2002) Association of sinking organic matter with various types of mineral ballast in the deep sea: Implications for the
rain ratio. Global Biochemical Cycles 16, 1116, doi: 10.1029/2001GB001765
Kleeberg A, Hupfer M, Gust G. (2008) Resuspension experiments on phosphorus entrainment in a lowland river, Spree, NE Germany. Aquatic
Sciences 70, in press
Labrenz M, Jost G, Jürgens K (2007) Distribution of abundant prokaryotic organisms in the water column of the central Baltic Sea with an
oxic-anoxic interface. Aquat Microb Ecol 46:177-190
Laguna R, Romo J, Read BA, Wahlund TM (2001) Induction of Phase Variation Events in the Life Cycle of the Marine Coccolithophorid
Emiliania huxleyi. Appl Environ Microbiol 67:3824–3831
Langer G, Geisen M, Baumann K-H, Kläs J, Riebesell U, Thoms S, Young JR (2006) Species-specific responses of calcifying algae to
changing seawater carbonate chemistry. Geochemistry, Geophysics and Geosystems 7:online early doi:10.1029/2005GC001227
LaRoche J, Boyd PW, McKay RML, Geider RJ (1996) Flavodoxin as an in situ marker for iron stress in phytoplankton. Nature 382:802-805
LaRoche J, McKay RML, Boyd P (1999) Immunological and molecular probes to detect phytoplankton responses to environmental stress in
nature. Hydrobiologia 401:177-198
Levitan O, Rosenberg G, Setlik I, Setlikova E, Grigel J, KLepetar J, Prasil O, Berman-Frank I (2007) Elevated CO2 Enhances nitrogen fixation
and growth in the marine cyanobacterium Trichodesmium, Global Change Biology, 13: 531–538, doi: 10.1111/j.13652486.2006.01314.x.
Løvdal T, Eichner C., Grossart HP, Carbonnel V, Chou L, Thingstad F. (2008). Competition for inorganic and organic forms of nitrogen and
phosphorous between phytoplankton and bacteria during an Emiliania huxleyi spring bloom (PeECE II). Biogeosciences, Spec.
Issue: Pelagic Ecosystem CO2 Enrichment Studies, 5: 371-383
Mari, X. (2008): Does ocean acidification induce an upward flux of marine aggregates? Biogeosciences Discussions, 5:1631-1654
Martin JH, Fitzwater SE (1988) Iron deficiency limits phytoplankton growth in the north-east Pacific subarctic. Nature 331:341-343
McKay RM, Geider RJ, LaRoche J (1997) Physiological and Biochemical Response of the Photosynthetic Apparatus of Two Marine Diatoms
to Fe Stress. Plant Physiol 114:615-622
Mendes PL, Thomsen B, Holscher H, Stigter G, Gust (2007). Pressure effects on the biological degradation of organo- mineral aggregates in
Submarine Canyons. Marine Geology 246, 165-175
Mock T, Samanta MP, Iverson V, Berthiaume C, Robison M, Holtermann K, Durkin C, Bondurant SS, Richmond K, Rodesch M, Kallas T,
Huttlin EL, Cerrina F, Sussman MR, Armbrust EV (2008) Whole-genome expression profiling of the marine diatom Thalassiosira
pseudonana identifies genes involved in silicon bioprocesses. Proc Natl Acad Sci U S A
Moseley JL, Allinger T, Herzog S, Hoerth P, Wehinger E, Merchant S, Hippler M (2002) Adaptation to Fe-deficiency requires remodeling of
the photosynthetic apparatus. Embo J 21:6709-6720
Müller MN, Antia AN, LaRoche J (2008a) Influence of cell cycle phase on calcification in the coccolithophore Emiliania huxleyi. Limnology
and Oceanography 53
Müller MN, Schulz K, Riebesell U (2008b) Influence of high CO2 on three coccolithophore key species over multiple generations. in prep
Naumann B, Busch A, Allmer J, Ostendorf E, Zeller M, Kirchhoff H, Hippler M (2007) Comparative quantitative proteomics to investigate the
remodeling of bioenergetic pathways under iron deficiency in Chlamydomonas reinhardtii. Proteomics 7:3964-3979
Naumann B, Stauber EJ, Busch A, Sommer F, Hippler M (2005) N-terminal processing of Lhca3 Is a key step in remodeling of the
photosystem I-light-harvesting complex under iron deficiency in Chlamydomonas reinhardtii. J Biol Chem 280:20431-20441
Nausch M (1998) Alkaline phosphatase activities and the relationship to inorganic phosphate in the Pomeranian Bight (southern Baltic Sea).
Aquat Microb Ecol 16: 87-94
Nausch M. (2000) Experimental evidence for interactions between bacterial peptidase and alkaline phosphatase activity in the Baltic Sea.
Aquat Ecol 34: 331-343
Nausch M, Nausch G. (2000) Stimulation of peptidase activity in nutrient gradients in the Baltic Sea. Soil Biol Biochem 32: 1973-1983
Nausch M, Nausch G (2004) Bacterial utilization of phosphorus pools after nitrogen and carbon amendment and its relation to alkaline
phosphatase activity. Aquat Microb Ecol 37: 237-245.
Nausch M, Nausch G (2006) Bioavailability of dissolved organic phosphorus in the Baltic Sea. Mar Ecol Prog Ser 321: 9-17
Nausch M, Nausch G (2007) Bioavailable dissolved organic phosphorus and phosphorus use by heterotrophic bacteria. Aquat Biol 1: 151-160
Obernosterer I, Herndl GJ (1995) Phytoplankton extracellular release and bacterial growth: Dependence on the inorganic N:P ratio, Mar. Ecol.
Progr. Ser., 116, 247–257
Orr JC, et al. (2005) Anthropogenic ocean acidification over the twenty-first century and its impact on calcifying organisma. Nature 437: 681686.
Passow U. (2002) Transparent exopolymer particles (TEP) in aquatic environments., Progress in oceanography, 55(3/4), 287-333
Piontek J, Händel N, Engel A (2007) Effects of rising temperature and pCO2 on bacterial degradation processes in marine systems, European
Geosciences Union, General Assembly 2007, Vienna, 15.-20.4.
Piontek J, Händel N, Lunau M, Engel, A. (2007). Investigating the effects of changing CO2 concentration on a natural marine plankton
community using a chemostat system, International Union of Geodesy and Geophysics, General Assembly, Perugia, July 2-13
Pomery LR, Wiebe WJ (2001): Temperature and substrates as interactive limiting factors for marine heterotrophic bacteria, Aquatic Microbial
Ecology, 23(2): 187-204
103
Ramos JBe, Biswas H, Schulz KG, LaRoche J, Riebesell U (2007) Effect of rising atmospheric carbon dioxide on the marine nitrogen fixer
Trichodesmium, Global Biogeochemical Cycles, 21: GB2028, doi:10.1029/2006GB002898
Reusch TBH, Veron A, Preuss C, Weiner J, Wissler L, Beck A, Klages S, Kube M, Reinhardt R, Bornberg-Bauer E (2008) Comparative
Analysis of Expressed Sequence Tag (EST) Libraries in the Seagrass Zostera marina Subjected to Temperature Stress. Marine
Biotechnology online early
Reusch TBH, Wood TE (2007) Molecular Ecology of global change. Molecular Ecology 16:3973–3992
Riebesell U., Bellerby, R., Grossart, H.-P., Thingstad, F.(2008) Mesocosm CO2 perturbation studies: from organism to community level.
Biogeosciences, Spec. Issue: Pelagic Ecosystem CO2 Enrichment Studies (accepted)
Riebesell U (2004) Effects of CO2 enrichment on marine phytoplanktons. Journal of Oceanography 60:719-729
Riebesell U, Schulz KG, Bellerby RGJ, Botros M, Fritsche P, Meyerhofer M, Neill C, Nondal G, Oschlies A, Wohlers J, Zollner E (2007)
Enhanced biological carbon consumption in a high CO2 ocean. Nature 450:545-U510
Riebesell U, Zondervan I, Rost B, Tortell PD, Zeebe RE, Morel FMM (2000) Reduced calcification of marine plankton in response to
increased atmospheric CO2. Nature 407:364-367
Rost B, Riebesell U, Burkhardt S, Sültemeyer D (2003) Carbon acquisition of bloom-forming marine phytoplankton. Limnology and
Oceanography 48:55-67
Rynearson TA, Armbrust EV (2000) DNA fingerprinting reveals extensive genetic diversity in a field population of the centric diatom Ditylum
brightwelli. Limnology and Oceanography 45:1329-1340
Schneider B, Schmittner A (2006) Simulating the impact of the Panamanian seaway closure on ocean circulation, marine productivity and
nutrient cycling. Earth and Planetary Science Letters 246: 367-380
Schneider B, Engel A, Schlitzer R (2004) Effects of depth- and CO2-dependent C:N ratios of particulate organic matter (POM) on the marine
carbon cycle. Global Biogeochemical Cycles 18, doi:10.129/2003GB002184
Schneider B, Bopp L, Gehlen M (2008b): Assessing the sensitivity of modeled air-sea CO2 exchange to the remineralization depth of
particulate organic and inorganic carbon. Global Biogeochemical Cycles: doi: 10.1029/2007GB003100, in press
Schneider B, Bopp L, Gehlen M, Segschneider J, Froelicher T, Joos F, Cadule P, Friedlingstein P, Doney S, Behrenfeld M (2008a) Climateinduced interannual variability of marine primary and export production in three global coupled climate carbon cycle models.
Biogeosciences 5: 597-614
Schneider B, Schlitzer R, Fischer G, Nöthig EM (2003): Depth-dependent elemental compositions of particulate organic matter (POM) in the
ocean. Global Biogeochemical Cycles 17 (2), doi:10.1029/2002GB001871
Schulz KG, Rost B, Burkhardt S, Riebesell U, Thoms S, Wolf-Gladrow DA (2007) The effect of iron availability on the regulation of
inorganic carbon acquisition in the coccolithophore Emiliania huxleyi and the significance of cellular compartmentation for stable
carbon isotope fractionation. Geochim Cosmochim Ac 71:5301-5312
Søndergaard M, Williams PJleB, Cauwet G, Riemann B, Robinson C, Terzic S, Woodward EMS, Worm J (2000) Net accumulation and flux of
dissolved organic carbon and dissolved organic nitrogen in marine plankton communities, Limnology Oceanography, 45, 1097–
1111
Strzepek RF, Harrison PJ (2004) Photosynthetic architecture differs in coastal and oceanic diatoms. Nature 431:689-692
Tanaka T, Thingstad TF, Lødal T, Grossart HP, Larsen A, Schulz KG, Riebesell U. (2007) Availability of phosphate for phytoplankton and
bacteria and of labile organic carbon for bacteria at different pCO2 levels in a mesocosm study. Biogeosciences Discuss 4: 39373936
Taylor GT, Iabichella M, Ho T-Y, Scranton MI, Thunell RC, Muller-Karger F, Varela R (2001) Chemoautotrophy in the redox transition zone
of the Cariaco Basin: A significant midwater source of organic carbon production. Limnol Oceanogr 46:148-163
Tengberg, A,Stahl H, Gust G, Müller U, Arning H, Andersson P, Hall OJ (2004) Intercalibration of benthic flux chambers I. Accuracy of flux
measurements and influence of chamber hydrodynamics. Progress in Oceanography 60), 1-28
Thomsen L, McCave IN (2000) Aggregation processes in the benthic boundary layer at the Celtic Sea Continental Margin. Deep-Sea Res. I,
47: 1389-1404
Thomsen L, Gust G. (2000) Sediment stability and characteristics of resuspended aggregates of the western European continental margin.
Deep-Sea Res. I, 47: 1881-1897
Thomsen L, van Weeringm T, Gust G (2002) Benthic boundary layer characteristics at the Iberian Continental Margin Prog. Oceanogr. 54,
315-329
Thomsen, L (2004) Organic rich aggregates: formation, transport behavior, and biochemical composition. In: Flocculation in Natural and
Engineered Environmental Systems, Droppo, I. et al. (Eds.), 143-154
Volk T. Hoffert, MI (1985): Ocean carbon pumps: Analysis of relative strengths and efficiencies in ocean-driven atmospheric CO2 changes. In:
E.T. Sundquist and W.S. Broecker (eds) The Carbon Cycle and Atmospheric CO2: Natural Variations Archean to Present.
Geophys. Monogr. Ser. 32, pp. 99-110., AGU, Washington, D.C.
Voss M, Bombar D, Loick N, Dippner JW (2006) Riverine influence on nitrogen fixation in the upwelling region off Vietnam, South China
Sea, Geophysical Research Letters, 33,(L07604): doi:10.1029/2005GL025569
Voss M, Croot P, Mills M, Lochte K, Peeken, I (2004) Patterns of Nitrogen Fixation along 10°N in the Tropical Atlantic, Geophysical
Research Letters, 31,(L23S09: doi: 10.1029/204GL020127
Wasmund N, Nausch G, Schneider B, Nagel K, Voss M (2005) Comparison of nitrogen fixation and primary production rates by different
methods: A study in the Baltic Proper, Marine Ecology Progress Series, 297: 23-31
Wasmund N, Voss M, Lochte K (2001) Annual nitrogen fixation patterns in the Baltic Proper, Marine Ecology Progress Series, 214: 1-14
Wegner KM, Kalbe M, Kurtz J, Reusch TBH, Milinski M (2003) Parasite selection for immunogenetic optimality. Science 301:1343
Wegner KM, Kalbe M, Reusch TBH (2007) Innate versus adaptive immunity in sticklebacks: evidence for trade-offs from a selection
experiment. Evolutionary Ecology 21:473-483
Williams PJleB (1995) Evidence for the seasonal accumulation of carbon-rich dissolved organic material, its scale in comparison with changes
in particulate material and the consequential effect on net C/N assimilation ratios. Mar Chem 51:17–29
Wolf-Gladrow DA, Riebesell U, Burkhardt S., Bijma J.(1999). Direct effects of CO2 concentration on growth and isotopic composition of
marine plankton, Tellus series b-chemical and physical meteorology, 51, 461-476.
Wölfel J, Schumann R, Adler S, Hübener T, Karsten U (2007) Diatoms inhabiting a wind flat of the Baltic Sea - species diversity and seasonal
succession. Estuar Coast Shelf Sci 75: 296-307
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11.3: Theme 2: Performance characters: reproduction, growth and
behaviours in animal species
i.
Common Background
Theme 2 will address the contribution of ocean acidification (OA) and hypercapnia (i.e. higher
than normal CO2 concentrations in water, animal blood or tissue) to climate change effects on
marine ecosystems through experimental studies in marine water breathing animals. Initial
findings suggest decreased growth and enhanced mortality of sensitive species, e.g. among
molluscs or echinoderms, in response to a doubling of CO2 levels from pre-industrial to 560 ppm
(Shirayama and Thornton 2005). This value is likely exceeded during this century. Marine
invertebrates are hypothesized to be among the organisms most sensitive to anthropogenic CO2
accumulation, especially those heavily calcified and with a hypometabolic mode of life.
Unravelling the physiological mechanisms responding to OA is required for the development of
cause and effect relationships (Pörtner et al., 2004, 2005). However, ecosystem effects of OA
will not be understood by solely analysing the direct influence of ocean physicochemistry on
individual organisms and species. The CO2 dependent modulation of responses to other changing
abiotic factors, in particular temperature, also have to be included.
Previous analyses of CO2 effects have frequently addressed short-term stress responses or lethal
thresholds. According to recent insight, however, performance decrements occur prior to the
onset of stress and are responsible for climate-induced reductions in population density (Pörtner
and Knust, 2007). Long-term performance and thus fitness is key to survival and success of a
species. CO2 induced limitation may occur firstly through a decrease in the capacity for growth,
reproductive output, or diverse behaviours including the ventilation of burrows or bioturbation
activities or exercise capacities in general. Secondly, considering the known range of
mechanisms affected by CO2, it appears that a shift of acid-base status, including a shift of
extracellular pH, likely reduces functional capacity of affected mechanisms (including
calcification) and the whole organism in due course (see overview in Fig. 2.1). Preliminary data
indicate that a narrowing of thermal windows results and the effect observed suggests a large
sensitivity of the width of thermal windows (Pörtner et al., 2005, Metzger et al., 2007) and
associated temperature dependent zoogeographical ranges to CO2. Physiological key processes
setting sensitivity to CO2 include the regulation of organismal and cellular acid-base and ion
status and their feedback on other processes associated with organismal performance (Pörtner et
al. 2004, 2005). The physiological principles shaping performance are likely also involved in
multi-step processes affecting marine food webs. Here, species-specific responses and
sensitivities cause various species of an ecosystem to be affected differently, resulting in changes
in species interactions, food web structure and associated carbon fluxes.
Physiological mechanisms at molecular to systemic levels are subject to long term modification
during acclimation (within individuals) and adaptation (between generations). This will, at the
whole organism level, cause shifting tolerance ranges and adjustments in performance optima,
capacities and limits. Conversely, limits become effective where species and firstly, their critical
life stages reach their limits of physiological plasticity and also, of acclimation capacity.
Acclimation or evolutionary adaptation implies genetic changes that can be detected using
genomic and transcriptomic tools. Preliminary experiments indicate in fact that long term
acclimation occurs through enhanced gene expression processes of mechanisms involved in ion
and acid-base regulation. However, it is presently unclear how and to what extent this type of
acclimation improves performance. Similarly, evolutionary adaptation may improve resilience
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over the time course of generations. As a proximate indicator of potential evolutionary
adaptation, acclimation capacity to CO2 may reveal heritable variation between populations of the
same species along a latitudinal cline. Differential acclimation may relate to temperature
dependent differences in CO2 exposure due to different CO2 solubilities and associated water
physicochemistry.
Low water pH
and reduced HCO3ion equilibria
Calcification site
calcification
-
and high CO2
-
Na+/H+-exchange etc.
Epithelia (gill, gut, kidney)
H+ Ω
3 Na+
ATPase
2 K+
H+e
Na+
-
H+
membrane
intracellular space
-
-
H+
gene
expression ( + or - )
Heart
HCO3-
Cl-
Na+
Adenosine
accumulation
and release
blood
pigment
extracellular space
Muscle
functional
capacity
-
+
(some groups) Operculum
metabolic equilibria
protein synthesis rate
-
HCO3H+i
Chemosensory
Neurons pHi ↓
H2O
ventilation rate
Brain
CO2
H2O
Tissues
Fig. 2.1: Hypothetical overview of processes and mechanisms potentially affected by CO2 in a generalized water
breathing animal, at different stages in the life cycle, from egg to adult, emphasizing a key role for extracellular pH
(top right) in defining sensitivity to ocean hypercapnia and acidification (after Pörtner et al. 2004, 2008). Some
regulatory factors and processes relevant for growth and behavioural performance are indicated by shaded areas. As in
thermal sensitivity, the first line of hypercapnia tolerance is likely set at the level of functional capacity of the intact
organism and defined e.g. by its capacity for performance. e.g. growth (bottom left). Hypercapnia is expected to cause
a reduction in performance and enhanced sensitivity to thermal extremes (conceptual model for impact of extracellular
pH was adopted from Pörtner 2008; picture sources: www.valdosta.edu, www.imagequest3d.com, B. Niehoff, J.
Pickering, www.fische.kanaren.org, O. Heilmayer, C. Bock & G. Lannig).
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Theme 2 will investigate various measures of performance as early indicators of sensitivity and
also addresses their ecosystem consequences in various climate scenarios. Our goal is to
elaborate the physiological mechanisms setting performance levels in various species and animal
phyla as well as their capacity to shift those mechanisms and associated tolerance thresholds
through acclimation or evolutionary adaptation at transcriptional and translational levels.
Comparative work focuses on those species that are key in the functioning of the respective
ecosystems or of commercial interest. Marine fish and invertebrates with likely different levels of
sensitivity to CO2 and pH will be exposed to conditions simulating climate scenarios as expected
from anthropogenic emission strategies (Caldeira and Wickett, 2003, 2005). Application of more
extreme conditions may be required for an identification and characterization of key
physiological mechanisms responding to CO2, from molecular to organismal levels of biological
organisation.
These efforts explore whether unifying physiological principles are operative across life stages
and are shaping various degrees of sensitivity in both calcifiers and non-calcifiers. Potential
methods and approaches include invasive (optodes and chromatography techniques) and noninvasive (NMR, optical techniques) studies of systemic and cellular acid-base regulation. They
also include analyses of metabolic and calcification rates, of energetics and of correlates or
functional rates of cellular processes (e.g. protein synthesis) associated with energy budget and
growth. Investigations of gene expression capacities using microarrays include those genes
shaping performance levels, in line with the concept of oxygen and capacity limited thermal
tolerance and the assumed key role for acid-base regulation capacity in shaping sensitivity to
CO2. CO2 effects on motor activity (e.g. grazing), oxygen consumption, circulatory and
ventilatory activity, fertilization, fecundity or egg production and hatching, development and
somatic growth will be studied. Temperature dependent acid-base and ion regulation will also be
explored in model species, e.g. through studies of the levels of haemolymph or blood ion and gas
variables (pH, PO2, PCO2, Mg2+, Ca2+, K+ levels, total CO2, succinate).
These efforts will include three projects, according to the functional role of marine fauna. Project
1 will study grazers and filtrators, project 2 will focus on benthic reptant decapod crustaceans and
project 3 will address aspects of ontogeny, allometry and mechanisms shaping performance and
sensitivities in top predators. The latter project will include the development of mechanism-based
modelling.
Objectives
•
•
•
•
•
•
•
•
Identify critical stages in the life cycle (e.g. eggs, larvae) of functionally important marine
organisms based on performance measures as indicators of sensitivity to OA
Analyse physiological mechanisms defining performance levels and sensitivity as well as
potential consequences for behaviours
Estimate acclimation capacity (gene expression capacity) for that mechanism as the
background of physiological plasticity
Compare species from various phyla and habitats with respect to evolutionary constraints
in acclimation or adaptation as well as differential sensitivities
Quantify impact and tolerance thresholds (tipping points)
Identify potential consequences for shifts in species composition of ecosystems and
associated changes in ecosystem functioning and food web structure
Assess interaction between effects of OA and ocean warming
Compare responses and mechanisms in different populations of a species (e.g. in a climate
gradient) reflecting potential for evolutionary adaptation (genetic differences)
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Building on previous experience (e.g. Pörtner et al. 2001, Bailey et al. 2003, Clemmesen et al.
2003, Michaelidis et al. 2005, Caldarone et al. 2006, Metzger et al. 2007) collaborative research
between projects (2.1., 2.2., and 2.3. as well as 3.1.3) will be carried out in populations of fish
species like cod (Gadus morhua) or molluscs like pectinids (Aequipecten opercularis, Chlamys
islandica) or mussels (Mytilus edulis) or crustaceans (Hyas aranaeus, Cancer pagurus) across a
northern hemisphere latitudinal cline, between the North Sea and the Barents Sea. Species
populations from various latitudes may display variable sensitivity to environmental hypercapnia
in relation to CO2 solubility and carbonate saturation, to temperature dependent shifts in steady
state acid-base status and associated decrements in e.g. growth performance (2.1.2, 2.1.3, 2.2.2).
After adequate testing of mechanistic hypotheses in the adults, the respective knowledge will be
transferred to juveniles or larvae (2.2.1; 2.3.1. and 2.3.2) for an investigation of the specific
sensitivity of developmental processes and of the allometry of CO2 effects.
Efforts will build on jointly used infrastructure developments (0.3.1. and 0.3.2.). Joint laboratory
studies will use common CO2 incubations, with levels covering preindustrial values as well as
those expected from IPCC emission scenario values. These values will be those commonly used
throughout the network. They will be combined with temperature scenarios adopting realistic
CO2 temperature combinations according to climate scenarios and local climates, adopting
similar steps of e.g. 3°C including e.g. either 0,3,6,9,12,15 °C etc.
Data from these joint experiments will provide a basis to understand the differences in
sensitivities to OA among various marine organisms of major Metazoan phyla (Mollusca,
Arthropoda, Echinodermata, Chordata (Pisces)). This allows testing of the hypothesis that the
capacity of ion and acid-base regulation in relation to metabolic capacities defines the level of
sensitivity in all life stages. Data will contribute to an understanding of tolerance thresholds
(tipping points), of the relationships of changes in performance and in biocalcification (3.1.3.,
3.1.4.), and to the development of an integrative molecular to ecosystem picture of CO2 effects.
Data will also improve the parameterization of mechanism based physiological, ecological and
biogeochemical models.
Project 2.1: Effects on grazers and filtrators
PI: Thomas Brey
i. Objectives
Project 2.1 tackles two questions:
- How will ocean acidification and temperature rise affect the invertebrate egg fertilisation
process as well as organism fitness and performance in calcifiers and non-calcifiers, and
- How will these effects translate into changes at the ecosystem level?
Eggs that are directly released into seawater – as in our model organism sea urchin - are exposed
unprotected to an acidified environment. Sea urchin eggs are dormant cells until fertilisation and
are activated within seconds via an external messenger, the invading sperm cell. As a
consequence, signal transduction pathways are induced in between seconds for the activation of
metabolic processes needed for rapid synthesis and hardening of the fertilisation membrane to
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avoid lethal polyspermy. Most processes of signal transduction involve ordered sequences of
biochemical reactions inside the cell, which are carried out by enzymes, activated by second
messengers. The second messenger calcium as a common regulator of many of these signal
transduction pathways has been shown to have a key role in the regulation of enzyme activation
and cell growth. Potential consequences of intracellular pH changes are depletion of intracellular
calcium stores and destruction of the acid-base balance. Enzymes working in a narrow pH
window are suspected to respond on intracellular pH changes by deactivation or inhibition.
In isoosmotic animals an increase of seawater CO2 concentration will translate quickly into a
corresponding CO2 concentration in the body fluids. This causes physiological responses at the
cell and the organism level, as the organism intends to maintain pH-homeostasis. Within certain
limits, higher costs of acid-base regulation may be compensated for by metabolic adjustments,
but organism fitness and performance may be reduced seriously beyond those limits, particularly
at elevated temperatures. Basically, OA represents the same problem for both non-calcifying and
calcifying taxa. In the latter, however, the energetics of shell/skeleton formation may be
particularly sensitive to OA, as may be the quality of the product. Therefore we target two
ecologically important taxa here, calanoid copepods and bivalve molluscs, albeit, owing to
differences in size and morphology, with different methodical approaches. In calanoid copepods
the focus is pH-homeostasis, i.e. the organism level, with the aim to create a multi-species
response matrix to OA and temperature rise that may facilitate our understanding of changes at
the food web level. In molluscs, we explore responses at the cellular and the organism level
simultaneously, with the intention to establish a physiological response model that may assist in
explaining OA induced modifications of physical and morphological properties of carbonate
shells/skeletons.
Across species, the rapid formation of a fertilisation membrane after the acrosom reaction with
one sperm is essential to avoid lethal polyspermy. The construction of this membrane occurs
within milliseconds and is induced by the release of calcium (Ca2+) from intracellular stores. The
resulting Ca2+ pools function as second messengers and induce a signal transduction cascade
leading to the initial cell divisions for forming the embryo (Santella et al. 2006). Ocean
acidification and high CO2 concentration are suspected to affect the function of membrane ion
channels, particularly in sodium, calcium and potassium (compare Jonz and Barnes 2007,
Doering and Mcrory 2007) and thus intracellular ion levels. In mammalian cells, Ca2+ levels and
signalling are strongly pH sensitive (Kostyuk et al. 2003, our unpublished results). Hence we
presume drastic effects of OA on the Ca2+ related signal transduction pathways during the
sensitive phase of fertilisation and initiation of embryogenesis. To our knowledge, however, the
impact of acidification on fertilisation success and embryonic development has not yet been
studied. Sea urchins are particularly vulnerable to acidification (Miles et al. 2007), and sea urchin
eggs have served as models for the analysis of fertilisation and development processes. However,
how calcium signalling in oocytes of echinoderms is affected remains to be elucidated.
Calanoid copepods contribute up to 80% to zooplankton biomass (e.g. Longhurst 1985) and are a
key component in pelagic marine ecosystems, linking primary production to higher trophic levels
(e.g. Runge 1988). Herbivorous species can control phytoplankton development (e.g. Bathmann
et al. 1990); as consumers of phyto- and microzooplankton, they have a strong impact on the
microbial loop in the upper water layers (e.g. Turner et al. 1998), and their faecal pellets
contribute to the carbon flow from the surface to deeper water layers (e.g. review by SchnackSchiel & Isla 2005). The role of CO2 for copepod ecophysiology will, however, not only be
109
understood through its direct influence, but even more so, through the modulation of responses to
thermal extremes. A recent study shows that synergistic effects of carbon dioxide and
temperature strongly affect the edible crab, Cancer pagurus, by narrowing its thermal tolerance
window (Metzger et al. 2007). In copepods, to our knowledge, only two studies focussed on the
effect of elevated CO2, indicating that hatching success and/or egg production of calanoid
copepod decrease at concentrations >8,000 ppm CO2 (Acartia spp. Kurihara et al. 2004; Calanus
finmarchicus, Mayor et al. 2007). Knowledge on the direct effects of CO2 on the organism
physiology and on possible synergistic effects with increasing temperature is lacking completely.
Elevated CO2 levels modify biogenic calcification. Most experiments demonstrated a reduction in
calcification and size at elevated pCO2 (Bijma et al. 1998; Leclercq et al. 2000; Riebesell et al.
2000; Michaelidis et al. 2005; Berge et al. 2006; Gazeau et al. 2007), but the opposite trend has
been observed, too (Iglesias-Rodriguez et al. 2008). Shell formation consumes energy through
active ion regulation but its integration into whole animal functioning and energy metabolism is
little understood. OA and associated hypercapnia requires acid-base regulation as extra- and
intracellular pH levels decrease (for review see Pörtner et al. 2005). Three main mechanisms are
involved in this regulatory process: (i) metabolic production and consumption of protons, (ii)
buffering of extra- and intracellular compartments e.g. by dissolution of CaCO3 exoskeletons of
bivalves and other calcifiers, and (iii) active transport of equivalent ion across cell membranes.
Chronic CO2 exposure has been shown to reduce shell growth rate and soft body growth in the
bivalve, Mytilus galloprovincialis combined with suppressed aerobic energy metabolism
(„metabolic depression“, Michaelidis et al. 2005), likely correlating with depressed protein
anabolism as observed in isolated cells (Langenbuch & Pörtner 2003). The shell stabilises body
form and function and protects the animal against predators and environmental forces and agents.
Thus, understanding the temperature-dependent physiological mechanisms that restrict aerobic
performance and thus fitness during hypercapnic conditions as well as its effect on shell
properties are crucial to assess the degree of sensitivity of calcifiers to the future global scenario.
2.1.1:
Angela Köhler is cell biologist and pathologist with emphasis on toxicology and is involved in
the development and implementation of biomarkers (OSPAR, AMAP, HELCOM, MEDPOL,
Viarengo et al. 2007). She is experienced in enzyme and immunocytochemistry, light-,
epifluorescence and confocal microscopy. Her group has 5 years experience in in vitro
fertilisation experiments with sea urchins and in metabolic mapping by enzyme kinetic studies
and immunolocalisation of proteins during fertilisation.
Ulf Bickmeyer is a cell biologist experienced in optical (CCD-imaging, confocal laser scanning
microscopy) and electrophysiological techniques (patch clamp and others) combined with cell
and tissue culture (vertebrate and invertebrate cells). Expertise: Properties and modulation of ion
channels, toxicity and cellular effects of secondary metabolites from marine organisms,
intracellular signal pathways (cAMP and calcium as second messenger, Bickmeyer et al 2007),
pH sensitive dyes.
2.1.2
Barbara Niehoff is an expert on zooplankton ecology and population dynamics. Since 1992, she
combines laboratory experiments with field observations and, to obtain the large picture of the
response of an organism to its environment, she conducts biochemical, histological and stable
110
isotope analyses (Kreibich et al. 2008). Currently, she is the head of the Helmholtz Young
Investigator Group “Trophic interactions in pelagic ecosystems: the role of zooplankton”.
Franz Josef Sartoris is an expert on the physiology of marine invertebrates, including acid-base
regulation, ion- and osmoregulation, energy metabolism and respiration (Metzger et al 2007). He
has broad experience with the implementation of physiological and biochemical methods in
small-sized animals like amphipods, copepods and decapod larvae.
2.1.3
Gisela Lannig is an expert in temperature and stress physiology of marine vertebrates and
invertebrates, in particular in organismic and cellular physiology where she has a broad
methodological spectrum. In the framework of global change her work focuses on the combined
effects of temperature and additional environmental stressors such as pollution or elevated pCO2
on energy metabolism of marine ectotherms (Lannig et al. 2006).
Thomas Brey is an expert for trophic relations and for energy budgeting at the organism,
population and ecosystem level (Heilmayer et al. 2004). He is experienced in the
sclerochronological analysis of biogenic carbonate archives, particularly mollusc shells (Epplé et
al. 2006).
Christian Bock is a biophysicist and expert in developing and applying Nuclear Magnetic
Resonance (NMR) techniques, particularly for in vivo measurements of muscular and circulatory
performance (Bock et al. 2008). His emphasis is the development and application of non-invasive
in vivo techniques (NMR imaging and spectroscopy, optical techniques) in adaptive and
comparative physiology of mainly marine animals.
Olaf Heilmayer is an expert in experimental bivalve ecophysiology with emphasis on pectinid
species. He is a marine biologist with a special interest in the ecology and physiology of marine
shellfish thriving under conditions that set limits to life, namely in the deep-sea and polar context.
His work on latitudinal adaptation mechanisms in pectinid bivalves was a key to understand
biogeographic shifts in commercially important species (Heilmayer et al. 2004).
Ragnhild Asmus is an expert in long term research on complex intertidal ecosystems,
particularly in experimental work at various scales (Asmus & Asmus 2005).
Subproject 2.1.1 Ocean Acidification and Reproduction: Is the beginning of Life in Danger?
PI: Angela Köhler / Ulf Bickmeyer
Work Programme
Sea urchin eggs are an excellent model to study early effects of environmental changes on vital
biological processes during fertilisation and embryogenesis, mechanisms that are present across
all eukaryotic species up to humans. We use Strongylocentrus sp and Psammechinus miliaris for
analysing (i) calcium related signal transduction and enzyme activation during fertilisation
processes, (ii) formation of a fertilisation membrane and (ii) embryogenesis at different CO2
concentrations (280 to 1,900 ppm). Our fertilization studies will complement the studies
proposed by Storch et al., Piatkowski, Melzner, Bleich et al., Tollrian as well as by Wahl
(subprojects 2.2.1, 2.3.1, 3.1.3, 3.1.4, 3.2.2, 4.1.2) on embryonal development..
111
- For exposure experiments gametes are harvested from female adults maintained in the
aquarium. Oocytes will then be exposed to the suggested CO2/pH conditions and in vitro
fertilisation experiments with appropriate replicates will be performed.
- Calcium signalling: Oocyte calcium levels will be measured optically using fluorescent calcium
chelators e.g. Fura 2 by means of CCD imaging and confocal microscopy (see Wertz et al. 2006,
2007). L-type calcium channels seem to be present in oocytes (Tosti & Boni 2004). Calcium
channels have been described to change function with changing pH; how small pH changes affect
calcium channel function in echinoderm oocytes and calcium dependent second messenger
cascades remains to be investigated in detail. Pharmacological tools like thapsigargin,
nitrendipine and brominated pyrrole-imidazole alkaloids (Hassenklöver & Bickmeyer, 2006,
Bickmeyer et al. 2007) are used to investigate calcium entry pathways.
- In vivo imaging of enzyme kinetics during in vitro fertilisation of NADPH producing pathways
and ovoperoxidase activation for metabolic at different CO2/pH and temperature regimes will be
conducted to analyse the influence of pH on Km and Vmax values. (Use of non-destructive
epifluorescence microscopy in a flow chamber for controlled gamete exposition).
- Immunolocalisation. Many of the potential target proteins have been conserved throughout
evolution. Therefore, a whole range of antibodies designed for mammalian studies can be applied
in sea urchins, as we have tested already. Sequencing of the genome of Strongylocentrotus
purpuratus is complete and supports immunocytochemical studies at the light, confocal and
transmission electronmicroscopic levels which are highly informative with respect to amount and
localisation of molecules at work and their changes in relation to environmental factors such as
pH.
Internal cooperation/networking with projects in other themes:
Subproject 2.1.1 is strongly connected, predominantly to projects in theme 2 via methodological
and scientific approaches (exchange of methodological expertise). Collaborative exchange with
Melzner [3.1.3], Bleich [3.1.4] and Wahl [4.1.2] will address CO2 impact on performance at
additional ontogenetic stages. Joint work is planned with Tollrian (3.2.2) in order to analyse CO2
and pH effects on fertilisation processes and embryonal development across species.
External cooperation/networking:
Jacobs University: Prof Klaudia Brix, Prof. Andrea Koschinsky; Stanford University: D. Epel,
A. Hamdoun; Brown University: G. Wessel; Bodega Marine Laboratory: G. Cherr.
Schedule
2.1.1
First Year
I
Collection and maintaining of organisms
Set-up of culturing facility
CO2 perturbation experiment,
Measurements of calcium signals in living
oocytes
Metabolic mapping of enzymes during in
vitro fertilisation
112
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
2.1.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Sample preparation for immunolocalisation
(LM/TEM)
Sample processing and microscopy
(LM/TEM,AFM)
Data analysis, statistical evaluation, data
interpretation
Milestones (2.1.2)
- Measurements of calcium signals in living oocytes
and description of basic oocyte properties completed
- Metabolic mapping of NADPH producing enzymes during in vitro fertilisation
1st set completed
- Sample preparation for immunolocalisation (LM/TEM) of NADPH producing
enzymes/tyrosin crosslinking 1st set completed
- Measurements of calcium signals in living oocytes during CO2 changes
- Metabolic mapping of NADPH producing enzymes during in vitro
fertilisation (CO2-T), 2nd set completed
- Cortical granule reaction to CO2, 2nd set completed
- Electron microscopy of protein localisation and cell pathology
- Measurements of calcium signals in living oocytes, 3rd set
- Data evaluation, manuscript preparation, presentation of results
month 12
month 12
month 12
month 24
month 24
month 24
month 28
month 33
month 24-36
Subproject 2.1.2 The response of zooplankton organisms to elevated CO2 concentrations
PI: Barbara Niehoff / Franz-Josef Sartoris
Work Programme
Our work will focus on three Calanus species, which dominate the zooplankton communities in
Arctic Seas. C. finmarchicus is advected into the Arctic Ocean from the Norwegian Sea while C.
glacialis and C. hyperboreus are endemic (Jaschnov 1970). All three species are adapted to the
environmental conditions prevailing in the respective water masses (Conover and Huntley 1992).
With increasing seawater temperature as climate models predict, the biogeographic boundaries of
these key species will potentially shift (Hirche and Kosobokova 2007), and this will have
consequences for the ecosystem functioning.
Calanus finmarchicus is one of the copepod species studied best. Many aspects of its life cycle
phenomena and their relation to environmental conditions are known (e.g. Marshall and Orr
1955, Hirche 1998, Niehoff 2007), and this provides an ideal basis for studying the physiological
response to CO2 in detail. In order to detect the species-specific responses to elevated CO2 and
increasing temperature (links to 2.1.3, 2.2.1, 3.1.3, 3.1.4), we will also include C. glacialis and C.
hyperboreus. All three species are large enough to perform measurements of acid-base
physiology parameters.
113
Copepods will be collected in the field, either during sea going expeditions or at land based
marine biological stations, e.g. in Kristineberg (Sweden) or Tromsø (Norway), and then
transported to the aquarium facilities at AWI. In incubation experiments (link to 0.3.1), the
animals will be exposed to different CO2 levels and temperatures according to values predicted
from climate models as used within BioAcid. Algal food, i.e. diatoms and flagellates, will always
be supplied in excess in order to avoid effects of food limitation. At start and during experiments,
metabolic activity (citrate synthase, respiration rates measured by means of optical O2
microelectrodes) as well as growth rates in terms of body weight (CN, protein content, dry
weight) will be determined. In female copepods, egg production rates and hatching success will
be studied to elucidate the combined effect of elevated CO2 concentrations and higher
temperature on reproductive success. For studying the acid-base status, the extracellular pH in the
copepods will be measured by optical pH microelectrodes suitable for small sample sizes (<0.5
µl). Also, ion concentrations (e.g. Cl-, SO42-, Na+, K+, NH4+, Mg2+, Ca2+) will be detected by ion
chromatography recently established at AWI by F. Sartoris. Extracellular CO2 status will be
measured either by gas chromatography (total CO2) or by means of CO2 optodes (pCO2), which
are currently in the process of being developed by PRESENS (link to 0.3.2). Combining these
methods is a novel approach, which allows to studying the copepods response to climate change
i.e. CO2 concentration and temperature increase, from different angles, including both
physiological and ecological aspects.
Internal cooperation with projects in other themes
Addressing the question of growth performance and physiological adaptation at different
scenarios of CO2 concentration and temperature, we will closely cooperate with other subprojects focussing on species-specific responses (2.1.3, 2.2.1, 3.1.3, 3.1.4) by means of joint
laboratory studies (0.3.1). Development and testing of CO2 optodes will be performed in close
cooperation with PRESENS (0.3.2). In addition, our data on the immediate impact of CO2
concentration and temperature on copepods will complement the studies of Boersma et al. (4.2.1)
who tackle the effect of elevated CO2 concentrations on food quality and community structure
effects on pelagic food chains.
Schedule
2.1.2
First Year
I
Set up of aquarium facilities and
cultures
Sampling of copepods
Incubation experiments
Sample analysis of copepod
biochemistry and enzyme activities
Evaluation of ecological and
physiological data
Reporting & publication
Theme workshop & conferences
114
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Milestones (2.1.2)
- Aquarium facilities and algal food cultures established
- Copepods sampled for first incubation experiments
- Copepods sampled for second incubation experiments
- First experimental series and sample analyses (Calanus finmarchicus) completed
- Second experimental series and sample analyses completed (C. glacialis and C.
hyperboreus)
- Data analysis and comparison of species completed
- Final BioAcid report
month 6
month 6
month 6
month 12
month 24
month 30
month 36
Subproject 2.1.3 Calcifying macroorganisms in acidifying & warming shallow waters
PI: Gisela Lannig / Thomas Brey / Christian Bock / Olaf Heilmayer / Ragnhild Asmus
Work Programme
- Analysing the physiological response is based on experiments using pectinid bivalves
(Aequipecten opercularis, 38° - 62°N, and Chlamys islandica, 64° - 70°N) as a model organism.
Pectinids grow comparatively fast and can swim, i.e. they can be exercised. Different populations
will be sampled along the latitudinal/ temperature gradient. Animals will be exposed to present
(control, 380 ppm) and future elevated CO2 levels (between 980 and 1,960 ppm) at ambient and
predicted temperature regimes [link to 0.3.1]. After 4-6 weeks metabolic key processes will be
determined in control and experimental animals: At the organism level (in vivo) we determine
oxygen consumption and metabolomics during routine metabolism, after exhausted exercise and
during recovery using flow-through respirometry in the NMR (Bailey et al. 2003), circulatory
capacity (heart rate, hemolymph pO2) using Doppler perfusion system and implantable O2-micro
sensors (Lannig et al. 2008), fitness parameters (contractile performance, lipid and glucose
metabolism, growth and shell structure (see below) and acid-base & ion regulation (changes in
pH and ion content in extrapallial fluid, hemolymph and tissues) using a force gauge/sensor
combination and NMR techniques incl. MAS spectroscopy and chromatography, and
pCO2/pO2/pH sensors (Bock et al. 2001, Brodte et al. 2007, Guderley et al. 2008) [link to 0.3.2,
2.1.2, 2.2.1, 2.2.2, 2.3.2]. Direct incorporation of calcium and CO32- will be measured using
labelling techniques (see Wheeler et al. 1975; Peck et al. 1996). At the cellular/organ level (in
vitro) we determine temperature-dependent changes in energy allocation and estimate capacities
for CaCO3 formation under internal milieu conditions inferred from organism level data and
under changing CO2 and extracellular pH values [link to 2.3.2, 3.1.3, 3.1.4, 3.4.1, 3.5.1]. We will
measure capacities and costs of key processes like ion regulation (Na+/K+-ATPase, Na+ Proton
exchanger, Na+-dependent HCO3-/Cl- co-transporter), protein synthesis, RNA/DNA synthesis,
ATP synthesis (mitochondrial efficiency) in gill, mantle and muscle tissue using CMOS cell
chip-technology (cellular respiration and acidification), NMR/MAS spectroscopy and
chromatography (see Wind et al. 2001, Lehmann et al. 2001, Mark et al. 2005, Bruno et al. 2006,
Cherkasov et al. 2006). Capacities of carbonic anhydrase will be determined in gill and mantle
tissue (Skakks & Henry 2002, Yu et al. 2006).
115
- The ecological effects will be evaluated by a “screening” approach that applies the whole
organism approach to a wide range of taxa (molluscs, barnacles, sea urchins) from the German
Bight. For this approach organisms will be maintained at defined temperatures and control and
elevated CO2 concentrations (see above) for one whole seasonal cycle in mesocosms systems at
the AWI field station Sylt. The selection of taxa will depend to a certain extent on the availability
of the various candidate species in the Sylt/Rønne area that may vary greatly from year to year.
Long-term effects of elevated CO2 levels on carbonate shell/skeleton properties are analyzed by
standardized procedures at levels of macrostructure (mass, density, stability), microstructure (µm
scale such nacre structure in bivalves, REM) and ultra structure (nm scale, crystallite structure,
Atomic Force Microscopy) [link to 3.2.4, 3.3.1, 3.3.2, 3.4.1, 3.5.1]. Short-term experiments will
be used to investigate synergistic effects of CO2 and temperature in early life stages (larvae,
juveniles) of selected species [link to 2.2.1, 2.3.1, 4.1.2]. Organism fitness for all ontogenic
stages is determined by standard measures such as metabolic activity, C, H, N, P content, and
enzyme activities (see above).
- The future scenario will combine our findings with existing evidence and with model
predictions of the marine environment development during the next century. As an example, a
predicted increase in wave action may be relevant for a species in which we find a certain
reduction in shell strength owing to reduced CO32- levels.
Internal cooperation/networking with projects in other themes:
Subproject 2.1.3 is strongly interconnected, predominantly to projects in theme 2 via
methodological approaches and incubation studies. Cooperation with projects in theme 3 will link
our “physiological performance” data to “biomineralization success”: in collaboration with Bijma
[3.3.1], Böttcher [3.4.1] and Immenhauser [3.5.1] we will determine CO2-dependent changes in
shell dissolution rates and structure of our experimental animals. Collaborative exchange with
Melzner [3.1.3], Bleich [3.1.4], Fitzke [3.2.4] and Wahl [4.1.2] will address CO2 impact on
additional aspects of calcification and stress resistance.
External cooperation/networking:
University Bremen, DFG project in SSP 1158 “Sclerochronology & Recent Climate Change” (PI
Heilmayer), EU project in EPOCA (PI Pörtner)
Schedule
2.1.3
First Year
I
Animal collection & culture
Sample processing & measurement organismic level
Sample processing & measurement cellular level
Physiological data evaluation organismic level
Physiological data evaluation - cellular
level
Mesocosm long term experiments
116
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
2.1.3
First Year
Second Year
Third Year
Mesocosm short term experiments &
analysis
Shell properties analysis & data
evaluation
Milestones (2.1.3)
- Pectinid species culture established
- Pectinid exposure & mesocosm experiments started
- Data evaluation at organism level completed
- Shell property analysis completed
- Coupled cellular energy allocation & animal metabolic models
- Development of future scenarios
- Final BIOACID report
month 3
month 6
month 20
month 28
month 30
month 33
month 36
First Year
Second Year
Third Year
Total
Personnel costs
2.1.1
2.1.2
2.1.3
Subtotal
Consumables
2.1.1
2.1.2
2.1.3
Subtotal
Travel
2.1.1
2.1.2
2.1.3
Subtotal
Investments
2.1.1
2.1.2
2.1.3
Subtotal
Other costs
2.1.1
2.1.2
2.1.3
Subtotal
Total
117
2.1.1
Personnel costs:
1 PhD position for 3 years
Consumables:
chemicals, antibodies, dyes, catching trawls
Travel:
€/y each for the PhD and the two PIs
Investment:
n/a
Other costs:
n/a
2.1.2
Personnel costs:
1 PhD position for 3 years
Consumables:
€/y for chemicals including ion concentration and enzyme activity
determination, microelectrodes and aquarium equipment
Travel:
€/y for participation in expeditions, animal transport, BIOACID
meetings and international conferences
Investment:
n/a
Other costs:
n/a
2.1.3
Personnel costs:
1 PhD position for 3 years – analysing the physiological response
1 PhD position for 3 years – ecological effects & future scenario
Consumables:
chemicals incl. NMR consumables, sensors and metabolic chips SC1000,
gas mixtures, aquarium equipment, sample preparation (REM, AFM)
Travel:
Animal transport ( €/transport), BioAcid meetings ( €/person), Int.
Conferences (e.g. Symposium “The Ocean in a High CO2 World”,
€/person), Transfer Sylt-Bremerhaven (ca. /trip)
Investment:
The MAS-NMR (Magic Angle Spinning NMR; €) spectra of intact
tissue samples yield high-resolution “fingerprints” of component
metabolites, while avoiding the risk of decomposition
Other costs:
Publication costs, e.g. American Journal of Physiology ( €)
118
vi. References
Asmus H, Asmus R (2005) Significance of suspension-feeder systems on different spatial scales, The comparative roles of suspension-feeders
in ecosystem : [proceedings of the NATO Advanced Research Workshop on the comparative roles od suspension-feeders in ecosystems,
Nida, Lithuania, 4 - 9 October 2003] / ed. by Richard F. Dame and Sergej Olenin, Dordrecht : Springer, 199-219 (NATO science series :
4, Earth and environmental sciences ;47)
Bailey DM, Peck LS, Bock C, Pörtner HO (2003) High energy phosphate metabolism during exercise and recovery in temperate and Antarctic
scallops: an in vivo 31P-NMR study. Physiol Biochem Zool 76(5):622-633
Bathmann U, Noji T, von Bodungen B (1990) Copepod grazing potential in late winter in the Norwegian Sea – a factor in the control of spring
phytoplankton growth? Mar Ecol Prog Ser 38: 45-51
Berge JA, Bjerkeng B, Pettersen O, Schaanning MT, Oxnevad S (2006) Effects of increased sea water concentrations of CO2 on growth of the
bivalve Mytilus edulis L. Chemosphere 62(4): 681– 687
Bickmeyer U, Grube A, Klings K-W, Köck M (2007) Disturbance of voltage induced cellular calcium entry by marine dimeric and tetrameric
pyrrole-imidazole alkaloids. Toxicon 50: 490-497
Bijma J, Hemleben C, Huber BT, Erlenkeuser H, Kroon D (1998) Experimental determination of the ontogenetic stable isotope variability in
two morphotypes of Globigerinella siphonifera (d'Orbigny). Mar Micropaleo 35:141-160
Bock C, Lurman GJ, Wittig RM, Webber DM, Pörtner HO (2008). Muscle bioenergetics of speeding fish: in vivo 31P-NMR studies in a 4.7T
scanner with an integrated swim tunnel. Conc Magn Reson B, 33B, 62-73
Bock C, Sartoris FJ, Wittig RM, Pörtner HO (2001) Temperature-dependent pH regulation in stenothermal Antarctic and eurythermal
temperate eelpout (Zoarcidae): an in vivo NMR study. Polar Biol 24:869-874
Brodte E, Graeve M, Jacob U, Knust R, Pörtner HO (2007) Temperature-dependent lipid levels and components in polar and temperate eelpout
(Zoarcidae). Fish Physiol Biochem (online) doi: 10.1007/s10695-007-9185-y
Bruno E, Digilio G, Cabella C, de Reggi A, Baroni S, Mainero V, Aime S (2006) Water exchange across the erythrocyte plasma membrane
studied by HR-MAS NMR spectroscopy. Magn Reson Med 56:978–985
Cherkasov AS, Biswas PK, Ridings DM, Ringwood AH, Sokolova IM (2006) Effects of acclimation temperature and cadmium exposure on
cellular energy budgets in the marine mollusk Crassostrea virginica: linking cellular and mitochondrial responses. J Exp Biol 209:12741284
Conover RJ, Huntley M (1991) Copepods in ice-covered seas. Distribution, adaptations to seasonally limited food, metabolism, growth
patterns and life cycle strategies in polar seas. J Mar Sys 2:1-42
Doering CJ, McRory JE (2007) Effects of extracellular pH on neuronal calcium channel activation. Neurosci 146(3):1032-1043
Epplé VE, Brey T, Witbaard R, Kuhnert H, Pätzold J (2006). Sclerochronological records of Arctica islandica from the inner German Bight.
The Holocene 16: 763-769
Gazeau F, Quiblier C, Jansen JM, Gattuso JP, Middelburg JJ, Heip C (2007) Impact of elevated CO2 on shellfish calcification. Geophys Res
Lett 34: L07603
Guderley H, Janssoone X, Nadeau M, Bourgeois M, Cortés HP (2008) Force recordings during escape responses by Placopecten magellanicus
(Gmelin): Seasonal changes in the impact of handling stress. J Exp Mar Biol Ecol doi:10.1016/j.jembe.2007.06.037
Hassenklöver T, Bickmeyer U (2006) The marine secondary metabolites 2,4-dibromophenol and 2,4,6-tribromophenol differentially modulate
voltage dependent ion currents in neuroendocrine (PC12) cells. Aquatic Toxicol 79:384-390
Heilmayer O, Brey T, Pörtner HO (2004) Growth efficiency and temperature in scallops: a comparative analysis of species adapted to different
temperatures. Funct Ecol 18: 641-647
Hirche HJ (1996) Diapause in the marine copepod, Calanus finmarchicus, - a review. Ophelia 44:129-143
Hirche HJ, Kosobokova KN (2007) Distribution of Calanus finmarchicus in the northern North Atlantic and Arctic Ocean - expatriation and
potential colonization. Deep Sea Res II 54: 2729-2747
P, Rehm E, Armbrust EV, Boessenkool KP (2008) Phytoplankton Calcification in a High-CO2 World. Science 320: 336-340
Jaschnov WA (1970) Distribution of Calanus species in the seas of the northern hemisphere. Int Revue ges Hydrobiol 55:197-212
Jonz MG, Barnes S (2007) Proton modulation of ion channels in isolated horizontal cells of the goldfish retina. J Physiol -London 581:529541
Kostyuk P, Potapenko E, Siryk I, Voitenko N, Kostyuk E (2003) Intracellular calcium homoeostasis changes induced in rat spinal cord neurons
by extracelluar acidification. Neurochem Res 10: 1543-1547
Kreibich T, Saborowski R, Hagen W, Niehoff B (2008) Short-term variation of nutritive and metabolic parameters in Temora longicornis
females (Crustacea, Copepoda) as a response to diet shift and starvation. Helgoland Mar Res doi:10.1007/s10152-008-0112-0
Kurihara H, Shimode S, Shirayama Y (2004) Effect of raised CO2 concentration on the egg production rate and early development of two
marine copepods (Acartia steueri and A. erythraea). Mar Pollut Bull 49:721-727
Langenbuch M, Pörtner HO (2003) Energy budget of hepatocytes from Antarctic fish (Pachycara brachycephalum and Lepidonotothen
kempia) a function of ambient CO2: pH-dependent limitations of cellular protein biosynthesis? J Exp Biol 206:3895-3903
Lannig G, Cherkasov AS, Pörtner HO, Bock C, Sokolova IM (2008) Cadmium-dependent oxygen limitation affects temperature tolerance in
eastern oysters (Crassostrea virginica Gmelin). Am J Physiol 249:R1338-R1346
Lannig G, Flores JF, Sokolova IM (2006) Temperature-dependent stress response in oysters, Crassostrea virginica: Pollution reduces
temperature tolerance in oysters. Aquat Toxicol 79: 278-287
Leclercq N, Gattuso JP, Jaubert J (2000) CO2 partial pressure controls the calcification rate of a coral community. Global Change Biol 6(3):
329– 334
Lehmann M, Baumann W, Brischwein M, Gahle HJ, Freund I, Ehret R, Drechsler S, Palzer H, Kleintges M, Sieben U, Wolf B (2001)
Simultaneous measurements of cellular respiration and acidification with a single CMOS ISFET. Biosens Bioelectron 16:195-203
119
Longhurst AR (1985) The structure and evolution of plankton communities. Prog Oceanogr 15: 1-35
Marshall SM, Orr AP (1955) The biology of a marine copepod Calanus finmarchicus Gunnerus. Oliver and Boyd, Edinburgh, pp. 188
Mayor DJ, Matthews C, Cook K, Zuur AF, Hay S (2007) CO2 -induced acidification affects hatching success in Calanus finmarchicus. Mar
Ecol Prog Ser 350: 91-97
Cancer pagurus. J Thermal Biol 32: 144-151
Michaelidis B, Ouzounis C, Paleras A, Pörtner HO (2005) Effects of long-term moderate hypercapnia on acid-base balance and growth rate in
marine mussels (Mytilus galloprovincialis). Mar Ecol Prog Ser 293:109-118
Miles H, Widdicombe S, Spicer JI, Hall-Spencer J (2007) Effects of anthropogenic seawater acidification on acid-base balance in the sea
urchin Psammechinus miliaris. Mar Poll Bull 54: 89-96
Niehoff B (2007) Life history strategies in zooplankton communities: The significance of female gonad morphology and maturation types for
the reproductive biology of marine calanoid copepods. Prog Oceanogr 74: 1-47
Peck LS, Parker AC, Conway LZ, (1996) Strontium labelling of the shell of the Antarctic limpet Nacella concinna (Strebel, 1908). J Moll Stud
62: 315-325
Pörtner HO, Langenbuch M, Michelidis B (2005) Synergistic effects of temperature extremes, hypoxia, and increases in CO2 on marine
animals: From Earth history to global change. J Geophys Res 110: doi:10.1029/2004JC002561
increased atmospheric CO2. Nature 407: 364-367
Runge JA (1988) Should we expect a relationship between primary production and fisheries? The role of copepod dynamics as a filter of
trophic variability. In Boxshall GA,. Schminke HK (eds), Biology of copepods, Proceedings 3. Int. Conf. On Copepoda, Aug 1987,
London. Hydrobiologia 167/168: 61-71
Santella L, Lim D, Moccia F (2006) Calcium and fertilisation: the beginning of life. Trends Biochem Scie 29: 400-408
Schnack-Schiel SB, Isla E (2005) The role of zooplankton in the pelagic-benthic coupling of the Southern Ocean. Scientia Marina 69: 39-55
Skakks HS, Henry RP (2002) Inhibition of carbonic anhydrase in the gills of two euryhaline crabs, Callinectes sapidus and Carcinus maenas,
by heavy metals. Comp Biochem Physiol C 133:605-612
Tosti E, Boni R, (2004) Electrical events during gamete maturation and fertilization in animals and humans. Humane Reproduction Update.
10: 53-65.
Turner J, Hopcroft R, Lincoln J, Huestis C, Tester P, Roff J (1998) Zooplankton feeding ecology: Grazing by marine copepods and
cladocerans upon phytoplankton and cyanobacteria from Kingston Harbour, Jamaica. Mar Ecol Prog Ser 19: 195-208
Viarengo A, Lowe D, Bolognesi C, Fabbri E, Koehler A (2007). The use of biomarker in biomonitoring: A 2-tier approach assessing the level
of pollutant-induced stress syndrome in sentinel organisms., Comp Biochem Physiol Part C 146: 281-300
Wheeler AP, Blackwelder PL, Wilbur KM (1975) Shell Growth in the Scallop Argopecten irradians. I. Isotope Incorporation with Reference
to Diurnal Growth. Biol Bull 148(3): 472-482
Wertz A, Rössler W, Obermayer M, Bickmeyer U (2006) Functional Neuroanatomy of the Rhinophore of Aplysia punctata. Front Zool 3: 6.
Wertz A, Rössler W, Obermayer M, Bickmeyer U (2007) Functional Neuroanatomy of the Rhinophore of Archidoris pseudorargus. Helgol
Mar Res 61: 135-142
Wind RA, Hu JZ, Rommereim DN (2001) High-resolution 1H NMR spectroscopy in organs and tissues using slow magic angle spinning.
Magn Reson Med 46:213–218
Yu Z, Xie L, Lee S, Zhang R (2006) A novel carbonic anhydrase from the mantle of the pearl oyster (Pinctada fucata). Comp Biochem
Physiol B 143:190-194
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Project 2.2: Long-term physiological effects on different life stages of benthic
crustaceans
PI: Felix Mark
i. Objectives
This project will investigate CO2 sensitivity in different life stages using crustaceans as a model
phylum to investigate common principles from the systemic to the molecular level. We intend to
examine whether and how quickly various life stages can adapt to or tolerate increased CO2
levels under exposition to carbon dioxide tensions from today’s up to the predicted extreme
levels of OA. As populations within a latitudinal gradient display different thermal sensitivities,
we will determine whether there is a synergy between CO2 and thermal challenges and what
implication this has for the future spatial distribution of a species. To explore the synergistic
effects of CO2 and temperature on the transcriptome, we will study whether and how relevant
gene clusters and regulatory networks are differentially expressed within and between
populations. We will use population genetics to include effects of genetic differentiation, and
analyse whether populations of different scope for adaptability exist and whether the genetic
variability of populations within a latitudinal gradient affect their physiological tolerance range.
These molecular data will be combined with analyses of systemic ecophysiological parameters
like behavioural and growth performance, acid-base regulation, oxygen delivery and
consumption and calcification. Ultimately, this will enable us to develop an integrative view of
how the necessity to regulate affects the animal´s energy budget.
The above objectives will be investigated in two osmoconforming and calcifying reptant
crustaceans (Anger 2001), that might be especially vulnerable to OA: Hyas araneus and Cancer
pagurus. They are easily accessible, uncomplicated to rear and much is known about their
biology (Anger and Jacobi 1985; Anger 2001; Truchot, 1980; Burnett and Bridges, 1981;
Metzger et al 2007). They occur with great abundance over a wide latitudinal range (H. araneus:
Barents Sea into the English Channel, C. pagurus: Norway to Northern Africa). The retreat in the
Southern North Sea indicates that H. araneus is already affected by global warming whereas C.
pagurus might benefit from the warming North Sea. This commercially exploited species is longlived and reaches sexual maturity at 10 years (Neal and Wilson, 2007). The juveniles spend most
of their time near the intertidal with extreme daily fluctuations of CO2 tensions and temperature
whereas the sexually mature adults inhabit deeper waters with less fluctuating abiotic factors.
Ocean acidification and global warming present - in evolutionary terms - extreme challenges to
environmental adaptation that summate onto the existing necessities of adaptation and ultimately
require evolution of adaptations de novo. For some aquatic and terrestrial animals, an effect of
global warming on the geographical distribution and even the risk of local extinction could be
demonstrated (Parmesan and Yohe 2003, Pörtner & Knust, 2007). Based on seminal data on the
decapod crustacean Maja squinado (Frederich and Pörtner, 2000), thermal tolerance windows of
aquatic organisms have been defined by limits in oxygen supply through ventilation and
circulation (Pörtner 2001, 2002). These limits exert their effects on the growth rate of individual
specimens and the abundance of a population thereby shaping the biogeography of a species.
Ocean acidification in combination with increasing temperatures thus even more adversely
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affects species survival: upper thermal limits have been shown to decrease under hypercapnia in
the decapod crustacean Cancer pagurus (Metzger et al 2007).
It has been assumed that sensitivity to CO2 may be highest in the smaller life stages (Ishimatsu et
al. 2004, 2005), such as eggs or the planktonic larvae, whereas thermal stress is felt especially by
the largest individuals of a species (Pörtner & Knust, 2007), the benthic adult. Decapod
crustaceans with complex life cycles develop through a planktonic larval and a benthic juvenile–
adult phase. The larvae show dramatic growth and morphogenetic changes, which are affected by
variation in environmental factors such as temperature (Anger, 1998) and salinity (Anger et al.,
1998). The Megalopa is the larval stage that might experience the greatest environmental
fluctuations, as it is the transition stage from the pelagic to the benthic mode of life. Among the
adults, brooding females might be especially affected by temperature extremes during cost
intensive brooding care (Fernandez et al., 2000; Brante et al., 2003), the extent of which has an
impact on the performance of the eggs and the hatching larvae.
By influencing the acid-base status of seawater, CO2 severely affects ion and osmoregulation of
higher marine animals and their life stages. This is of special importance in crustaceans, as they
differ with respect to their ability to regulate the extracellular osmolality against the seawater,
which significantly impacts species distribution: whereas osmoregulating species are able to enter
coastal and estuarine environments, osmoconformer are usually limited to full seawater and more
sensitive to varying salinity and are normally found in deeper water. The capability of
osmoregulation can differ between early developmental life-history stages and adults in both
directions (Charmantier 1998). Extracellular osmoregulation is associated with energy
expenditure for active ion transport, which becomes apparent in higher Na+/K+ ATPase capacities
usually found in gills of osmoregulating species like Carcinus maenas, when acclimated to
diluted seawater (Lucu & Flic, 1999; Henry et al., 2002). Thus the capacity of the sodium pump
in branchial epithelia may be a suitable marker to determine limited acid-base regulatory function
in certain species or life stages. In line with this assumption, the hypometabolic deep-sea decapod
crustacean, Chionoecetes tanneri, displays higher sensitivity towards short-term, severe
hypercapnia than the shallow-living Pacific Dungeness crab, Cancer magister, which is able to
restore extracellular pH to normocapnic levels during 24 h hypercapnic challenge (Pane and
Barry, 2007). Similar sensitivities may become especially effective in early life stages when
development of the ion regulatory inventory may be incomplete.
Classic examples of how osmolality, pH, CO2 and oxygen levels act in concert influencing
central aspects of the energy metabolism, can be found in the respiratory pigment of the
crustaceans, haemocyanin. As systemic oxygen supply and thus haemolymph oxygen levels
strongly affect thermal tolerance windows (Frederich and Pörtner, 2000), respiratory protein
function under hypercapnia and elevated temperatures is of central importance. CO2 can affect
haemocyanin function by binding to the respiratory protein itself and by increasing the acid load
of the haemolymph, both effects leading to decreased haemocyanin oxygen binding capacities
(Bohr shift, cf. Mangum, 1980; Bridges, 2001). Decreasing levels of physically dissolved oxygen
at warmer temperatures cause a further reduction in haemolymph oxygen tension. This can in
part be compensated at the molecular level by expression and rearrangement of haemocyanin
subunit composition. The expression of haemocyanin monomers in decapod crustaceans is thus
very variable and influenced by ontogenetic stage (Durstewitz & Terwilliger, 1997, Terwilliger &
Dumler, 2001) and many factors that exert physiological stress, such as pH, temperature,
hypoxia. Recently, even microarray approaches have been developed to analyse the complex
expression of a whole suite of haemocyanin functional units (Terwilliger et al. 2007). Other
molecular mechanisms defining the limits of acclimation to environmental challenges have so far
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been studied mainly by describing effects on single proteins. Genome wide transcriptome studies
with their model-independent, inductive form of analysis contrast with the more conventional
hypothesis-driven gene-by-gene or protein-by-protein form of analysis. Due to advantages in
technologies, genomic studies become more and more attractive in non-model animals, where
essentially no genetic information has been available and which were selected for special
adaptation of their physiology to answer questions in an ecological, environmental or
phylogenetic context (e.g. Fish: Gracey et al., 2001; 2004; frogs: Storey, 2004; turtles: Storey,
2007). Recently, an EST library for the porcelain crab, Petrolisthes cinctipes, has been
established, which serves as a basis for genomic studies in related crustaceans (Stillman et al.
2006). An important advantage of these screening techniques is their ability to identify groups of
proteins/enzymes that can have interrelated functions. Using profiling techniques based on
categorisation of gene function, regulatory pathways and tissue-specific functions become visible
and help to identify functions of so far unknown genes. Furthermore, novel (candidate) genes and
pathways essential in the response to certain stressors can be detected (Gracey and Cossins, 2003;
Cossins et al., 2006) and thus help to identify common molecular principles resulting in
phenotypic plasticity of the whole organism. Since transcriptome studies cannot detect regulation
at consecutive levels such as translation, protein turnover and post-translational modifications,
the complex pattern of transcript responses has to be interpreted within the framework of known
physiology to provide new hypotheses. Those have then to be tested for their functional
significance with regard to common principles in evolutionary adaptation.
During adaptation to their habitats in a climate gradient, marine animals encounter a wide variety
of local conditions that over time lead to differentiation of populations within the distribution
range of a species. Little is known whether genetic differentiation of populations is directly
linked to the scope for physiological adaptability of a given species, but it may be assumed that
population differentiation is key to fine tuning the match between genotype and environment
along climate gradients. Adaptation to local environmental conditions on a molecular basis are
typically identified by comparing expression patterns in populations experiencing known,
different conditions. This indirect inference of the adaptive value of the observed differences in
expression implicitly ignores other factors, most notably gene flow, that also structure the spatial
distribution of alleles within a species. This argument can, however, be reversed in that the
existence of different alleles and/or expression patterns among populations - despite high levels
of genetic exchange - is a strong argument in favour of a stabilizing selection being responsible
for maintaining the observed differences between them (Hemmer-Hansen et al 2008). Hence
evidence, unbiased by colonization history, is needed, which corroborates the adaptive value of
the candidate genes independent from interpretations derived from physiological pathways they
are involved in. This can be obtained by contrasting the spatial distribution of alleles potentially
involved in a response to OA with a baseline derived from selectively neutral microsatellite
markers.
2.2.1
Daniela Storch, systemic ecophysiologist, has been working intensely on the thermal tolerance
of crab larvae in a latitudinal gradient along the Chilean coast in the past years financed by the
Alexander von Humboldt foundation (Storch et al., submitted a,b,c). As PI in a recently approved
DFG funded project, she will investigate the thermal limits of early life history stages and their
relevance for the biodiversity and biogeography of reptant decapod crustaceans. She is interested
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in how behavioural and physiological performance of the different life history stages within the
ontogeny of crustaceans may determine distribution limits.
Christoph Held is head of a working group focussing on population genetics, phylogeny and
phylogeography of marine crustaceans. By interpreting spatio-temporal patterns of molecular
variation they investigate the processes driving diversification and speciation over ecological and
evolutionary timescales (Held, 2000). They developed and successfully applied a novel approach
for isolating microsatellite markers from unknown genomes (Leese & Held, 2008). They have
also co-developed an integrated software environment aimed at facilitating high-throughput
detection and classification of suitable microsatellites including multiplexed primer design (Held
& Leese, 2007).
Magnus Lucassen, molecular biologist, investigates the genetic basis of climate driven evolution
in higher marine animals and aims to identify the underlying networks and signals involved. He
focuses in his work on key processes of the central energy metabolism, in particular ion/pH
regulation, aerobic/anaerobic capacities and oxygen carriers (Lucassen et al., 2006). Realtime
PCR in combination with immunodetection of proteins and functional assays are being used to
characterize key genes from fish, crustaceans and molluscs (Deigweiher et al., 2008). Currently,
he investigates as PI in a DFG funded project the entire transcriptome in cold-adapted versus
eurythermal fish in response to environmental challenges. These genomic approaches in different
marine phyla are being introduced for the detection of unifying and specific regulatory networks
essential for the adaptability to certain abiotic factors and their consequences for whole animal
performance (Eckerle et al., 2008).
Felix Mark, molecular physiologist, has a strong ecophysiological and biochemical background
and has worked in various ecophysiological projects with fish, crustaceans and molluscs
investigating energy metabolism at systemic (Mark et al., 2002), cellular (Mark et al., 2005) and
molecular levels (Mark et al., 2006). As PI in a DFG funded project, he currently investigates
haemocyanin adaptation in polar and temperate cephalopods and is specifically interested in how
molecular structural design relates to physiological function (Melzner et al., 2007).
2.2.2:
The previous work of the group around Christopher Bridges has been extensively directed
towards the study of ecophysiological adaptations in marine organisms in both the intertidal and
sublittoral areas. Crustacean studies have led to the development of numerous techniques for
catheterisation (Burnett and Bridges, 1981) and investigation of both oxygen and carbon dioxide
transport (Bridges et al, 1979) in detail at the whole animal level. The properties of haemocyanin
in terms of the structure and function of respiratory proteins and control of respiratory affinity
have also been studied together with the influence of carbon dioxide (Bridges, 2001, Chausson et
al, 2001). Ongoing studie involve the moulting of rock lobsters and extensive work on moulting
hormones with colleagues in South Africa (Marco and Gade UCT).
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H. araneus is a well-established model species at the AWI for examination of larval
development. C. pagurus is a well-established model species in the Bridges lab in Düsseldorf
with regard to acid-base regulation. The two species will be subject to the same BIOACID
standard pCO2 levels of 380 (present, base line), 700 (2,5 * preindustrial, IPCC ‘business as
usual’ for 2100), 1120 (4* preindustrial) and 1960ppm CO2 (7* preindustrial) and BIOACID
standard temperatures in the range between 6 and 21°C. To ensure standardised experimental
conditions, most long-term acclimations of both species will be carried out in the aquarium
system in Bremerhaven. CO2 incubation of adult animals will take place in the AWI mesocosms,
the hatching and rearing of larvae under CO2 in the recirculated AquaInno Pond-in-Pond systems
[0.3.1: AWI infrastructure development plan]. Common parameters that will be measured in both
subprojects 2.2.1 and 2.2.2 on H. araneus and C. pagurus include:
- oxygen consumption (larval stages, juveniles and adults)
- acid-base parameters (haemolymph pCO2, pH, total alkalinity, CCO2 and buffer titration)
(juveniles and adults)
- ion composition (larval stages, juveniles and adults)
- haemocyanin oxygen affinity and subunit composition (larval stages, juveniles and adults)
Subproject 2.2.1 Hyas araneus: Sensitivity, adaptive capacities and evolutionary
consequences in populations from different latitudes
PI: Daniela Storch / Christoph Held / Magnus Lucassen / Felix Mark
Work Programme
The H. araneus model will look at the mechanisms from systemic (A) to molecular levels (B) on
the background of population differentiation.
(A) System physiology
We will study the combined effect of acidification and temperature on the whole organism across
populations including all life history stages from the brooding female and egg to the larval stages.
Eggs are included since they may be affected in two ways: first, less oxygen supply by the
females because of thermal stress and or higher costs for extra cellular acid-base regulation and,
second, high CO2 sensitivity for the eggs.
(1) Long-term accumulative effects of acidification
Long-term exposure experiments of H. araneus females with and without eggs will be conducted
to determine the costs of parental care at high temperature and different CO2 levels. Oxygen
consumption will be measured to identify differences in the investment of brooding at varying
CO2 levels at high temperature. Haemolymph flow of females and water flow through the embryo
mass, as indicators for oxygen provision to the eggs, will be obtained by using magnetic
resonance imaging techniques [2.1.3]. The loss of embryos, survival/development of the eggs and
the hatching rates from egg masses of long-term CO2 incubated females will be detected. Larval
performance of hatchlings from these differently incubated females will be monitored with regard
to mortality rates during successive larval stages. Haemolymph ion composition in the various
larval stages will be measured [2.1.2] and related to the Na+/K+ ATPase function (see below).
125
Additionally, oxygen-binding characteristics of haemolymph samples from the long time scale
perturbation incubations of H. araneus and C. pagurus (sub-project 2.2.2) will be analysed
spectrophotometrically at AWI and gas diffusion chambers (cf. work programme 2.2.2) in
Düsseldorf. Purified haemocyanin samples of H. araneus and C. pagurus will be further analysed
biochemically by 2D gel electrophoresis and native and SDS PAGE to examine putative changes
in quaternary structure and expression of different isoforms (further characterised at the genetic
level (B)).
(2) Life history stages most vulnerable to acidification in synergy with temperature
To meet this goal, larvae of populations along the latitudinal temperature gradient will be reared
at varying CO2 levels and then acute temperature tolerance windows will be determined in the
varying larval stages by measuring swimming performance, oxygen consumption rate and cardiac
performance.
(B) Molecular physiology and genetic basis of hypercapnic vulnerability
(1) Functional characterisation of ion regulatory capacities
To estimate ion regulatory sensitivities, Na+/K+ ATPase functional capacities and mRNA
expression will be compared in adults of different populations. Due to the small sample size
Na+/K+ ATPase abundance in larval stages will be assessed using antibodies, which have already
been used in several crustaceans (Lucassen et al. unpublished). The data serves as baseline
information as in several other projects [2.3.2; 3.1.3] and will be related to the acid-base
parameters.
(2) Isolation of key genes determining hypercapnic sensitivity/adaptability
For analyses of the transcriptome we will generate a normalized cDNA library for one population
under control conditions to ensure high genome coverage. Using suppression subtractive
hybridization (SSH), further cDNA libraries will be constructed for the identification of
differentially expressed genes within the time course of hypercapnic acclimation. This technique
has been successfully used in our laboratory for copepods, cephalopods and fish. Additionally,
the life stages that turn out to be most sensitive in the part (A), will be tested against the least
sensitive stage. Sequencing of clones from the normalised and the subtractive libraries will be
performed at the AWI, IFM-GEOMAR or service companies using pyrosequencing (normalized
library) and conventional Sanger technologies (subtractive libraries), respectively. Genbank
based assignment of protein function will allow identification of physiological processes
involved in the response to hypercapnia. For unknown genes we expect several overlaps with
other genomic studies [2.3.2; 3.1.3], so that at least the functional framework of the isolated gene
can be attributed.
Based on these approaches, oligonucleotide based micro-arrays will be developed for
representative gene clusters that mirror physiologically important metabolic pathways and
regulatory processes, with special emphasis to the ion regulatory inventory, aerobic/anaerobic
metabolism and oxygen transport system. With this micro-array, the time course of the
expression profile of all spotted genes will be determined under hypercapnia. Hybridisation, data
collection and analyses will be performed at AWI (in coop. with Dr. Uwe John). For promising
candidate genes differential expression will be validated using real-time PCR (ABI7500), and
profiled in detail over the whole time series, for all acclimation conditions and in all populations
126
and life stages. We will thus be able to assess differences between populations (=genetic markers
under selection) and to detect general fluctuations over time within populations.
(3) Population genetics
We will enrich and isolate candidate microsatellite loci from the genome of H. araneus using the
reporter genome protocol (Nolte et al 2005; Held & Leese 2007). The high efficiency of the
protocol allows careful selection of suitably variable loci for the question under study (Leese &
Held, 2008). Contrasting presumably adaptive genetic differences between populations against
selectively neutral microsatellites allows us to gather independent evidence for the adaptive
nature of candidate genes identified using the functional and transcriptome approach. We will
identify genetic evidence for natural selection by calculating pairwise differences for a matrix of
n populations in the standardized F’ST for coding and neutral markers, respectively. Those point
estimates for which the F’ST(coding) significantly exceeds the F’ST(neutral) are taken to be under natural
selection. They will be tested to what degree environmental similarity (temperature) can explain
genetic similarity - regardless of geographic proximity or background genetic similarity
(Hemmer-Hansen et al 2007). These loci can then be considered prime candidate genes for indepth functional and physiological characterisation not only in crustaceans, but also in other
animal phyla [fish: 2.3.2; molluscs: 3.1.3]. A nested clade analysis (Templeton 1998) will enable
us to separate the impact of contemporary gene flow from the effects of events in the
evolutionary history of H. araneus. The variability of microsatellites typically resolves more
recent events (tens of thousand years), a period in which both the range shift in the German Bight
took place presumably in response to ongoing climate change as well as a recolonization of
higher latitudes after the last glaciation of the Northern hemisphere [4.1.2]. Thus, the combined
approach of systemic to molecular approaches on the background of population genetics and
history will allow for meaningful predictions of the future impacts of OA in this model species,
which can be seminal for studies in other phyla.
Schedule
2.2.1
First Year
I
II
Second Year
III
IV
I
II
III
Third Year
IV
I
II
III
IV
Animal collection, establishment of lab
populations
Incubation experiments, tissue/body
fluid sampling, sample preparation
Female performance with and without
eggs (NMR)
Larval
performance
(swimming
activity, oxygen consumption cardiac
parameters)
Collaborative measurements of acidbase parameters, ion composition of
body fluids, enzyme capacities,
Western blots
cDNA library construction/analysis,
sequencing, gene identification, SSH
libraries
Gene expression analysis (micro array)
and validation (real-time-PCR)
Population genetics
127
2.2.1
First Year
Second Year
Third Year
data interpretation
results at conferences, synthesis
Milestones (2.2.1)
- Implementation of incubation experiments, base-line data on acid-base + gas
month 6
transport status
- Data set on female performance, base-line data on larval performance
month 12
- Data sets on activity and expression of ion regulatory proteins
month 15
- Data set on transcriptome
month 21
- Data set on synergistic effects of CO2 and temperature (first population)
month 24
- Data set on population structure
month 27
- Data set on larval performance (second population)
month 29
- Synthesis, evaluation of combined data sets, sensitivities and uncertainties
month 35
Subproject 2.2.2 Cancer pagurus: Chronic and acute responses – Adaptation versus
Tolerance
PI: Christopher Bridges
Work Programme
In the C. pagurus model, three key physiological areas will be examined:
1. Acid-base balance of the whole organism under varying carbon dioxide load (chronic and
acute) together with varying temperature regimes
2. Functionality of both oxygen and carbon dioxide transport at different life stages and the
influence of both temperature and environmental carbon dioxide on this parameter.
3. The influence of acidification, through higher carbon dioxide levels on both the hormonal
control of moulting and at the same time calcium deposition within the carapace.
Initially both juvenile and adult populations will be established together with suture tagging of
individuals for experimental purposes and baseline parameters measured. These will involve
acid-base status of the two populations (adult and juvenile), the gas transport properties of the
haemolymph, their moulting stage, hormonal status and calcium incorporation rates. Acute (short
time scale, 6 hrs) laboratory exposure to graded level(s) taken from the perturbation schemes will
also be investigated if possible, probably using maximum exposure (1960ppm CO2; 7*
preindustrial) only if time is limited. As out-lined above (work programme 2.2.1), long time scale
perturbations experiments will then be commenced using the BIOACID standard levels of 380,
700, 1120 and 1960ppm CO2. These will consist of parallel groups and also some specimens
prepared for serial sampling from the base line groups. Sampling of both the haemolymph and
acute exposure experiments for each perturbation level will be carried out at 6 and 12 month
intervals. Acid-base status and gas transporting properties will then be examined using similar
methodology as shown in subprojects 2.1.3 for bivalve molluscs and 3.1.3 for cephalopods. At
the same time calcium incorporation rates will be studied for each of the groups. During
perturbation incubation, growth and moulting will be regularly monitored by weighing and
collection of exuvia.
128
A further set of synergy experiments will be carried out involving both carbon dioxide and
temperature commencing 6 months after the start of the single parameter perturbations or 12
months if space is limited. This will involve the use of standard carbon dioxide levels and
standard temperature levels of 9, 15 and 21°C. Acid base status will be determined by standard
measurements of haemolymph PCO2, pH, TA, CCO2 and buffer titration of the haemolymph for
perturbated animals (Düsseldorf). CO2 transport properties can at the same time be established
and oxygen transport properties of the haemolymph under various CO2 levels via the “diffusion
chamber” (Bridges et al, 1979) technique (Düsseldorf). These analyses will also be applied to
blood samples of H. araneus from parallel incubation experiments at AWI (cf. work programme
2.2.1). The temperature sensitivity of oxygen transport can then be determined together with any
specific effect of carbon dioxide on oxygen affinity. After the new base line determination from
perturbated organisms these can be acutely exposed to the maximum levels of carbon dioxide
perturbations (1960ppm CO2; 7* preindustrial) and again acid-base status and gas transport
determined. Moulting hormone levels will be measured using standard HPLC methods and
bioassays. For calcium incorporation, radioactive Ca2+ incubations at set carbon dioxide levels
will be measured and at the same time the role of carbonic anhydrase investigated using specific
inhibitors to look at both the cellular and whole animal level (Düsseldorf) similar measurements
will be made in project 2.1.3 for bivalve molluscs.
Data from both matrices of carbon dioxide perturbation experiments and combined temperature
carbon dioxide experiments with then be examined for synergistic effects. After completion of all
sampling and analytical work, a possible scenario for C. pagurus acid-base balance, gas transport
and calcium deposition in the face of long-term changes in environmental pH will be elaborated.
The magnitude of the acute response before and after long-term perturbation will be assessed.
The role of tolerance or adaptation in the physiological adaptation of C. pagurus, H. araneus and
other crustacean species will then be considered for consequences on a larger scale.
Schedule
2.2.2
First Year
I
II
Second Year
III
IV
I
II
III
Third Year
IV
I
II
III
IV
Establishment of juvenile and adult
populations – Parallel and serial groups
ID.-Tagging
Collaborative analysis of base-line
acid-base, gas transport and growth
status
Laboratory
CO2
Perturbations
experiments
Laboratory
Combined
CO2
Temperature Perturbations
Sampling – Serial sampling + Parallel
group
Sample Analysis (Serial-) and acute
test (parallel-group)
Calcium incorporation experiments
(Parallel group)
data interpretation
129
Milestones (2.2.2)
- Establishment of populations and begin perturbation acclimation
- Baseline data on acid-base, gas transport and growth status
- Data set on CO2 perturbations experiments
- Data set on synergy of temperature and CO2 perturbations
- Completion of data evaluation and synthesis of model for crustaceans
month 3
month 6
month 13
month 24
month 33
First Year
Second Year
Third Year
Total
Personnel costs
2.2.1
2.2.2
Subtotal
Consumables
2.2.1
2.2.2
Subtotal
Travel
2.2.1
2.2.2
Subtotal
Investments
2.2.1
2.2.2
Subtotal
Other costs
Subtotal
Total
2.2.1
Personnel costs: personnel cost stated here are for two PhD students over a period of three years
to be paid at the official rate of
. The success and feasibilty of this comprehensive
subproject is dependent on the approval of the two PhD students as they will both be responsible
for the time-consuming animal collection, perturbation experiments, rearing of the larval stages
and maintenance of the adult populations aside from the experimental parts. One of the PhD
projects will have a systemic, the other one a more molecular bias. Both PhD students will be
supervised by all PIs involved, according to needs and expertise. This should lead to the
presentation of two PhD theses in the form of published papers at the end of a period of
approximately 3 years.
Consumables: The annual consumables would involve the standard requirements for the
laboratories: Animal collection and maintenance: ; Systemic physiology: optodes, gases,
chemicals: molecular biology: library construction and sequencing: expression analyses:
130
Travel: Since some of the analyses will be based at the University of Düsseldorf, costs of trips to
Düsseldorf would be included, also collecting trips for juveniles and adults (Helgoland, Millport,
Norway (Bergen, Tromsö) and Spitsbergen). Support of travelling and subsistence is also needed
for attending project meetings and workshops, for presenting project results on national and
international conferences.
Investment: None
Other costs: None
2.2.2
Personnel costs: personnel cost stated here are for one PhD student over a period of three years
to be paid at the official rate of . This person will be responsible for collecting and setting up of
the juvenile and adult populations within the mesocosm system which will lead to the
perturbation experiments over a period of six to 12 months. He/she will be responsible for
sampling at the given six-month intervals and analysing both chronic and acute responses. This
should lead to the presentation of a PhD thesis in the form of published papers at the end of a
period of approximately 3 years.
Consumables: the consumables outlined here are graded over a three-year period with a high
during the second year. The consumables would involve the standard requirement for the
laboratory e.g. gases, chemicals and isotopes, test kits, aquaria, electrodes, spare parts, animal
supply and feeding
Travel: Since most of the perturbation work will based in AWI (Bremerhaven), costs of trips to
the centre would be included, also collecting trips for juveniles and adults (Millport, Roscoff,
Helgoland). Support of travelling and subsistence is also needed for attending project meetings
and workshops, for presenting project results on national and international conferences.
Investment: Most of the necessary equipment is present and we have a well-equipped technical
workshop, which can provide most of our needs. Specialized systems may have to be purchased
for carbon dioxide measurement systems together with acid-base balance determinations and also
an optode system. Again a graded use of these courses is proposed with a high investment in the
first two years.
Other costs: None
vi. References
Anger K (1998) Patterns of growth and chemical composition in decapod crustacean larvae. Invertebr Reprod Dev 33:159– 176
Anger K (2001) The biology of decapod crustaean larvae. Crustacean Issues 14, AA Balkema Publishers, Lisse, 1-417
Anger K, Spivak E, Luppi E (1998) Effects of reduced salinities on development and bioenergetics of early larval shore crab, Carcinus
maenas. J Exp Mar Biol Ecol 220:287– 304
Anger K, Jacobi CC (1985) Respiration and growth of Hyas araneus L. larvae (Decapoda: Majidae) from hatching to metamorphosis. J exp
Mar Biol Ecol 88(3):257-270
Brante A, Fernández M, Eckerle L, Mark FC, Poertner HO, Arntz W (2003) Reproductive investment in the crab Cancer setosus along a
latitudinal cline: egg production, embryo losses and embryo ventilation. Mar Ecol Prog Ser 251:221-232
Bridges, CR (2001) Modulation of haemocyanin oxygen affinity: properties and physiological implications in a changing world. J Exp Biol
204:1021-1032
Bridges CR, Bicudo JEW, Lykkeboe G (1979) Oxygen content measurement in blood containing haemocyanin. Comp Biochem Physiol
62A:457-462
Burnett LE, Bridges CR (1981) The physiological properties and function of ventilatory pauses in the crab Cancer pagurus. J Comp Physiol
145:81-88
Charmantier, G (1998) Ontogeny of osmoregulation in crustaceans: A Review. Inv reprod development 33(2-3):177-190
131
Chausson F, Bridges CR, Sarradin PM, Green BN, Riso R, Caprais JC, Lallier FH (2001) Structural and Functional Properties of Hemo-cyanin
from Cyanagraea praedator, a Deep-Sea Hydrothermal Vent Crab. Proteins : Structure, Function, and Genetics 45:351–359.
Cossins A, Fraser J, Hughes M, Gracey A (2006) Post-genomic approaches to understanding the mechanisms of environmentally induced
phenotypic plasticity. J Exp Biol 209: 328-2336
Deigweiher K, Koschnick N, Pörtner HO, Lucassen M (2008) Adaptation of ion regulatory capacities in gills of marine fish under hypercapnic
acidosis. Submitted to Am J Physiol
Durstewitz G, Terwilliger NB (1997) Developmental Changes in Hemocyanin Expression in the Dungeness Crab, Cancer magister. J Biol
Chem 272:4347-4350
Eckerle LG, Lucassen M, Hirse T and Pörtner HO (2008) Cold induced changes of adenosine levels in common eelpout (Zoarces viviparus): a
role in modulating cytochrome-c-oxidase expression. J Exp Biol 211:1262-1269
Fernández M, Bock C, Pörtner HO (2000) The cost of being a caring mother: the ignored factor in the reproduction of marine invertebrates.
Ecology Letters 3(6):487–494
Frederich M, Pörtner H O (2000) Oxygen Limitation of Thermal Tolerance Defined by Cardiac and Ventilatory Performance in Spider Crab,
Maja squinado. Am J Physiol 279:R1531-R1538
Gracey AY, Troll JV, Somero GN (2001) Hypoxia-induced gene expression profiling in the euryoxic fish Gillichthys mirabilis. Proc Natl Acad
Sci U S A 98:1993-1998
Gracey A, Cossins A (2003) Application of microarray technology in environmental and comparative physiology. Annu Rev Physiol 65:231259
Gracey AY, Fraser EJ, Li W, Fang Y, Taylor RR, Rogers J, Brass A, Cossins AR (2004) Coping with cold: An integrative, multitissue analysis
of the transcriptome of a poikilothermic vertebrate. Proc Natl Acad Sci USA 101:16970-16975
Held C (2000) Phylogeny and biogeography of serolid isopods (Crustacea, Isopoda, Serolidae) and the use of ribosomal expansion segments in
molecular systematics. Mol Phylogen Evol 15(2):165-178
Held C, Leese F (2007) The utility of fast evolving molecular markers for studying speciation in the Antarctic benthos. Polar Biol 30(4):513521
Hemmer-Hansen J, Nielsen EE, Frydenberg J, Loeschcke V (2007) Adaptive divergence in a high gene flow environment: Hsc70 variation in
the European flounder (Platichthys flesus L.). Heredity 99:592-600
Henry RP, Garrelts EE, McCarty MM, Towle DW (2002) Differential induction of branchial carbonic anhydrase and Na+/K+ ATPase activity
in the euryhaline crab, Carcinus maenas, in response to low salinity exposure. J Exp Zool 292:595-603
Hewitt G (2000) The genetic legacy of the quaternary ice ages. Nature 405(6789):907-913
Ishimatsu A, Kikkawa T, Hayashi M, Lee KS, Kita J (2004) Effects of CO2 on marine fish: Larvae and adults. J Oceanogr 60:731–741
Ishimatsu A, Hayashi M, Lee KS (2005) Physiological effects on fishes in a high-CO2 world. J Geophys Res 110:C09S09
Leese F, Held C (2008) Identification and characterization of microsatellites from the Antarctic isopod Ceratoserolis trilobitoides: nuclear
evidence for cryptic species. Conservation Genetics, online first, doi 101007/s10592-007-9491-z
Lucassen M, Koschnick N, Eckerle LG, Pörtner, HO (2006). Mitochondrial mechanisms of cold adaptation in cod (Gadus morhua L.)
populations from different climatic zones. J Exp Biol 209:2462-71
Lucu C, Flik G (1999) Na+-K+-ATPase and Na+/Ca2+ exchange activities in gills of hyperregulating Carcinus maenas. Am J Physiol
276:R490-499
Mangum CP (1980) Respiratory function of the hemocyanins. Amer Zool 20:19-38
Mark FC, Bock C, Pörtner HO (2002). Oxygen limited thermal tolerance in Antarctic fish investigated by magnetic resonance imaging (MRI)
and spectroscopy (31P-MRS), Am J Physiol 283(5):R1254- R1262
Mark FC, Hirse T, Pörtner HO (2005). Thermal variability of cellular energetics in Antarctic fish hepatocytes. Polar Biol 28(11):805-814
Mark FC, Lucassen M, Pörtner, HO (2006). Thermal sensitivity of uncoupling proteins in polar and temperate fish. Comp Biochem Physiol D
1:365-374
Melzner F, Mark FC, Pörtner HO (2007) Role of blood-oxygen transport in thermal tolerance of the cuttlefish, Sepia officinalis. Int Comp Biol
47(4):645-655
Cancer pagurus. J therm Biol 32:144-151
Neal KJ, Wilson E (2007) Cancer pagurus Edible crab. Marine Life Information Network: Biology and Sensitivity Key Information Subprogramme [on-line]. Plymouth: Mar Biol Assoc UK. <http://www.marlin.ac.uk/species/Cancerpagurus.htm>
Nolte AW, Stemshorn KC, Tautz D (2005) Direct cloning of microsatellite loci from Cottus gobio through a simplified enrichment procedure.
Mol Ecol Notes 5:628-636
Pane EF, Barry JP (2007) Extracellular acid-base regulation during short-term hypercapnia is effective in a shallow-water crab, but ineffective
in a deep-sea crab. Mar ecol prog ser 334:1-9
Parmesan C, Yohe G (2003) A globally coherent fingerprint of climate change impacts across natural systems. Nature 421 37-42
Pörtner HO (2001) Climate Change and Temperature-Dependent Biogeography: Oxygen Limitation of Thermal Tolerance in Animals.
Naturwissenschaften 88:137-146
Pörtner HO (2002) Physiological basis of temperature-dependent biogeography: trade-offs in muscle design and performance in polar
ectotherms. J Exp Biol 205:2217-2230
Pörtner HO, Knust R (2007) Climate change affects marine fishes through the oxygen limitation of thermal tolerance. Science 315:95-97
Stillman JH, Teranishi KS, Tagmount A, Lindquist EA, Brokstein PB (2006) Construction and characterization of EST libraries from the
porcelain crab, Petrolisthes cinctipes. Integr Comp Biol 46(6):919–930
Storch D, Santelices P, Barria J, Cabeza K, Pörtner HO, Fernández M (2008) Thermal tolerance of crustacean larvae (Zoea I) in two different
populations of the kelp crab Taliepus dentatus (Milne-Edwards). Submitted to J Exp Biol
Storch D, Fernández M, Navarrete SA, Pörtner HO (2008) The effect of tempertature on the physiology of Megalopae from Taliepus dentatus.
Submitted to Funct Ecol
Storch D, Cabeza K, Fernández M (2008) Larval performance of Taliepus dentatus (Milne-Edwards, 1834): A comparison between
populations. Submitted to Mar Biol
Storey KB (2004) Strategies for exploration of freeze responsive gene expression: advances in vertebrate freeze tolerance. Cryobiology 48:
134-145
132
Storey KB (2007) Anoxia tolerance in turtles: Metabolic regulation and gene expression. Comp Biochem Physiol A 147(2): 263-76
Templeton AR (1998) Nested clade analyses of phylogeographic data: testing hypotheses about gene flow and population history. Mol Ecol
7(4):381-397
Terwilliger NB, Dumler K (2001) Ontogeny of decapod crustacean hemocyanin: effects of temperature and nutrition. J Exp Biol 204:1013–
1020
Terwilliger NB, Ryan M, Phillips M R (2007) Crustacean hemocyanin gene family and microarray studies of expression change during ecophysiological stress. Integr Comp Biol 46: 991-999
Truchot JP (1980) Lactate increases the oxygen affinity of crab hemocyanin. J Exp Zool 214:205-208
133
Project 2.3: Effects on top predators (fishes, cephalopods)
PI: Catriona Clemmesen
i. Objectives
Early developmental stages of fish and cephalopods have been shown to be generally more
susceptible to environmental toxicants than adults (McKim 1977, Ishimatsu et al. 2004) leading
to the hypothesis that these are the life stages most sensitive to changes in oceanic pH and
hypercapnia. At the same time, the allometry of thermal limitation shows that tolerance to
temperature extremes is lowest in the largest specimens of a species (e.g. Pörtner and Knust
2007). Exposure to combined temperature and CO2 scenarios are needed to show how OA and
warming interact to shape fitness windows in the natural environment. This involves testing of
the hypothesis that OA causes a narrowing of thermal windows (Pörtner et al. 2005, Metzger at
al. 2007).
Comparative analyses of the growth performance and biomineralization of calcified structures
(otoliths, statoliths) of these two important groups (fish and cephalopods) will allow the
comparison of effects of changing pH in the environment on processes involved in development,
growth, metabolic reaction, energy budget allocations and calcification responses. These data will
be compared to analyses of the capacity of ion and acid base regulation in order to evaluate their
role in setting tolerance to the CO2 induced acidification of ambient media (Pörtner et al. 2005,
2008). The mechanisms involved are specifically those setting and defending the parameters of
acid-base status in extracellular and then intracellular compartment, which in turn strongly
influence the levels of metabolic and functional performance of cells, tissues and the whole
organism including growth.
In addition to experimental studies a quantitative approach requires integrated mathematical
modelling of the functioning of contributing acid-base exchangers as identified by experimental
molecular and physiological analyses. The complexity of the model can be reduced to a
manageable size by introducing parameters derived from experiments. At the same time the
experimental design can be substantially simplified by the parallel development of a model, since
the model defines exactly the parameters that are required for the comprehension of the
processes. The project will concentrate on molecular and membrane biology functional studies,
on patterns of extracellular and intracellular acid base regulation, on modelling the integrated
functioning of the acid-bas exchangers as well as on acclimation and adaptation and will
evaluate the reaction of the organisms with different performance indicators.
For each species or life stage the conceptual model (Fig. 2.1) outlined above needs testing and
complementation on various scales. Firstly, the mechanisms of acid-base regulation have to be
identified. Long term incubations will reveal their immediate response and their capacity to
acclimate under OA scenarios at various temperatures and depending on pre-adaptation to
various climate regimes. Secondly, the relative contribution of these carriers to cellular, tissue
and whole organism functioning and the level of energy turnover and performance needs to be
explored under these scenarios. Thirdly, the response of the acid-base machinery to ocean
physicochemistry requires mechanism based modelling as a basis for future quantitative
scenarios of CO2 impacts at the whole organism level. As a perspective, process models may be
used in a spatial ocean model, where past, present, and future scenarios are provided by model
simulations of physical and biogeochemical states as reported by IPCC. This would allow
examination of spatio-temporal scales for the tolerance of organisms to CO2 induced
acidification.
134
Emerging knowledge indicates different sensitivities to ocean acidification in various marine
invertebrate groups and marine fishes, with a key role for the capacity of ion and acid-base
regulation in setting tolerance to the CO2 induced acidification of ambient media (Pörtner et al.,
2005, 2008, Fig. 2.1). The mechanisms involved are specifically those setting and defending the
parameters of acid-base status, firstly in extracellular and then intracellular compartments, which
in turn strongly influence the levels of metabolic and functional performance of cells, tissues and
the whole organism during growth, reproductive periods or foraging (Fig. 2.1). The principles of
how setpoints of acid-base regulation influence function and energetics at cellular, tissue or
whole animal level, have been most comprehensively explored in the sipunculid, Sipunculus
nudus, with a role for shifting use of individual acid-base exchangers (e.g. Pörtner et al. 2000).
Data available for an Antarctic zoarcid indicate similar principles in operation in fish
(Langenbuch and Pörtner 2003). However, our understanding of how individual transport
proteins contribute to setting the levels of acid-base parameters and define their response to
environmental change is still in its infancy. Similarly, detailed study is needed to quantify the
feedback of acid-base status on cellular and tissue functioning as well as whole organism
performance.
Changes in whole organism performance are key to an understanding of stress effects at marine
ecosystem level. A clear example is one where temperature extremes cause a loss in species
abundance primarily through a weakening of performance at the limits of the window of oxygen
and capacity limited thermal tolerance (Pörtner and Knust 2007). CO2 likely weakens the
capacity of oxygen supply mechanisms further (Metzger et al. 2007). Sensitivity to CO2 may be
thus highest at the limits of the thermal window of a species and, vice versa, sensitivity to
temperature extremes may rise with increasing ambient CO2 levels (Pörtner et al. 2005, Metzger
et al. 2007).
During early life history the development of mechanisms involved in acid-base regulation may be
a critical stage with respect to maintenance of growth and sensitivity to ocean hypercapnia.
Larval growth is crucial for winter survival. Larval fish and cephalopod growth follow
genetically determined patterns that are modified by environmental conditions including
photoperiod, oxygen, temperature, prey availability and possibly CO2 induced changes in water
physicochemistry. RNA concentration and RNA/DNA ratios in tissues of a wide variety of
organisms have been related to growth rate and feeding condition. This approach appears to work
particularly well for fish and cephalopod larvae that typically grow rapidly in weight mainly
through high rates of protein synthesis during the larval period (Clemmesen 1994, 2003,
Clemmesen & Doan 1996, Melzner et al 2005). Recent research findings indicate reduced
translational activity (measured as a decrease in RNA/DNA ratio) in newly hatched herring
larvae as a response to increased pCO2 during embryonic development (Franke, Clemmesen &
Riebesell in prep.).
Otoliths and statoliths are calciform ear stones of fishes and cephalopods, respectively. They are
already available in early larval stages and involved in spatial orientation and movement, as well
as in the perception of acceleration. They also function as life history recorders and under
“normal environmental conditions” daily increments are added to their ring-like structure, with
the amount of deposited material being dependent on growth and condition of the animals
(Zumholz et al. 2006, 2007). Under stressful conditions like OA, deposition may decrease.
Element incorporation into ear stones is mainly influenced by chemical parameters of the
surrounding water, but it is unknown if and how changes in the pH in the marine environment
will effect formation and daily incorporation of in these internal calcified structures.
135
Inorganic carbonate species (CO2, HCO3-) in different body compartments mirror the response of
acid-base physiology to changes in oceanic pH and hypercapnia. For instance, intracellular pH is
controlled by the velocity of the interconversion of CO2 and HCO3 and the transport rates for
carbonate species or protons across the compartment boundary (e.g. HCO3-/Cl- antiport, CO2
source via respiration, diffusive CO2 efflux). Mathematical modeling of the regulation of
intracellular pH involving CO2 and ion exchange processes between body compartments and the
environment rely on a sound understanding of carbonate chemistry in open buffered systems.
Properties of the equilibrium state of the carbonate system in aqueous solution are well known.
However, it can be shown that in cellular systems, where we have to deal with small
compartments and short time scales the steady state of carbonate chemistry can deviate
appreciably from equilibrium (Thoms et al. 2001, Thoms & Wolf-Gladrow in prep). Hence, for
modeling intracellular pH regulation this chemical non-equilibrium is of particular interest.
Hitherto there is little detailed work on the steady states of an open buffered carbonate system.
The development of analytical and numerical techniques to determine the steady states of the
carbonate system within body compartments is mandatory for the mathematical description of the
acid-base status of the organism under different environmental conditions.
2.3.1.
Uwe Piatkowski is a zooplankton and fish ecologist with broad expertise in macrozooplankton
community studies and in the biology of cephalopods focusing on reproductive and feeding
ecology, on growth patterns, and on distribution and taxonomy. Recent studies investigated the
influence of temperature and salinity on the trace element incorporation into statoliths of
cephalopods and the reproductive adaptation of sepiolid cephalopods to an oceanic lifestyle
(Zumholz et al. 2006, 2007a,b,c). He is a member of the Kiel University cluster “Future Ocean”
and a partner in several EU projects such as EPOCA, the Marie Curie ITN network CalMarO,
and the European network of excellence MarBEF and he has broad experience in
interdisciplinary research.
Catriona Clemmesen’s primary research interest has been studying environmental effects on the
distribution and composition of fish communities, and the biology and importance of early life
history stages of fish. The applicant has extensive knowledge and experience in using
zooplankton and ichthyoplankton studies to interpret changes in the dynamics of marine
ecosystems based on climate change and in biochemical and otolith analysis in relation to growth
performance and animal well-being (Caldarone et al., 2006, Clemmesen, 1994, 1996, Clemmesen
et al., 2003). The proponent is a member of the excellence Cluster “Future Ocean”, partner in the
EU project EPOCA and the Marie Curie ITN network CalMARO and has initiated the
installation of the CO2 manipulation system at IFM-GEOMAR.
2.3.2.
Magnus Lucassen is a molecular biologist with broad experience in the isolation and
characterization of the responses of key genes in marine fishes, crustaceans and mollusks to
temperature, CO2 and oxygen, using quantitative realtime PCR in combination with the
immunodetection of proteins and functional assays. He investigate the genetic bases of climate
driven evolution and responses to environmental challenges in marine animals, by focusing on
key processes in energy metabolism as well as ion- and acid-base regulation, specifically on
136
aerobic/anaerobic pathways, oxygen carriers and transmembrane ion exchangers (Lucassen et al.
2006; Eckerle et al. 2008; Deigweiher et al. 2008). Genomic approaches are currently being
introduced for the detection of unifying and specific regulatory networks essential for the
response to certain abiotic factors and their consequences for whole animal performance.
Ulf Bickmeyer is a cell biologist experienced in optical (CCD-imaging, confocal laser scanning
microscopy) and electrophysiological techniques (patch clamp and others) partly combined with
cell and tissue culture (vertebrate and invertebrate cells). Interests: Properties and modulation of
ion channels, especially calcium channels. Toxicity and cellular effects of secondary metabolites
(natural products) from marine organisms. Intracellular signal pathways using cAMP and calcium
as second messenger. pH sensitive dyes. Chemoreception in marine invertebrates (Bickmeyer et.
al.2002; 2007; Wertz et al. 2006).
Gerrit Lohmann is experienced in large scale modelling as well as the setting up of modelling
tools including statistical analysis of observational and proxy data. His working Group
‘Paleoclimate Dynamics’ at AWI has a long experience in simulating the different components of
the Earth system in various climate stages. In addition, geostatistical methods are developed and
applied for the analysis of environmental and climate data. Emphasis is on the ocean circulation
and data-model comparison. Statistical analysis of paleo-environmental data and direct
simulation of the data are essential for the integrated use of models and data. Models of different
complexity are developed and used, ranging from conceptual, models of intermediate complexity
to full circulation models (Lohmann, 2003; Knorr and Lohmann, 2003; Lohmann et al. 2008).
Hans O. Pörtner has a long standing history in addressing the physiological bases of ecological
processes and ecosystem functioning, particularly the roles of climatic factors like temperature,
CO2 and oxygen in animal evolutionary history and ecology as well as biogeography. Together
with his division he elaborated the concept of oxygen and capacity limited thermal tolerance
across animal phyla, as well as its ecosystem implications. He is also an expert in quantitative
studies of acid-base regulation. Current interests cover the interaction between climatic factors,
the mechanisms shaping cellular and whole-animal energy budgets under various thermal and
carbon-dioxide regimes, and the molecular mechanisms of environmental adaptation and
limitation (Pörtner et al. 1998, 2000, 2005; Pörtner and Knust, 2007; Metzger et al., 2007;
Pörtner, 2008). The proponent has coordinated the EU project CLICOFI and is currently a work
package leader in the EU project EPOCA.
Silke Thoms is a theoretical physicist with broad experience in interdisciplinary work and in
modelling of biochemical processes involved in NADPH/ATP generation and carbon acquisition
in (photoautotrophic) marine organisms. She has introduced the use of mathematical models to
understand the role of spatial compartmentalization of chloroplasts for the carbon concentrating
mechanism (CCM) in eukaryotic algal cells and to simulate phytoplankton growth under different
light and atmospheric CO2 conditions. Currently, she is principle investigator of a DFG-project
that has the aim to contribute to a better understanding of the regulation of elemental fluxes
(carbon (C), nitrogen (N), divalent metal-ions (Ca2+, Mg2+, Sr2+) on the level of individual (algal)
cells. In the future, we are developing improved parametrizations of biochemical cellular
processes as a platform from which a higher - level ecosystem simulation model can be
constructed (Thoms et al 2001; Engel et al. 2004; Kroons and Thoms, 2006).
137
Subproject 2.3.1: Effects of changes in ocean pH on the development, growth, metabolism
and otolith/statolith formation and composition of fish and cephalopod early life stages: a
comparative approach
PI: Uwe Piatkowski / Catriona Clemmesen
Work Programme
Perform laboratory and mesocosm experiments with marine fish (herring, cod and gobiids) and
cephalopod (European cuttlefish, long-finned squid) eggs, larvae and juveniles at different pH
levels in the range of expected changes from climate forecasting models (0.3.1). A CO2
manipulation system has just been installed at IFM-GEOMAR and will allow for controlled and
stable CO2 concentrations to be applied, which will enable to use the same pCO2 scenarios e.g.
present (380), 2x pre-industrial (560), 2.5x pre-industrial, business as usual scenario for 2100
(700), 3.5x pre-industrial (980) and 5x pre-industrial (1400) and temperature scenarios according
to local climate. It is postulated that gill chloride cells are involved in pH regulation in marine
species and exceptionally CO2 tolerant larval fish have been shown to have a high density of
chloride cells (Ishimatsu et al., 2004). Therefore histological studies on the development and
abundance of these gill cells in juvenile fish in relation to pH changes will be performed (in
cooperation with EPOCA workpackage 6).
Examine synergistic effects of changes in temperature, oxygen and pH on development, growth
and survival of larval and juvenile fish and cephalopods, with the temperature tolerance being
expected to decrease under lower pH levels. Detailed studies:
•
Measure fertilization rate, egg development (2.1.1, 4.1.2), changes in the morphology of
fish/cephalopod eggs and embryos, size at hatch, size of the otolith/statolith at hatch and
during larval development.
•
Determine RNA/DNA ratios as a biochemical indicator for protein metabolism in early fish
and cephalopods (larvae and juveniles) and determine which stages are most susceptible to
pH changes (3.1.4, 2.3.2)
•
Determine the effects of different pH levels on the formation and elemental composition of
otoliths and statoliths in larval and juvenile fishes and cephalopods (3.3.1).
•
Study short- and long-term effects of low pH on larval fish and cephalopods to define the
vulnerability (2.1.3, 2.3.2, 5.3).
Elementary composition of statoliths and otoliths will be determined by using modern Laser
Ablation techniques (LA-ICP-MS) available in IFM-GEOMAR Research division Marine
Biogeochemisty in cooperation with Prof. Eisenhauer within the Marie Curie ITN Network
CalMarO.
Internal cooperation: 0.3.1, 0.3.2, 0.4, 2.1.1, 2.1.3, 2.3.2, 3.1.3, 3.1.4, 3.3.1, 4.1.2, 5.3
138
Schedule
2.3.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
calibration
Collection and rearing of organisms
CO2 perturbation experiment with fish
& cephalopods
Combined CO2/temperature
perturbation experiments with fish &
cephalopods
data interpretation
Milestones (2.3.1)
- Experimental data set on CO2/pH sensitivity on development, growth
and metabolism of fish & cephalopods
- Statolith and otolith composition of cephalopods and fish
- Data set on synergistic effects of CO2 and temperature
of fish and cephalopods
- Evaluation and comparison of combined data sets on
fish & cephalopods
- Completion of work, submission of manuscripts
month 6
month 18
month 24
month 30
month 33
month 36
Subproject 2.3.2 Mechanisms setting and compensating for animal sensitivity to ocean
acidification: functional capacities, thermal interactions and mechanism-based modelling
PI: Magnus Lucassen / Ulf Bickmeyer / Gerrit Lohmann / Hans O. Pörtner / Silke Thoms
Work Programme
The project will focus on teleost fish and their life stages from various climates, specifically
Atlantic cod (Gadus morhua). Comparative data will be elaborated for an elasmobranch, e.g.
Scyliorhinus canicula. For some principle questions and as a preliminary test of unifying
principles in acid-base metabolism across phyla, data should be contrasted to those available for
animal models characterized by incomplete acid-base compensation, calcifiers like Mytilus edulis
or non-calcifiers like S. nudus. Accordingly, the project will coordinate with others with respect
to such comparative analyses and the principle investigations of adult and juvenile life stages
[0.3.1, 2.1.1, 2.1.3, 2.2.1, 2.2.2, 2.3.1, 3.1.3, Coordination with EPOCA, work package 6].
Experiments will use the BIOACID standard levels of 380, 700, 1120 and 1960ppm CO2 at
variable temperatures considering the thermal windows of the species analysed.
139
Central hypotheses driving the comparative approach:
• Marine animals display various sensitivities to elevated CO2 levels according to the
degrees of acid-base compensation in association with their phylogenetic constraints,
metabolic capacities, life history stage, and the capacities of contributing transporters,
as influenced by thermal acclimation and adaptation including the thermal influence on
membrane structure.
• Fish are less sensitive to ocean acidification than marine invertebrates due to a
quantum leap in acid-base regulation capacity and efficiency. Early life stages are more
sensitive than adults especially during the transition phase to adult physiologies. Adults
may become more sensitive than juveniles at the edges of their thermal windows.
• There is a climate dependent setting of thermal windows which has its bearing on the
capacity and efficiency of acid-base regulation and thus, sensitivity to elevated CO2
levels is higher at the edge of thermal windows; vice versa, thermal sensitivity is higher
at elevated CO2 levels.
A combination of molecular, physiological and modelling approaches will be applied in order to
test these hypotheses under controlled conditions of water physicochemistry and temperature.
Responses of larvae and adults will be titrated at various CO2 levels comprising those expected
from IPCC emission scenarios. Detailed studies include:
Studies of molecular and membrane biology coordinated with functional assays
• identification of transporters according to sequence information and inhibitor effects:
The presently about 150,000 ESTs published from G. morhua transcriptome projects
will serve as a basis for identification and characterisation of transport proteins.
Subtractive libraries (SSH) [2.2.1, 3.1.3] and determination of full-length sequences
(RACE; see Mark et al. 2006, Deigweiher et al. 2008) will focus on gill transporters.
• assessment of pH dependent gene expression capacity and protein concentrations of
individual transporters using real-time PCR: The corresponding protein concentration
will be monitored with available antibodies specifically developed for cod. Labelling
the transporter protein with specific inhibitors will be another method of choice [2.2.1,
3.1.3].
• analyses of the interaction of pH and temperature effects on gene expression:
Transporters identified as essential for acclimation due to their mode of regulation will
be further characterised through functional studies. After determination of the fulllength sequence, the gene will be cloned into an appropriate vector system for
heterologous expression and functional assays applying optical and
electrophysiological techniques [3.1.4].
• assay of membrane structures (changes in lipid composition) and functional
consequences for ion and acid-base regulation depending on thermal acclimation and
adaptation (climate regime)
Functional studies, coordinated with molecular work
• whole organism patterns of extra- and intracellular acid-base regulation at thermal
optima as well as at low and high thermal extremes depending on acclimation (and
adaptation) [0.3.2, 2.1.2]
• influence of acclimation on the capacity of cellular pH regulation
140
•
•
•
analysis of the capacity of cellular regulation mechanisms using pH sensitive
intracellular dyes. Transporters will also be functionally characterised in isolated gill
preparations (Deigweiher et al. unpubl.) using specific inhibitors.
pH dependent changes in the contribution of ion exchangers involved in transepithelial
proton equivalent ion exchange, and in associated energy turnover in isolated perfused
gills, following the rationale of Pörtner et al. 2000.
analysis of CO2 dependent performance indicators (e.g. protein synthesis, growth,
larval development) [0.3.1, 2.1.1, 2.1.3, 2.2.1, 2.3.1, 3.1.3] and regulators (e.g.
adenosine, neurotransmitters) at low and high temperatures and thermal optima, at
various levels of acid-base parameters to define the vulnerability of important fish
species [5.3].
Mechanism based modelling, building on functional studies
Mechanism based modelling of the complex acid-base regulatory system and responding
metabolic and functional characters will support predictions of set-points in acid-base regulation
and their relevance in whole organism functioning. In thinking generally about the effects of
temperature and CO2 on acid-base status, it is helpful to start by modeling a single compartment,
which represents either particular cells or all cells collectively. We intend to investigate acid-base
regulation in relation to aerobic / anaerobic metabolism and CO2/ion exchange between the
organism and the environment. The model is described in terms of rate equations, being a system
of coupled ordinary differential equations (ODEs). Based on this model we will use the metabolic
control analysis (MCA) theory to examine the regulatory aspects of the acid-base status,
including a shift of extracellular pH, and to infer the systemic control properties of CO2 and
temperature variations on the level of the functional protein.
• model of transport of inorganic ions (e.g. Na+, H+, Cl-, HCO3-) based on the transport
kinetics of identified transporters and on analyses of energy turnover attributed to
individual ion transport mechanisms as well as pH dependent changes in such energy
turnover
• model of acid-base regulation on short time scales, based on an open CO2 system and
starting with a one compartment model comprising all known biochemical reactions
(including buffering and net proton equivalent ion exchange)
• simulation of acid-base regulation on long time scales: introduction of feedback
phenomena caused by long term acclimation of the organism to environmental stresses
in terms of adjusted model parameters reflecting the short term behaviour
Internal cooperation:
Projects 0.3.1, 0.3.2, 2.1.2, 2.1.3, 2.2.1, 2.2.2, 2.3.1, 3.1.3, 3.1.4, 5.3
External cooperation:
University Bremen, DFG project “Calcification” (Heilmayer)
141
Schedule
2.3.2
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Set up of aquarium and culture
facilities
Sampling of fish, larval cultures
Incubation experiments, in vivo data,
tissue sampling
Isolation of transporter genes, library
construction
Tissue specific expression
Analysis of gill ion exchanger
activities in gill tissues and after
heterologous expression
Set-up of mechanistic model for
integrative analysis of acid-base status
Evaluation of physiological data and
running of mechanism based model
Milestones (2.3.2)
- Aquarium facilities and animal cultures established
- Completion of first incubation experiments in vivo
- Set-up of tissue and cell preparations for functional studies
- Isolation of genes and proteins of relevant transporters
- First experimental series and sample analyses completed
- Set-up of integrated model of carbonate chemistry and ion transport
- Second series and sample analyses completed
- Data analysis and comparison of species completed, publication
- Simulation of acid-base regulation and sensitivity studies, publication
- Final BioAcid report
month 6
month 6
month 9
month 9
month 18
month 24
month 30
month 36
month 36
month 36
First Year
Personnel costs
2.3.1
2.3.2
Subtotal
Consumables
2.3.1
2.3.2
Subtotal
142
Second Year
Third Year
Total
First Year
Second Year
Third Year
Total
Travel
2.3.1
2.3.2
Subtotal
Services
2.3.1
2.3.2
Subtotal
Other costs, studentships
2.3.1
2.3.2
Subtotal
Total
2.3.1
Personnel costs: 1 PhD position for 2 years for the studies on fish,
1 PhD position for 2 years for the studies on cephalopods. The missing two years for both PhD
students will be supplied from the EU-Project EPOCA
Consumables: €/year for chemicals (biochemical analysis), aquarium equipment…
Travel: €/year for animal collection and transport, meetings and conferences…
Services: €/year for the measurements of the elementary composition of statoliths and otoliths
using Laser Ablation techniques
Other costs: Student help:
otolith/statolith preparation
€/year for assistance in the rearing of larval fish and cephalopods,
2.3.2
Personnel costs: 3 x
(3 PhD-students, total
per year). Different skills and
specializations are needed for each of the three fields of molecular and membrane physiology, of
acid-base and functional studies up to organismal levels and of mechanism based modelling.
Consumables: Chemicals, molecular biology (e.g. Sequencing:
,- €; antibodies €, inhibitors etc., total: per year)
,-€; expression studies:
Travel: field work, animal collection and transport; meetings etc. per year
Investment: NA
Other costs: NA
143
vi. References
Caldarone EM, Clemmesen CM, Berdalet E, Miller TJ, Folkvord A, Holt GJ, Olivar MP and Suthers IM (2006) Intercalibration of four
spectrofluorometric protocols for measuring RNA/DNA ratios in larval and juvenile fish. Limnol Oceanogr Methods 4: 153-163.
Clemmesen C (1994) The effect of food availability, age or size on the RNA/DNA ratio of individually measured herring larvae: laboratory
calibration. Mar Biol 118:377-382
Clemmesen C and Doan T (1996) Does the otolith structure reflect the nutritional condition of a fish larva? - A comparison of otolith structure
and biochemical index (RNA/DNA ratio) determined on cod larvae. Mar Ecol Prog Ser 138:33-39.
Clemmesen C, Buehler V, Carvallo G, Case R, Evans G, Hauser L, Hutchinson WF, Kjesbu OS, Mempel H, Moksness E, Otteraa H, Paulsen
H, Thorsen H and Svaasand T (2003):Variability in condition and growth of Atlantic cod larvae and juveniles reared in mesocosms:
environmental and maternal effects. J Fish Biol 62:706-723.
Deigweiher, K, Koschnick, N, Pörtner HO and Lucassen M (2008). Adaptation of ion regulatory capacities in gills of marine fish under
hypercapnic acidosis. submitted.
Franke A, Clemmesen C and Riebesell U. The effect of changes in ocean pH on the development and early larval phase of herring. Manuscript
in prep.
Ishimatsu A, Kikkawa T, Hayashi M, Lee K-S and Kita J (2004) Effects of CO2 on marine fish: larvae and adults. J Oceanogr 60, 731-741.
Kroon, B. and Thoms, S. (2006). From electron to biomass: a mechanistic model to describe phytoplankton photosynthesis and steady state
growth rates. J. Phycol. 42: 593-609.
Langenbuch, M and Pörtner, HO (2003). Energy budget of hepatocytes from Antarctic fish (Pachycara brachycephalum and Lepidonotothen
kempi) as a function of ambient CO2: pH-dependent limitations of cellular protein biosynthesis? J. exp. Biol. 206: 3895-3903.
Larsen, BK, Pörtner HO and Jensen FB (1997) Extra- and intracellular acid-base balance and ionic regulation in cod (Gadus morhua) during
combined and isolated exposures to hypercapnia and copper. Mar. Biol. 128: 337-346.
Lucassen, M, Koschnick, N, Eckerle, LG and Pörtner, HO (2006). Mitochondrial mechanisms of cold adaptation in cod (Gadus morhua L.)
populations from different climatic zones. J. Exp. Biol. 209: 2462-71.
Mark FC, Lucassen M and Pörtner HO (2006) Thermal sensitivity of uncoupling proteins in polar and temperate fish Comp. Biochem. Physiol.
D 1, 365–374.
McKim JM (1977) Evaluation of tests with early life stages of fish for predicting long-term toxicity. J Fish Res Board Can 34:1148-1154
Melzner F, Forsythe JW, Lee PG, Wood JB, Piatkowski U and Clemmesen C (2005) Estimating recent growth in the cuttlefish Sepia
officinalis: Are nucleic acid based indicators for growth and condition the method of choice? J Exp Mar Biol Ecol 317:37-51.
Metzger R, Sartoris FJ, Langenbuch M and Pörtner HO (2007) Influence of elevated CO2 concentrations on thermal tolerance of the edible
crab Cancer pagurus. J. therm. Biol. 32: 144-151.
Michaelidis, B, Ouzounis, C,Paleras, A and Pörtner HO (2005) Effects of long-term moderate hypercapnia on acid-base balance and growth
rate in marine mussels (Mytilus galloprovincialis). Mar. Ecol. Progr. Ser. 293: 109-118.
Michaelidis B, Spring ,A and Pörtner HO (2007). Effects of long-term acclimation to environmental hypercapnia on extracellular acid-base
status and metabolic capacity in Mediterranean fish Sparus aurata. Mar. Biol. 150: 1417-1429.
Pörtner, HO, Reipschläger, A, and Heisler, N (1998) Metabolism and acid-base regulation in Sipunculus nudus as a function of ambient carbon
dioxide. J. exp. Biol. 201: 43-55.
Pörtner, HO, Bock, C, and Reipschläger, A (2000) Modulation of the cost of pHi regulation during metabolic depression: a 31P-NMR study in
invertebrate (Sipunculus nudus) isolated muscle. J. Exp. Biol. 203: 2417-2428.
Pörtner, HO, Langenbuch, M and Michaelidis, B (2005) Synergistic effects of temperature extremes, hypoxia, and increases in CO2 on marine
animals: From Earth history to global change, J. Geophys. Res. 110: C09S10
Pörtner, HO, and Knust R (2007) Climate change affects marine fishes through the oxygen limitation of thermal tolerance. Science 315: 95 97.
Pörtner HO (2008) Ecosystem effects of ocean acidification in times of ocean warming: a physiologist’s view. Mar. Ecol. Progr. Ser. in review
Thoms, S, Pahlow, M, and Wolf-Gladrow, DA (2001). Model of the carbon concentrating mechanism in chloroplasts of eukaryotic algae. J.
Theor. Biol. 208: 295-313.
Thoms, S and Wolf-Gladrow, DA Relaxation time conctants of the carbonate system at steady states. Manuscript in prep.
Zumholz K, Hansteen TH, Klügel A and Piatkowski U (2006) Food effects on statolith composition of the common cuttlefish (Sepia
officinalis). Mar Biol 150:237-244
Zumholz K, Hansteen TH, Piatkowski U and Croot PL (2007a) Influence of temperature and salinity on the trace element incorporation into
statoliths of the common cuttlefish (Sepia officinalis). Mar Biol 151: 1321-1330
Zumholz K, Hansteen, T, Hillion F, Horreard, F and Piatkowski, U (2007b) Elemental distribution in cephalopod statoliths: NanoSIMS
provides new insights into nano-scale structure. Rev Fish Biol Fisheries 17: 487-491
Zumholz K, Klügel, A, Hansteen T and Piatkowski U (2007c) Statolith microchemistry traces the environmental history of the boreoatlantic
armhook squid Gonatus fabricii. Mar Ecol Prog Ser 333: 195-204
144
11.4: Theme 3: Calcification - sensitivities across phyla and ecosystems
Marine calcification processes, especially those of dominant planktonic autotrophs (e.g.
coccolithophores), have a considerable impact on the marine carbon cycle . While heterotrophic
calcifiers are less important in terms of their contribution to the ocean carbon cycle, they play a
crucial role in structuring ecosystems, mainly by creating complex three-dimensional habitats
(‘ecosystem engineers’: reef building warm- and cold-water corals, bivalve beds). Others (e.g.
sea urchins; pteropods) control ecosystems by means of grazing (e.g. on kelp forests or
phytoplankton of high latitudes).
The acidification of the ocean will have a variety of effects on its inhabitants and their metabolic
performance, particularly on the calcifying organisms. Most of these effects will occur through
the shifts in the carbonate system, where acidification leads to an increase of CO2 and a decrease
in CO32-.
CO2 + H2O ↔ H+ + HCO3- ↔ 2H+ + CO32Marine photosynthesis and organic matter mineralization are closely linked to calcification and
CaCO3 dissolution:
⇐dissolution
⇐ respiration
Ca2+ + 2HCO3- ↔ CaCO3 + H2O + CO2 ↔ CaCO3 + O2 + biomass (CH2O)
calcification ⇒
photosynthesis ⇒
In words, calcium carbonate precipitation buffers to some extend the pH increase due to
photosynthetic CO2 fixation, and decalcification buffers the pH decrease due to respiratory CO2
release.
The marine calcification process is almost entirely biogenic and, in the absence of
photosynthesis, releases CO2 to the atmosphere. Calcification can occur when the concentration
product ([Ca2+]x[CO32-]) exceeds the saturation constant. The saturation constant decreases with
temperature, i.e. warmer water is stronger oversaturated at the same calcium and carbonate
concentrations. The state of seawater with respect to calcium carbonate saturation is often
conveniently expressed as Ω, below 1 indicates under-saturation and above 1 over-saturation.
Seawater of pH 8.2 is oversaturated for calcite, at 25oC ca. 7 times (Ω=7), at 50C ca. 3 times
(Ω=3), under which condition no precipitation occurs. For precipitation a higher over-saturation
is needed, either driven by shifts in the carbonate system or by active transport of protons and
Ca2+ by ion-pumps at the calcifying sites. Altered carbonate chemistry of future oceans will
probably change calcification rates in most ecosystems. Calcification rates in many marine
invertebrates correlate well with declining sea water Ω, while some are surprisingly tolerant of
acidification induced changes in Ω (see Fig. 3.2, Ch 3.1). Conversely, calcium carbonate
dissolution can only occur in under-saturated seawater, but the kinetics are even less well known
and depend on surface area.
145
This theme is composed of projects studying calcification and decalcification mechanisms,
signals in biogenic carbonates to obtain insights in formation, as well as evidence from past and
recent events and the effects of pH stress on calcification both on ecosystem- and organism level.
The projects on calcification mechanisms will study the pH-sensitivity of the transport of ions
across membranes of marine calcifying organisms, and the sensitivity of the coupling between
benthic microbial photosynthesis and calcification. By both mechanisms the over-saturation must
be enhanced, but whereas benthic photosynthesis leads to a simple shift in carbonate equilibria,
the biochemical mechanisms will include a more complete control by the organisms over the
local chemistry of the calcifying site. The mechanism of calcification is studied from the biogenic
carbonate side as well. Calcifying organisms maintain their own distinct trace metal homeostasis,
which results in characteristic and species specific elemental ratios as well as in peculiar isotope
fractionation (“vital effect”) which is significantly different from any inorganic-thermodynamic
expectations. This “vital effect” is most likely due to the development of biochemical
mechanisms to keep the trace metal homeostasis of the most important divalent cations, and other
trace elements, e.g. strontium, in narrow limits in order to meet certain physiological needs. It is
thus expected that trace metal concentrations in the biogenic carbonates will hold clues on its
formation, and is effected by the oceanic pH. The effect of pH decrease on several highly
sensitive calcifiers will be studied on the organism level. Since the calcium carbonate saturation
constant is temperature dependant, a future sub-saturation with respect to aragonite will begin in
the Southern Ocean and in sub-Arctic regions. Therefore, the local calcifying key-organisms,
pteropods, may be most vulnerable for ocean acidification. Beside the vulnerability of subpolar
regions also tropical coral reef ecosystems are globally threatened by a twofold effect of the
increasing CO2 concentration in the atmosphere: Global warming as a major cause of coral
bleaching and ocean acidification. Although it is generally assumed that corals are negatively
affected at a certain level of pCO2, differential effects in corals and other calcifiers indicate that
the picture may be much more complicated than previously thought. Of particular interest is how
different ontogenetic stages respond to pH stress.
The ongoing acidification appears not to be a unique event, but is documented for at least 4 times
in the period between 52.5 till 58.4 Ma BC. These events were marked by global warmings, 4x
current CO2 concentrations and a low seawater pH. The hypothesis will be tested that
nannoplankton is less sensitive to acidification as bivalves, and compared to fossil records of
these periods. Research directed towards decreasing the CO2 concentrations has focused on
increasing marine or terrestrial net productivity (e.g. the oceanic iron fertilization experiment).
However, also oceanic calcification is a potential atmospheric CO2 modifier: reduced
calcification may at least partially compensate the increase of CO2, provided the decreased
calcification does not lead to a decreased organic carbon burial. Decalcification leads to a pH
increase and thus can buffer the effects of increasing CO2. This hypothesis will be tested by
experiments with Antarctic- and North Sea sediments.
Calcification and decalcification mechanisms
3.1 Cellular mechanisms of calcification
4 subprojects. PI: Melzner, co-PIs: Bleich, Form, Schulz
Within this project we will study the physiological mechanisms of calcification in a variety of
calcifying marine organisms: echinoderms, bivalves, cold-water corals, coccolithophores, ranging
from temperate to arctic regions. The methodology includes molecular biological, biochemical
146
and cell physiological approaches, and the groups will strongly profit from each others expertise.
The project is internally well integrated as explained in Ch 3.1.iv. The project will study
calcification mechanisms from the organism side, i.e. ion transport to the calcifying sites. It is
thus complementary to project 3.3, which focuses on mechanistic research on the solid phase.
3.2 Calcification under pH-stress: Impacts on ecosystem and organismal levels
4 subprojects. PI: Tollrian, co-PIs: Riebesell, Richter, Fietzke
In this project we will study the effect of pH stress on organismal level. Investigated will be reef
survival under pH stress (on reef and colony level), pH effects on coral recruitment, calcification
of pteropods during critical development stages, and calcification in red coralline algae. A wide
range of methods will be used, ranging from organisms physiological techniques, imaging of
individuals and colonies, and solid phase chemistry and physics. A strong synergy will be
obtained from collaboration with project 3.4 and 3.3, e.g. by comparing studies on solid
chemistry, metabolic rate and microenvironment assessments using microsensors. Organisms will
be studied from temperate and tropical areas.
3.3 Ultra-structural changes and trace element / isotope partitioning in calcifying organisms
(foraminifera, corals)
2 subprojects. PI: Bijma, co-PI: Eisenhauer
This project will investigate the isotope composition and physical parameters of the shells of
corals and foraminifera to obtain information on the mechanism of calcification. The
methodology is based on a state of the art solid phase analyses. The organisms will be sampled
from temperate and tropical regions. The solid phase analyses will strongly support project 3.4,
and the studies towards the mechanisms clearly interlink with Project 3.1. However, different
from project 3.1, mechanistic information is derived primarily from the biogenic carbonates.
3.4 Microenvironmentally controlled (de-)calcification mechanisms
3 subprojects. PI: Böttcher, co-PIs: de Beer, Hoppema
Within this project we will study benthic calcium and carbonate cycling, controlled by the
metabolic activity of microorganisms, and in how far this is influenced by water column
chemistry. The benthic conversions are subjected to a transport resistance in the sediments and
boundary layer, which has profound effects on the sediment and porewater chemistry. The
methodologies are mainly geochemical, including solid phase physics and chemistry, porewater
composition, and measurements of transport- and metabolic rates. A modeling approach will be
implemented and applied in several of the subprojects. Collaborative microsensor experiments
are planned with Melzner and Form (3.1), Fietske and Richter (3.2). Joint solid phase analyses
are planned with Eisenhauer (3.3). The studies will be done on benthic systems from tropical,
temperate and Arctic regions.
3.5 Impact of present and past ocean acidification on metabolism, biomineralization and
biodiversity of pelagic and neritic calcifiers
3 subprojects. PI: Immenhauser, co-PIs: Mutterlose, Meier.
This project aims to assess traces of past pH events in bivalve shells, foraminifera and
coccolithophores. The fossil record from past hyperthermal and acidification events combined
147
with analyses on experimentally grown shells will give insight in how environmental parameters
influence the shell chemistry and physics. This project includes solid phase chemistry and
physics, thus a collaboration with the partners in project 3.3 is evident. The samples originate
from temperate and tropical areas.
Project 3.1: Cellular mechanisms of calcification
(PI: Frank. Melzner)
i. Objectives
Research in four sub–projects will focus on elucidating the mechanisms and sensitivities of
calcium (Ca2+) and dissolved inorganic carbon (DIC) transfer to calcification sites. Emphasis will
be placed on identifying the role ion transporters and channels play therein, both in autotroph
(coccolithophores) and heterotroph (corals, molluscs, echinoderms) taxa. Comparing relatively
sensitive with tolerant calcifiers will help identify the limiting components in biogenic
calcification machinery. In addition, the differing DIC requirements of calcification vs.
photosynthesis will be investigated in autotroph model organisms.
Figure 3.1: An overview of how the proponents combine expertise and resources to characterize cellular calcification
responses, from the gene regulatory level to cell and tissue function.
148
Fig. 3.2: The dependence of calcification on seawater
calcium carbonate saturation (Ωarag) in marine
invertebrates: Long-term coral reef data set recorded in
the Biosphere 2 mesocosm (Langdon et al. 2000;
Milliman 1993), the bivalve Mytilus edulis calcification
(Gazeau et al. 2007), the cephalopod Sepia officinalis
(Gutowska, Pörtner, Melzner, submitted). Control
calcification rates were set at 100%.
Altered carbonate chemistry of future oceans will probably change calcification rates in both,
marine auto- and heterotroph taxa. Coral, echinoderm and bivalve calcification rates correlate
extremely well with declining sea water calcium carbonate saturation state, Ω (Fig 3.2), while
others (cephalopods) are surprisingly tolerant of acidification induced changes in Ω. It is not
known yet as to why, in many species, Ω gives the best correlation with calcification
performance, as the carbonate ion is not considered the biologically active form of dissolved
inorganic carbon (DIC): Rather, CO2 can cross biological membranes, and protons, as well as
bicarbonate ions (HCO3-), are chief substrates of ion–transport proteins important for cellular
homeostasis and carbon acquisition (e.g. Na+/H+ exchangers, Na+ dependent Cl-/HCO3exchangers etc). It is still a matter of debate, which transport proteins are involved in Ca2+
transport towards the calcification site and if the paracellular or transcellular route is used.
Involvement of Ca2+ ATPases and calcium channels has been demonstrated at least in some
groups (Allemand et al. 2004). However, studies that investigate the ion transporters involved in
combination with the functional properties of epithelia and whole cells in detail are scarce.
(1) Autotrophs: calcification sensitivity might be influenced by photosynthesis
Unicellular coccolithophores are complicated model systems at the base of the marine food web,
as both calcification and photosynthesis have a high demand for DIC. They have been shown to
actively take up CO2 and HCO3-, operating a so-called carbon concentrating mechanism (CCM)
(Rost et al. 2003; Schulz et al. 2007). However, similar to heterotrophs, differing sensitivities
towards elevated CO2 have been observed in this taxon: While cellular calcification in Emiliana
huxleyi was found to linearly decrease with increasing CO2 levels (Riebesell et al. 2000),
Coccolithus pelagicus was hardly affected by changes in the carbonate system, and Calcidicus
leptoporus showed an optimum curve with maximum particulate inorganic carbon (PIC)
production rates at present day CO2 (Langer et al. 2006). A species specific response in
particulate organic carbon (POC) production with varying CO2 was also observed, urgently
calling for a process based understanding of CO2 dependent carbon acquisition for both
photosynthesis and calcification in coccolithophores (see sub project 3.1.1).
149
(2) Basal heterotrophs: coral models without photosynthetically active symbionts.
Complex interactions between calcification and photosynthesis complicate the study of ion
sources and supply for calcification in hermatypic warm–water corals (Allemand et al. 2004;
Gattuso et al. 1999). During biomineralisation, corals have to supply calcium and inorganic
carbon to the calcification site (to the extracellular calicoblastic fluid, ECF) but also need to
eliminate protons resulting from CaCO3 production (McConnaughey & Whelan 1997). Transport
of molecules across the calicoblastic membrane into the ECF could be achieved by two
mechanisms: a paracellular pathway (driven by a chemical or electrochemical gradient:
diffusional), or a transcellular pathway through the cell (active transport with specific membrane
transporters such as carriers and pumps). While there is some evidence for carrier mediated (e.g.
Ca2+/H+ ATPase, L-type Ca2+ channels) calcium transport across the calicoblastic epithelium in
warm-water corals (Tambutte et al. 1996; Zoccola et al. 2004), the pathways for DIC are largely
unknown. Inhibitor studies point to the involvement of anion exchangers (e.g. HCO3-/Clexchangers) in these epithelia (Tambutte et al. 1996). Carbonic anhydrase (CA) is also a key
component of the calcification machinery, inhibition of this enzyme resulted in an 80% reduction
of calcification rate in tropical corals (Tambutte et al. 2007). Within calicoblastic cells, it may
primarily mediate between metabolically produced CO2 and HCO3- destined for the ECF. A
significant part of the DIC incorporated by coral calcification probably stems from metabolically
produced CO2, either produced by calicoblastic cells or by photosynthetically active symbiont
cells(Furla et al. 2000). Cold-water corals, such as Lophelia pertusa, do not contain
photosynthetic symbionts, making them ideal study objects to solely consider calcification related
ion transport. Experiments with tropical reef building corals demonstrated that a lowering of the
carbonate ion concentration significantly reduces coral calcification and growth (Langdon et al.
2003; Marubini et al. 2003; Schneider & Erez 2006) (see Fig 3.2).
In the middle of this century, many tropical coral reefs may well erode faster than they can
rebuild. Cold-water corals are living in an environment (high latitude, cold and deep waters) with
carbonate saturation already close to a critical carbonate saturation (i.e. Ω=1) below which
CaCO3 will dissolve. Projections indicate that about 70% of the currently known Lophelia
pertusa reef structures will be exposed to sub-saturating conditions by the end of the century
(Guinotte et al. 2006). This illustrates the necessity to learn more about the processes that
influence calcification in these organisms (see sub project 3.1.2).
(3) Complex heterotrophs: calcification sensitivity gradients within a phylum
Certain molluscs may react similarly sensitive towards acidified sea water, especially the class
bivalvia (see Fig 3.2). Exposed to elevated seawater pCO2, the genus Mytilus displays (1) an
inability to fully compensate hemolymph pH, (2) decreased somatic growth rates, (3) decreased
rates of calcification, (4) an inability to control blood Ca2+ concentration, which might be an
indicator of internal shell dissolution at low hemolymph pH (Gazeau et al. 2007; Michaelidis et
al. 2005). Quite the contrary, the cephalopod mollusc Sepia officinalis does not suffer from any
of these physiological perturbations when exposed to comparable sea water pCO2 (Gutowska,
Melzner, Pörtner et al., submitted, Melzner et al. unpublished). In fact, S. officinalis is the only
marine invertebrate studied so far, that does not show a decrease in calcification rate at elevated
pCO2 values predicted for the next 300 years (Gutowska, Pörtner, Melzner, submitted). A high
ion–regulatory capacity in combination with a certain natural pre–adaptation to transiently high
pCO2 values, as can be observed in cephalopods, might be the key to tolerance. Little is known
about the exact ion transport mechanisms related to calcification in molluscs. A higher
organismic complexity (with respect to coccolithophores and corals) led to the evolution of
150
specialized ion exchange organs, the gills. Several candidate genes / proteins with a relevance to
calcification processes have already been identified in both, Mytilus spec. and S. officinalis gills
(e.g. Na+/K+ ATPase, Na+/HCO3- cotransporters (NBC) and HCO3-/Cl- exchangers, CAs, Ca2+
ATPases, Ca2+ channels (Medakovic 2000; Piermarini et al. 2007; Melzner, Lucassen, Gutowska,
Hu unpublished), their expression patterns in response to high pCO2 have, however, not yet been
considered. A recent study on CO2-induced acclimation processes relevant for acid-base
regulation in fish gills demonstrated that several important gill ion transporter mRNAs were
differentially expressed during a six week time-course experiment (e.g. Na+/K+ ATPase, NBC,
AE, NHE, Deigweiher et al., submitted). Elevated Na+/K+ ATPase and NBC capacities at the end
of the experiment may be reflected in elevated costs of ion and acid base regulation under
elevated CO2. As several of the differentially regulated transporters also play a role in cellular
DIC accretion, these results indicate how intricately calcification and general pH regulatory
processes are intertwined, and suggests that their metabolic machinery should be studied
simultaneously (see subproject 3.1.3).
(4) Larval heterotrophs: ontogenetic calcification sensitivity gradients
Similarly, sea urchin embryos and larvae are suited for studies that investigate both, calcification
and pH homeostasis due to their use as model organisms for the past 100 years. There is a large
amount of data available on developmental processes and gene regulatory networks in this taxon
(Poustka et al. 2007). Still, functional data examining pH and ion regulation on a cellular level
are limited. Sequencing of the genome of Strongylocentrotus purpuratus has now been
completed (Sodergren et al. 2006) and renders molecular genetic approaches highly attractive to
identify candidate genes for proteins involved in homeostatic processes for ion composition,
volume regulation and pH. Larval stages of marine ectotherms have been shown to react much
more sensitively towards ocean acidification than their respective adult stages (Ishimatsu et al.
2005). This indicates that compensatory mechanisms which are recruited in adults are not
functionally active in early development. In several sea urchin species, stark decreases of larval
calcification rate with water pCO2 have been recorded (Fig 3.5, Kurihara & Shirayama 2004.
Stumpp & Melzner, unpublished). Mechanisms leading to retarded growth are largely unknown,
as are the major pathways for Ca2+ and DIC transport in echinoderms. How these pathways
function and how they are being modified in the course of ontogeny, thereby altering sensitivity
towards high water pCO2, constitutes an important research challenge (see subproject 3.1.4).
In summary, the ion transport pathways relevant for calcification are poorly understood in many
marine phyla, indicating that basic research efforts have to be undertaken to (i) identify gene
products that code for important ion transport proteins, (ii) monitor levels and / or activities of
mRNAs and proteins in response to abiotic stress, (iii) study the function of cells and epithelia
where these proteins are active, (iv) study fluxes of Ca2+ and DIC from the sea water into
organisms and cells and, finally (v) estimate the capacity of key model organisms to acclimate
their ion transport systems to an altered carbon system (see Fig 3.1).
3.1.1: Kai Schulz combines all the expertise necessary to successfully study the proposed
hypothesis, e.g., coccolithophore culturing, carbonate system manipulations, mass spectrometry
(Schulz et al. 2006; Schulz et al. 2007; Schulz et al. 2004). Furthermore, employed as a research
scientist at IFM-GEOMAR in Kiel he has full access to unique infrastructure, essential for an
effective experimental implementation (centralized CO2 aeration system, membrane-inlet-massspectrometer lab).
151
3.1.2: During the last three years Armin Form has established and improved a world-wide
unique cultivation facility for high latitude marine organisms and has developed many new
methods for laboratory deep-sea coral research (Form et al., in prep.). Based on these methods
the proponent's primary research focus has been on measurements of calcification rates of coldwater corals (Form and Riebesell, in prep.) and arctic coralline red algae (Form and
Büdenbender, in prep.) in relationship to elevated CO2-levels and temperatures.
3.1.3: Frank Melzner and Magdalena Gutowska have considerable expertise in physiological
studies on mollusc stress physiology and the effects of ocean acidification on marine ectothermic
animals (Melzner et al. 2006a; Melzner et al. 2006b; Melzner et al. 2007a; Melzner et al. 2007b;
Melzner et al. 2007c), Melzner et al. submitted, Gutowska, et al., submitted). Melzner leads the
Junior Research Group ‘Ocean Acidification’ within the Kiel Excellence Cluster ‘Future Ocean’
and has access to state of the art culturing and CO2 manipulation facilities. Gutowska currently
finishes her PhD thesis at the AWI and will thereafter move as a Postdoc to Markus Bleich’s lab.
Magnus Lucassen is an expert in molecular physiology and functional genomics of marine fish
and invertebrates (molluscs, crustaceans), with a strong background in isolation and quantitative
expression analyses of key genes and their functional impact under different environmental
challenges (Eckerle et al. 2008; Heise et al. 2006; Lucassen et al. 2006; Lucassen et al. 2003),
Deigweiher et al. submitted; see also subproject 2.3.2). Hans-Otto Pörtner has a long standing
history in studying acid-base regulation and proton equivalent ion exchange of marine ectotherms
in relation to ambient water conditions, as well as the interaction of acid-base parameters with
metabolic processes (e.g. Pörtner 2002; Pörtner et al. 2000; Pörtner et al. 1990; Pörtner et al.
2005; Pörtner et al. 2004; Pörtner et al. 1993, see also subproject 2.3.2). Current interest
comprises the interaction between CO2 levels and other climatic factors, and the mechanisms
shaping cellular and whole animal energy budgets.
3.1.4: Markus Bleich has a long standing expertise in experimental ion transport physiology
(Bleich et al. 1990; Kosiek et al. 2007; Schroeder et al. 2000). In previous projects on
mammalian cells as well as on marine organisms, electrophysiology of cellular transport
processes has been characterized from the single ion channel to the systemic level (Bleich et al.
1999; Hou et al. 2007). Equipment and technology for patch clamp analysis of cells and isolated
cell membranes is in place. In addition, microfluorimetry is used for the measurement of
intracellular ion concentrations (H+, Ca2+, Na+, Cl-) and membrane voltage. From human and
experimental pharmacology in close collaboration with pharmaceutical industry a collection of
pharmacological tools is available to manipulate ion channels and transporters which are directly
involved in transport or which are responsible for the generation of driving forces (Bleich &
Greger 1997; Reuter et al. 2008). Bleich and co-workers have significantly contributed to
genome research projects for the identification of disease related genes (Barth et al. 2005) and
used molecular biological techniques to identify, clone and characterize ion channels in epithelial
transport (Waldegger et al. 1999). Kerstin Suffrian, PhD student in Bleich’s working group, has
a profound expertise in microfluorimetric experiments on nano- and micrometric marine
organisms (Suffrian, Bleich et al., unpublished) and will help train the proposed PhD student.
Frank Melzner and working group (see also 3.1.3) have established a sea urchin culture at IFMGEOMAR and recently performed first experiments on embryonic and larval development of the
sea urchin Strongylocentrotus droebachiensis under elevated pCO2 (Stumpp & Melzner,
unpublished).
152
Collaborative research within project 3.1
While the four subprojects will each focus on one important aspect of cellular calcification
mechanisms, some collaborative experiments will be performed simultaneously by all PIs (1st
quarter of years two and three) to gather comparative biochemical indicators for calcification
performance for all studied taxa. Recent findings indicate that carbonic anhydrase (CA) activity
might be a meaningful indicator for ion-regulatory capacity in marine organisms and, possibly,
sensitivity towards ocean acidification (Fabry et al. 2008, in press). CA is a crucial enzyme
important both for general acid-base regulation, calcification and especially photosynthesis, in
auto- and heterotrophic organisms (see e.g. Henry 1996). Experiments will be conducted using
the state of the art membrane inlet mass spectrometry (MIMS) setup in the lab of Ulf Riebesell
and Kai Schulz. At present, meaningful CA measurements can only be conducted using MIMS
technology, as classic biochemical CA assays operate under un-physiological conditions (cold
temperatures, low pH). CA capacity analysis will also be strongly related to studies of in vivo
acid base physiology carried out in theme II.
In addition, gene expression studies will be performed in the central molecular biology labs of
IFM-GEOMAR and Magnus Lucassen’s lab at the AWI. Using transcripts that might be of
importance for calcification processes in all organisms (CA, Na+/K+ ATPase, Ca2+ ATPase and
Ca2+ channels), we will try to answer the question, whether ocean acidification activates a
common, compensatory gene response and how their regulation capacity defines organisms
sensitivity. Which transcripts will be monitored following the first year will depend on the results
of the transcriptomic work conducted in subproject 3.1.3. Gene expression will be studied using
real-time PCR techniques. Instrumentation is available at both AWI and IFM-GEOMAR,
technical support can be provided by Frank Melzner’s working group (e.g. lab technician).
Eventually, we will correlate calcification performance of the various taxa with the
genomic/enzymatic indicators, to develop a conceptual framework of how sensitivity to future
acidification might be related to the phenotypic flexibility of the metabolic machinery. Output of
this collaborative effort will be a synthesis / review type article, co-authored by all PIs.
153
Subproject 3.1.1: Inorganic carbon acquisition for calcification and photosynthesis in
marine coccolithophores: towards a unifying theory
(Kai Schulz, IFM-GEOMAR Kiel)
Work Programme
Within the framework of the project 3.1 'Cellular Mechanisms of calcification', the apparently
different modes of carbon acquisition for particulate inorganic and organic carbon production
(PIC/POC) in marine key coccolithophores,
such as Emiliania huxleyi, Coccolithus
pelagicus and Calcidiscus leptoporus will be
investigated. The experiments planned will be
based on the hypothesis that the seemingly
different calcification and photosynthesis
dependence
on
CO2
in
various
coccolithophores are the result of species
specific thresholds for DIC demand and pH
sensitivity. Therefore we hypothesize all
species to possess certain CO2 and pH optima
for both processes. On one side of the optimum
curves photosynthesis and calcification would
be limited by the availability of DIC. While on
Fig. 3.3: Schematic diagram of the proposed effects
the other, low pH values would hinder their
of seawater and pH on photosynthesis and
calcification of coccolithophores. The process
maximum functioning (Fig. 3.3). Hence,
sensitivity observed for a certain coccolithophore
coccolithophores will be cultured under a broad
crucially depends on the experimental CO2 / pH
range and can even change its sign.
range of well defined carbonate chemistry
conditions, extending the core CO2 levels between 280 and 980 ppm towards both ends. This
allows assessment of individual CO2 sensitivities of photosynthesis and calcification.
Furthermore, employing sophisticated carbonate chemistry manipulations, it will be possible to
separate the usually tight coupling of CO2,
respective DIC concentration, and pH. Standard
measurement techniques (POC, PIC, inorganic
nutrient, growth rate determinations) will be
combined
with
membrane-inlet-mass
spectrometry (MIMS), allowing for carbon flux
measurements on the cellular level (Fig. 3.4).
Moreover, the MIMS setup will be used to
quantify cellular carbonic anhydrase (CA)
activity, a key component of many CCMs. The
MIMS setup will also be available for other
Fig. 3.4: Schematic representation of a
project partners interested in CCM activity
coccolithophorid cell taking up dissolved
measurements such as subproject 1.1.3, 1.1.4, inorganic carbon. Internally this carbon is fixed as
1.2.5, 3.4.2. Frequent experience exchange and CO2 in the chloroplast (green) while it is
discussions with project 4.2.2 will ensure high precipitated as CaCO3 in the coccolith production
vesicle (grey).
data quality. Collaboration with subproject 3.5.3
will give additional information regarding the nature of the calcification response towards
changing carbonate chemistry by automatic quantification of coccolith numbers, calcite contents
154
and morphology which allows for comparison with results obtained in 3.5.2. Finally, the results
on coccolithophorid photosynthesis and calcification in response to increasing CO2 will directly
feed into the biogeochemical modelling projects 1.3 and 5.2.
Together, the results will greatly enhance our understanding of the effects of ocean acidification
on calcification and photosynthesis. This in turn is a prerequisite for predicting the future of
marine coccolithophores in a high CO2 ocean.
Work Schedule
3.1.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Calcification / Photosynthesis
dependence of the three
coccolithophores on CO2 / DIC
Identifying their pH sensitivities
Assessment of inorganic carbon fluxes
under selected CO2 / pH conditions
(MIMS)
Data assessment and identification of
potential optima and thresholds
Collaboration within 3.1
Milestones (3.1.1)
- Identifying the calcification and photosynthesis dependencies of Emiliania
huxleyi, Coccolithus pelagicus and Calcidiscus leptoporus on CO2, respective
DIC concentration.
Month 18
- Identifying the pH sensitivity of particulate inorganic (PIC) and organic
carbon (POC) production in these three species.
Month 24
- Assessment of dissolved inorganic carbon uptake fluxes under selected CO2 /
pH conditions.
Month 30
- Data assessment and identification of potential optima and thresholds
Month 36
- Collaborative work: Gene expression patterns / MIMS
Month 30
155
Subproject 3.1.2: Transepithelial calcification processes in the hermatypic cold-water coral
Lophelia pertusa (Scleractinia)
(Armin Form, IFM-GEOMAR)
Work Programme
The hermatypic cold-water coral (CWC) Lophelia pertusa (Fig. 3.5) will be cultivated under
standardized conditions and challenged by different pCO2manipulative long- and short-term experiments (IFM-GEOMAR,
Kiel):
Long-term experiments (according to the BIOACID pCO2 core levels):
- Phase I: 12 months: 380, 560, 700 and 980 ppm CO2 at 7°C
- Phase II: 12 months: 380, 560, 700 and 980 ppm CO2 at 11°C
Short-term experiments (to accommodate synergistic aspects):
- 4 phases ranging from weeks to max. 3 months: various pCO2
levels and temperatures
Physiological functioning of the calicoblastic epithelium from the corals Fig. 3.5: Lophelia pertusa
during calcification
cultivated under these conditions will then be determined in ex-vivo measurements under
measurements in Ussing chamber and patch-clamp experiments elevated pCO2's using
(collaboration with M. Bleich & K. Suffrian, Institute of Physiology - modified alkalinity
anomaly technique
CAU, Kiel). These techniques allow the determination of transepithelial
voltage, transepithelial resistance and equivalent short-circuit currents as characteristics of
epithelial and/or cellular function (for methodical details see subproject: 3.1.4). Using
pharmacological tools and changes in the solute composition on either site of the epithelium, we
aim to uncover the transepithelial pathways for calcium and carbon during the calcification
process (milestone 2, M2) and their sensitivities to ocean acidification by identifying the involved
ion transport proteins for the corals (M3). In this context we will further address the issues of
how corals can regulate pH in the calicoblastic cells, and how they remove protons produced
during the calcification process, respectively.
In addition to the core questions, the experimental setup is designed for a maximum of crosscollaborations with several BIOACID subprojects outside 3.1:
- For characterizing the pH-gradients and Ca2+-concentrations on the different sites of the
calicoblastic epithelia, microsensor measurements will be done in collaboration with Dirk de
Beer (MPI Bremen, subproject 3.4.2). These measurements should complete our
understanding of the epithelial functioning (M2 & M3), and are invaluable precursors for
the Ussing chamber and patch-clamp experiments.
- During the long-term experiments carbonate substratum (coral skeleton) will be built
through the calcification process. This material will be analysed for microchemical and
structural changes of crystal growth (M5) in a collaboration with Jan Fietzke and Thor
Hansteen (IFM-GEOMAR Kiel, subproject: 3.2.4).
- In a collaboration with Alban Ramette (MPI Bremen, subproject 4.1.4) whole coral
branches from long- and short-term experiments will be analyzed for their microbial
community composition and their sensitivities to alterations of environmental parameters
(pH, CO2 concentration, temperature). Molecular techniques will be used to obtain a high
156
resolution description of microbial community shifts in the different experimental
treatments (M6).
- During short-term experiments the synergistic effects between elevated pCO2-levels and
hydrostatic pressures of 0.1 – 5 MPa to coral physiological performance (PP) will be
investigated in a joint study with Laurenz Thomsen and Giselher Gust (Jacobs University
Bremen, subproject 1.2.5) (M7).
For complementary comparisons between cold- and warm-water corals, results of this project
will be set in context with those of Ralph Tollrian (Ruhr University Bochum, subproject 3.2.2),
Jelle Bijma (AWI Bremerhaven, subproject 3.3.1) and Anton Eisenhauer (IFM-GEOMAR Kiel,
subproject 3.3.2).
All these collaborations in addition to the subproject internal findings should result in a
comprehensive integrated view about the effects of ocean acidification on the main hermatypic
cold-water coral Lophelia pertusa (M8).
Schedule
3.1.2
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Method development & improvement
Long-term experiments (Phase I & II)
Short-term experiments on selected
aspects
Collaborative Research within project
3.1 (MIMS, real-time PCR)
Sample processing & Measurements
Data analysis and interpretation
Joint publication of results
Milestones (3.1.2)
- Transfer and modification of physiological methods/tools (M1)
- Characterization of coral epithelial ion transport mechanisms (M2)
- Quantification of changes in ion transport mechanisms in response to prolonged
elevated CO2 (M3)
- Collaborative work: gene expression patterns / MIMS (M4)
- Quantification of microchemical and -structural changes due to prolonged
elevated CO2 (M5)
- Quantification of microbiological community sensitivity due to elevated CO2
(M6)
- Characterization of synergistic effects between pressure and elevated CO2 on PP
(M7)
- Comprehensive data set on the response of cold-water corals to ocean
acidification (all experiments) (M8)
Month 06
Month 18
Month 36
Month 30
Month 33
Month 33
Month 36
Month 36
157
Subproject 3.1.3: Calcification & ion homeostasis in the phylum mollusca in response to
ocean acidification
(Frank Melzner, (IFM-GEOMAR), Magdalena Gutowska, Magnus Lucassen, Hans O. Pörtner
(AWI Bremerhaven))
Work Programme
We will study model organisms from both, classes bivalvia and cephalopoda, integrating
genomic and physiological approaches. According to areas of expertise, three work packages
have been assembled, which will be targeted by two PhD students that will be jointly supervised
by the WP–leaders (Frank Melzner (FM), Magdalena Gutowska (MG), Magnus Lucassen (ML),
Hans Pörtner (HP).
A good tolerant model species is available with the
cephalopod S. officinalis, currently in culture at the
AWI Bremerhaven (see Fig. 3.6). A subtracted cDNA
library (elevated pCO2 vs. control) has recently been
established for S. officinalis gill tissue (Melzner,
Lucassen, et al., in prep.), further sequence
information will be obtained using the next-generation
sequencing technology available to the Kiel
Excellence Cluster ‘Future Ocean’. Two Baltic Sea
bivalve species (Mytilus edulis and Arctica islandica)
are presently emerging as sensitive model organisms
within the Excellence Cluster working groups of F.
Melzner (Ocean acidification, A1) and P. Rosenstiel
(Marine Medicine, B2). For both organisms, large
fractions of the transcriptome are presently being
sequenced (Rosenstiel, Phillip, Melzner, work in
progress). Availability of large amounts of genomic
information is an absolute prerequisite for meaningful Fig. 3.6: Growth and calcification in the
cuttlefish Sepia officinalis incubated under
hypothesis-driven research targeting selected crucial ∼6,000 ppm CO2 (red) and control
processes. Incubations of animals for extended periods conditions (black). CaCO3 accretion shown
of time under realistic pCO2 (e.g. 380 – 1400 ppm) as bars, calcified structure shown shaded
grey in the schematic drawing.
and temperature regimes will be performed using the
IFM-GEOMAR / AWI CO2 incubation systems and wet labs and in close collaboration with
several projects of theme 2 (i.e. 2.1.1, 2.1.3, 2.2.1, 2.2.2, 2.3.1, 2.3.2), as the calcification
machinery strongly depends on intracellular ion- and pH homeostasis.
WP I: Gene expression profiling (ML, FM): Based on available sequence information,
representative transcripts will be monitored in relevant tissues (gill, calcifying tissues of the
mantle margin) of model organisms exposed for various periods (time course series of up to 8
weeks) to various intensities of abiotic stressors (CO2, T). Main processes targeted using
techniques such as realtime PCR and in situ hybridization will be the (i) ion regulatory
transcriptome in the gills (e.g. Na+/H+ exchanger (NHE), Na+/K+ ATPase, Cl-/HCO3--exchanger
(AE1), Na+/HCO3--cotransporter (NBC1) etc.), (ii) mRNA of proteins that are important for Ca2+
transport / calcification, in both, gills and calcifying epithelia (e.g. Ca2+ ATPases, Ca2+ channels,
AEs, etc.). Based on their plastic expression patterns, essential transporters involved in the
response to high CO2 / T stress will be identified. Regulated transcripts then will be studied in
158
more detail on the protein / cellular level (see below). A second approach will be more
explorative and focus on differentially expressed genes using approaches such as cDNA library
generation via subtractive suppression hybridization (SSH). This will help to identify (novel)
essential ion regulatory proteins involved and aid in developing novel hypotheses on how
elevated pCO2 might affect yet unconsidered physiological processes. Results of this approach
will also be verified by realtime PCR and further (functional) characterization. There will be a
strong collaborative link to sub-project 2.3.2, as similar approaches will be used in both projects.
WP II: Ion regulatory proteins and epithelia function (MG, ML, FM): In a first step, possible
changes in ion transport protein composition of important epithelia (gill, calcifying interfaces)
will be assessed using Western Blots and immuno-histochemical methods (e.g. Deigweiher et al.
submitted, Melzner et al., submitted). Further, enzyme ion-transport capacity (Ca2+ ATPase,
Na+/K+ ATPase) will be determined employing photometric enzyme tests (FM, ML). In a second
step, important epithelia will be isolated, and their functional characteristics will be determined
using Ussing-chamber techniques (MG, in close collaboration with Markus Bleich and his
working group, as well as with partners from sub-projects 2.1.1, 2.1.3, 2.3.2). Finally, changes in
mRNA, transporter protein concentration and transporter activities can be correlated with
functional changes in ion-transporting epithelia.
WP III: Ionic composition, acid – base physiology, calcification (HOP, FM): The third work
package will focus on how the ion regulatory machinery studied in WPs I and II affects wholeanimal performance, namely (i) ion composition of body fluids (all major cations and anions in
hemolymph, coelomic fluid, renal fluid, extrapallial fluid) (HOP, FM), (ii) acid base parameters
of extra- and intracellular compartments (HOP), (iii) oxygen transport protein physiology (FM,
link to , 2.2.1 and 2.2.2), and (iv) calcification rates at various time points during acclimation to
elevated pCO2 (FM, collaboration with sub-project 4.1.2). Calcification rates will be determined
using the ∆TA method, or by directly determining CaCO3 contents of shells in fast calcifiers
(cephalopods). Calcification performance in our model organisms will be compared to that of
other molluscs (pectinids, 2.1.3, pteropods 3.2.1). In a collaboration with Dirk DeBeer (see
3.4.2), it is further planned to measure the ionic composition of extrapallial fluid directly at the
calcification site using microsensors. This work package will provide information on the degree
of acidification that can be compensated by the ion-regulatory and physiological machinery.
Schedule
3.1.3
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Incubation experiments, tissue / body
fluid sampling, calcification rates,
sample preparation
cDNA library construction / analysis,
sequencing, gene identification
Gene expression analysis (qRT PCR)
Protein functional properties (Enzyme
activities, Western blots, immuno histochemistry)
Epithelial function (Ussing chambers)
Acid – base parameters (intra /
extracellular), ion composition of body
fluids
159
3.1.3
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Collaborative work within the project
(MIMS, Gene expression studies)
data interpretation
Milestones (3.1.3)
- Dataset on calcification rates, tissue / body fluid samples
month 09
- Dataset on gene expression patterns
month 15
- Dataset on protein functional properties
month 21
- Dataset on acid - base / pH regulation
month 21
- Dataset on epithelial function
month 33
- Completion of PhD theses / defense
month 36
160
Subproject 3.1.4 Sea urchin membrane transport mechanisms for calcification and pH
regulation
Markus Bleich, Kerstin Suffrian (CAU Kiel), Frank Melzner (IFM-GEOMAR)
Work Programme
Preliminary work for the project:
Animals: We have collected Strongylocentrotus droebachiensis from the western Baltic Sea and
maintain them for continuous use in the aquarium facilities of the IFM-GEOMAR in Kiel. In
addition continuous supply of sea urchins has been organised using routine cruises of IFMGEOMAR research vessels to substitute the existing stock. Fertilized eggs are routinely
generated and different embryonic stages and larvae are available. The maintenance of adult as
well as embryonic to larval sea urchins can be performed under any CO2 partial pressure between
380 and 1,400 ppm in a newly established facility.
Genomics: In a collaborative project within the cluster of excellence “Future Ocean” (Frank
Melzner, Philip Rosenstiel), sequencing maps of S. droebachiensis are currently aligned with the
genome of S. purpuratus and provide the data base for the search of homologous genes coding
for membrane transport proteins involved in H+, HCO3- and Ca2+ transport, and for proteins
involved in the generation of membrane voltage as a key driver of electrogenic transport
processes.
Infrastructure: One lab is available to the group and equipped for all measurements described in
the work plan. A setup is ready for the installation of the imaging equipment which has been
applied for.
Key questions: i. which membrane proteins are responsible for ion transport? ii. which ion
channels are pH sensitive? iii. which mechanisms are involved in pH and Ca2+ homeostasis? iv.
does chronic environmental CO2 incubation change the functional properties of the cells? v. does
chronic environmental CO2 incubation change the expression of transport proteins?
Methods:
Patch clamp analysis: Whole cell conductance measurements will be performed on isolated
cells from embryonic stages to determine the basic electrophysiological properties and to
characterize the ion transport mechanisms involved in pH homeostasis, Ca2+ metabolism and
membrane voltage generation. The sensitivity of the respective ion channels towards changes in
pH/CO2 will be investigated on the single channel level in isolated membrane patches.
Microfluorimetry: Cells from embryonic stages will be challenged by acute changes in ambient
pH/CO2. They will be monitored via pH sensitive fluorescent dye indicators for their ability to
counter-regulate as a measure for the expression of pH regulatory mechanisms. The same
experiments will be performed after chronic incubation at increased CO2 partial pressures.
Fluorescence measurements of cytosolic Ca2+ will reveal the status of Ca2+ metabolism with
respect to its role for calcification and as a second messenger molecule.
Molecular biology: Realtime PCR of membrane transport protein candidate gene mRNAs will
be performed to validate the results suggested from functional studies and database mining. The
influence of chronic CO2 elevation on the expression level will be investigated.
161
Networking with projects and sub-projects in other themes:
The projects under themes 1-5 provide an excellent platform for interactive networking on a
methodological as well as on a data driven basis. Obvious contact points are within project 1.1
(acclimation versus adaptation; subproject 3/4) with respect to the analysis of genetic and
proteomic changes in calcifiying organisms on long-term exposure to CO2. Project 2.1 is
especially well suited for a strong interaction since similar questions are asked and similar
methods are used here for the analysis of sea urchin oocytes, while subproject 3.1.4 takes over
after fertilization. We expect exchange on cellular mechanisms and technical progress. Also
project 2.3 has one focus on cellular mechanisms of CO2/pH sensitivity on development, growth
and metabolism and finally interaction with subproject 4.1.2 could provide added value in the
discussion how survival and performance of early life stages are affected.
Schedule
3.1.4
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Set-up of facilities and instrumentation,
hiring of PhD student (M1, M2)
Patch clamp experiments (M3)
Microfluorimetry (M4)
Data base mining and gene
identification (M5)
Quantification of specific membrane
transport mechanisms (M6)
Gene expression analysis (M7)
data interpretation (M3 – M8)
Milestones (3.1.4)
- Setup of a new microfluorimetric imaging equipment (M1)
Month 03
- Characterization of embryonic cells by patch-clamp technique (M2)
Month 12
- Characterization of embryonic cells by microfluorimetry (M3)
Month 18
- Database mining to identify candidate genes for the respective transporters and
channels (M4)
Month 21
- Quantification of specific membrane transport mechanisms in embryonic cells
after chronic pre-incubation at different CO2 concentrations (M5)
Month 33
- Quantification of the expression level of the candidate genes which might be
functionally relevant at different CO2 concentrations (M6)
Month 36
- Generation of a cell model for membrane transport mechanisms in embryonic cells
(M7)
Month 36
Optional: Expansion of measurements to cells from other developmental stages
according to M2-M7
Month 36
162
First Year
Second Year
Third Year
Total
Personnel costs
3.1.1 PhD student
3.1.2 Postdoc position
3.1.2 Student contracts
3.1.3 PhD student AWI
3.1.3 PhD student IFMGEOMAR
3.1.4 PhD student
Subtotal
Consumables
3.1.1
3.1.2
3.1.3 AWI
3.1.3 IFM-GEOMAR
3.1.4
Subtotal
Travel
3.1.1 meetings
3.1.2 meetings
3.1.3 meetings AWI
3.1.3 meetings IFMGEOMAR
3.1.4 meetings
Subtotal
Investment
3.1.1 MIMS maintenance
3.1.4
Subtotal
TOTAL
163
3.1.1
Personnel costs: Most of the work will be done by a PhD candidate, to be employed for three
years.
Consumables, chemicals, gases, analyzes: Money for consumables will be needed for various
analyzes such as elemental composition analyzes on a mass-spectrometer, nutrient analyzes,
chemicals, gases and isotopic analyzes.
Investments and maintenance: The membrane-inlet mass-spectrometer will need frequent (at
least annual) maintenance to ensure high data quality.
Travel costs and publication charges: Travel of the PhD student and one supervisor to an
international conference and publication charges.
3.1.2
Personnel costs:
Postdoc position
for Armin Form: Due to the complex nature of the proposed
research program and the very challenging animal model, an experienced researcher is needed for
this project. Armin Form is currently finishing his Ph.D. thesis on effects of elevated pCO2-levels
to cold-water corals and arctic coralline red algae and has thus in-depth understanding of both,
the relevant biogeochemical processes and the cultivation methods (see 3.1.2.iii).
Student contracts (HiWi): Student helpers are needed to support the extensive cultivation work as
well as for the routine analysis in the lab (e.g. water chemistry monitoring). They will also be
trained to assist the experiments and will help with sample preparation and processing. For
adequate assistance, two continuous contracts per long-term experiment will be necessary
.
Consumables:
Coral cultivation: The costs for coral cultivation are estimated to be
per year, including
mainly purchases for food and water treatment (such as filter materials and adsorbers). Additional
funds of
per year will be needed for culturing experiments (aquarium supplies, incubation
vessels, gas, sea salt). The costs for chemicals are estimated to be
per year. These costs
include purchases of gases, acids, enzyme inhibitors.
Travel: One international conference per year will be visited by the Postdoc in order to
disseminate the project results to the scientific community.
3.1.3
Personnel costs: Two PhD students will be needed owing to the complexity of the proposed
research projects. We envision one PhD student to focus on the bivalve and one on the
cephalopod model. Formally, one student will be based at IFM-GEOMAR, the other at the AWI.
However, work plans will be arranged in parallel, as to facilitate collaborations between the
students.
Consumables: For both model organisms, one sequencing run on the next generation sequencing
/ 454-platform (P. Rosenstiel, CAU Kiel) needs to be performed on gill tissue extracts to gain
164
sufficient amounts of transcript information to devise meaningful gene expression experiments.
One run will cost approximately
and will generate 200.000 sequences of 200-300 bp
length. Reagents for realtime PCR experiments / in situ hybridization need to be purchased
(approximately
). Significant funds will be used purchasing molecular biological
reagents (ca.
). Additional funds will be needed for culturing experiments
(aquarium supplies, incubation vessels, gas, sea salt), for Ussing-chamber experiments (ion
channel / transporter inhibitors, chemicals) and tissue acid-base analysis (chemicals, pH
electrodes).
Travel: Travel of the PhD students to an international conference in years two and three.
Additional costs for travel between the Institutions (AWI Bremerhaven, CAU Kiel / IFMGEOMAR) will be covered using the home institution’s funds.
3.1.4
Personnel costs: The experimental approach utilizes difficult and complicated techniques which
need a substantial training period. The supervision and training is provided by the PIs. A three
year contract guaranties a sustained and effective exploitation of the techniques by the PhD
student.
Consumables: All consumables covered by the given sum are necessary to perform the planned
experiments. The higher needs in second and third year are caused by molecular biological
experiments.
Travel: Travel of the PhD student and one supervisor to an international conference. Costsharing with home institution.
Investment: The lab is already equipped with a complete experimental setup, which, however,
has to be upgraded for microfluorimetric measurements. The availability of a separate setup for
microfluorimetric measurements is critical for the success of the project since about 50% of the
data will be produced using this equipment. Due to the high workload, time sharing at the
existing setup is not feasible.
vi. References
Allemand D, Ferrier-Pages C, Furla P, Houlbreque F, Puverel S, Reynaud S, Tambutte E, Tambutte S & Zoccola D (2004) Biomineralisation
in reef-building corals: from molecular mechanism to environmental control. C R Palevol 3: 453-467
Barth AS, Merk S, Arnoldi E, Zwermann L, Kloos P, Gebauer M, Steinmeyer K, Bleich M, Kaab S, Hinterseer M, Kartmann H, Kreuzer E,
Dugas M, Steinbeck G & Nabauer M (2005) Reprogramming of the human atrial transcriptome in permanent atrial fibrillation: expression
of a ventricular-like genomic signature. Circ Res 96: 1022-1029
Bleich M & Greger R (1997) Mechanism of action of diuretics. Kidney Int Suppl 59: S11-15
Bleich M, Schlatter E & Greger R (1990) The luminal K+ channel of the thick ascending limb of Henle's loop. Pflügers Arch 415: 449-460
Bleich M, Warth R, Schmidt-Hieber M, Schulz-Baldes A, Hasselblatt P, Fisch D, Berger S, Kunzelmann K, Kriz W, Schutz G & Greger R
(1999) Rescue of the mineralocorticoid receptor knock-out mouse. Pflügers Arch 438: 245-254
Eckerle LG, Lucassen M, Hirse T & Pörtner HO (2008) Cold induced changes of adenosine levels in common eelpout (Zoarces viviparus): a
role in modulating cytochrome c oxidase expression. J Exp Biol 211: 1262-1269
Furla P, Galgani I, Durand I & Allemand D (2000) Sources and mechanisms of inorganic carbon transport for coral calcification and
photosynthesis. J Exp Biol 203: 3445-3457
Gattuso JP, Allemand D & Frankignoulle M (1999) Photosynthesis and calcification at cellular, organismal and community levels in coral
reefs: A review on interactions and control by carbonate chemistry. Am Zool 39: 160-183
Gazeau F, Quiblier C, Jansen JM, Gattuso JP, Middelburg JJ & Heip CHR (2007) Impact of elevated CO2 on shellfish calcification. Geophys
Res Lett 34:
Guinotte JM, Orr J, Cairns S, Freiwald A, Morgan L & George R (2006) Will human-induced changes in seawater chemistry alter the
distribution of deep-sea scleractinian corals? Front Ecol Environ 4: 141-146
Heise K, Puntarulo S, Nikinmaa M, Lucassen M, Portner HO & Abele D (2006) Oxidative stress and HIF-1 DNA binding during stressful cold
exposure and recovery in the North Sea eelpout (Zoarces viviparus). Comp Biochem Physiol A Mol Integr Physiol 143: 494-503
Henry RP (1996) Multiple roles of carbonic anhydrase in cellular transport and metabolism. Ann Rev Physiol 58: 523-538
Hou J, Shan Q, Wang T, Gomes AS, Yan Q, Paul DL, Bleich M & Goodenough DA (2007) Transgenic RNAi depletion of claudin-16 and the
renal handling of magnesium. J Biol Chem 282: 17114-17122
165
Ishimatsu A, Hayashi M, Lee KS, Kikkawa T & Kita J (2005) Physiological effects on fishes in a high-CO2 world. J Geophys Res-Oceans 110:
Kosiek O, Busque SM, Foller M, Shcheynikov N, Kirchhoff P, Bleich M, Muallem S & Geibel JP (2007) SLC26A7 can function as a chlorideloading mechanism in parietal cells. Pflügers Arch 454: 989-998
Kurihara H & Shirayama Y (2004) Effects of increased atmospheric CO2 on sea urchin early development. Mar Ecol Prog Ser 274: 161-169
Langdon C, Broecker WS, Hammond DE, Glenn E, Fitzsimmons K, Nelson SG, Peng TH, Hajdas I & Bonani G (2003) Effect of elevated CO2
on the community metabolism of an experimental coral reef. Global Biogeochem Cycles 17:
Langdon C, Takahashi T, Sweeney C, Chipman D, Goddard J, Marubini F, Aceves H, Barnett H & Atkinson MJ (2000) Effect of calcium
carbonate saturation state on the calcification rate of an experimental coral reef. Global Biogeochem Cycles 14: 639-654
Langer G, Geisen M, Baumann KH, Klas J, Riebesell U, Thoms S & Young JR (2006) Species-specific responses of calcifying algae to
changing seawater carbonate chemistry. Geochem Geophys Geosys 7:
Lucassen M, Koschnick N, Eckerle LG & Pörtner HO (2006) Mitochondrial mechanisms of cold adaptation in cod (Gadus morhua L.)
populations from different climatic zones. J Exp Biol 209: 2462-2471
Lucassen M, Schmidt A, Eckerle LG & Pörtner HO (2003) Mitochondrial proliferation in the permanent vs. temporary cold: enzyme activities
and mRNA levels in Antarctic and temperate zoarcid fish. Am J Phys-Reg I 285: R1410-R1420
Marubini F, Ferrier-Pages C & Cuif JP (2003) Suppression of skeletal growth in scleractinian corals by decreasing ambient carbonate-ion
concentration: a cross-family comparison. Proc Biol Sci 270: 179-184
McConnaughey TA & Whelan JF (1997) Calcification generates protons for nutrient and bicarbonate uptake. Earth-Sci Reviews 42: 95-117
Medakovic D (2000) Carbonic anhydrase activity and biomineralization process in embryos, larvae and adult blue mussels Mytilus edulis L.
Helgoland Mar Res 54: 1-6
Melzner F, Bock C & Portner HO (2007a) Coordination between ventilatory pressure oscillations and venous return in the cephalopod Sepia
officinalis under control conditions, spontaneous exercise and recovery. J Comp Physiol [B] 177: 1-17
Melzner F, Bock C & Pörtner HO (2006a) Critical temperatures in the cephalopod Sepia officinalis investigated using in vivo
spectroscopy. J Exp Biol 209: 891-906
31
P NMR
Melzner F, Bock C & Pörtner HO (2006b) Temperature-dependent oxygen extraction from the ventilatory current and the costs of ventilation
in the cephalopod Sepia officinalis. J Comp Physiol [B] 176: 607-621
Melzner F, Bock C & Pörtner HO (2007b) Allometry of thermal limitation in the cephalopod Sepia officinalis. Comp Biochem Physiol A Mol
Integr Physiol 146: 149-154
Melzner F, Mark FC & Pörtner HO (2007c) Role of blood-oxygen transport in thermal tolerance of the cuttlefish, Sepia officinalis. Integr
Comp Biol 47: 645-655
Michaelidis B, Ouzounis C, Paleras A & Pörtner HO (2005) Effects of long-term moderate hypercapnia on acid-base balance and growth rate
in marine mussels Mytilus galloprovincialis. Mar Ecol Prog Ser 293: 109-118
Milliman JD (1993) Production and accumulation of calcium-carbonate in the ocean - budget of a nonsteady state. Global Biogeochem Cycles
7: 927-957
Piermarini PM, Choi I & Boron WF (2007) Cloning and characterization of an electrogenic Na/HCO3- cotransporter from the squid giant fiber
lobe. Am J Physiol Cell Physiol 292: C2032-2045
Pörtner HO (2002) Environmental and functional limits to muscular exercise and body size in marine invertebrate athletes. Comp Biochem
Physiol A Mol Integr Physiol 133: 303-321
Pörtner HO, Bock C & Reipschlager A (2000) Modulation of the cost of pHi regulation during metabolic depression: A
invertebrate (Sipunculus nudus) isolated muscle. J Exp Biol 203: 2417-2428
31
P-NMR study in
Pörtner HO, Boutilier RG, Tang Y & Toews DP (1990) Determination of intracellular pH and PCO2 after metabolic inhibition by fluoride and
nitrilotriacetic acid. Respir Physiol 81: 255-273
Pörtner HO, Langenbuch M & Michaelidis B (2005) Synergistic effects of temperature extremes, hypoxia, and increases in CO2 on marine
animals: From Earth history to global change. J Geophys Res-Oceans 110:
Pörtner HO, Mark FC & Bock C (2004) Oxygen limited thermal tolerance in fish? Answers obtained by nuclear magnetic resonance
techniques. Respir Physiol & Neurobiol 141: 243-260
Pörtner HO, Webber DM, O'Dor RK & Boutilier RG (1993) Metabolism and energetics in squid (Illex illecebrosus, Loligo pealei) during
muscular fatigue and recovery. Am J Physiol 265: R157-165
Poustka AJ, Kuhn A, Groth D, Weise V, Yaguchi S, Burke RD, Herwig R, Lehrach H & Panopoulou G (2007) A global view of gene
expression in lithium and zinc treated sea urchin embryos: new components of gene regulatory networks. Genome Biol 8: R85
Reuter S, Velic A, Edemir B, Schroter R, Pavenstadt H, Gabriels G, Bleich M & Schlatter E (2008) Protective role of NHE-3 inhibition in rat
renal transplantation undergoing acute rejection. Pflügers Arch:
Riebesell U, Zondervan I, Rost B, Tortell PD, Zeebe RE & Morel FM (2000) Reduced calcification of marine plankton in response to
Rost B, Riebesell U, Burkhardt S & Sultemeyer D (2003) Carbon acquisition of bloom-forming marine phytoplankton. Limnol Oceanogr 48:
55-67
Schneider K & Erez J (2006) The effect of carbonate chemistry on calcification and photosynthesis in the hermatypic coral Acropora
eurystoma. Limnol Oceanogr 51: 1284-1293
Schroeder BC, Waldegger S, Fehr S, Bleich M, Warth R, Greger R & Jentsch TJ (2000) A constitutively open potassium channel formed by
KCNQ1 and KCNE3. Nature 403: 196-199
Schulz KG, Riebesell U, Rost B, Thoms S & Zeebe RE (2006) Determination of the rate constants for the carbon dioxide to bicarbonate interconversion in pH-buffered seawater systems. Mar Chem 100: 53-65
Schulz KG, Rost B, Burkhardt S, Riebesell U, Thoms S & Wolf-Gladrow DA (2007) The effect of iron availability on the regulation of
inorganic carbon acquisition in the coccolithophore Emiliania huxleyi and the significance of cellular compartmentation for stable carbon
isotope fractionation. Geochim Cosmochim Ac 71: 5301-5312
Schulz KG, Zondervan I, Gerringa LJ, Timmermans KR, Veldhuis MJ & Riebesell U (2004) Effect of trace metal availability on
coccolithophorid calcification. Nature 430: 673-676
Sodergren E, Weinstock GM, Davidson EH, Cameron RA, Gibbs RA, Angerer RC, Angerer LM, Arnone MI, Burgess DR, Burke RD,
Coffman JA, Dean M, Elphick MR, Ettensohn CA, Foltz KR, Hamdoun A, Hynes RO, Klein WH, Marzluff W, McClay DR, Morris RL,
Mushegian A, Rast JP, Smith LC, Thorndyke MC, Vacquier VD, Wessel GM, Wray G, Zhang L, Elsik CG, Ermolaeva O, Hlavina W,
Hofmann G, Kitts P, Landrum MJ, Mackey AJ, Maglott D, Panopoulou G, Poustka AJ, Pruitt K, Sapojnikov V, Song X, Souvorov A,
166
Solovyev V, Wei Z, Whittaker CA, Worley K, Durbin KJ, Shen Y, Fedrigo O, Garfield D, Haygood R, Primus A, Satija R, Severson T,
Gonzalez-Garay ML, Jackson AR, Milosavljevic A, Tong M, Killian CE, Livingston BT, Wilt FH, Adams N, Belle R, Carbonneau S,
Cheung R, Cormier P, Cosson B, Croce J, Fernandez-Guerra A, Geneviere AM, Goel M, Kelkar H, Morales J, Mulner-Lorillon O,
Robertson AJ, Goldstone JV, Cole B, Epel D, Gold B, Hahn ME, Howard-Ashby M, Scally M, Stegeman JJ, Allgood EL, Cool J, Judkins
KM, McCafferty SS, Musante AM, Obar RA, Rawson AP, Rossetti BJ, Gibbons IR, Hoffman MP, Leone A, Istrail S, Materna SC,
Samanta MP, Stolc V, Tongprasit W, Tu Q, Bergeron KF, Brandhorst BP, Whittle J, Berney K, Bottjer DJ, Calestani C, Peterson K, Chow
E, Yuan QA, Elhaik E, Graur D, Reese JT, Bosdet I, Heesun S, Marra MA, Schein J, Anderson MK, Brockton V, Buckley KM, Cohen
AH, Fugmann SD, Hibino T, Loza-Coll M, Majeske AJ, Messier C, Nair SV, Pancer Z, Terwilliger DP, Agca C, Arboleda E, Chen N,
Churcher AM, Hallbook F, Humphrey GW, Idris MM, Kiyama T, Liang S, Mellott D, Mu X, Murray G, Olinski RP, Raible F, Rowe M,
Taylor JS, Tessmar-Raible K, Wang D, Wilson KH, Yaguchi S, Gaasterland T, Galindo BE, Gunaratne HJ, Juliano C, Kinukawa M, Moy
GW, Neill AT, Nomura M, Raisch M, Reade A, Roux MM, Song JL, Su YH, Townley IK, Voronina E, Wong JL, Amore G, Branno M,
Brown ER, Cavalieri V, Duboc V, Duloquin L, Flytzanis C, Gache C, Lapraz F, Lepage T, Locascio A, Martinez P, Matassi G, Matranga
V, Range R, Rizzo F, Rottinger E, Beane W, Bradham C, Byrum C, Glenn T, Hussain S, Manning G, Miranda E, Thomason R, Walton K,
Wikramanayke A, Wu SY, Xu R, Brown CT, Chen L, Gray RF, Lee PY, Nam J, Oliveri P, Smith J, Muzny D, Bell S, Chacko J, Cree A,
Curry S, Davis C, Dinh H, Dugan-Rocha S, Fowler J, Gill R, Hamilton C, Hernandez J, Hines S, Hume J, Jackson L, Jolivet A, Kovar C,
Lee S, Lewis L, Miner G, Morgan M, Nazareth LV, Okwuonu G, Parker D, Pu LL, Thorn R & Wright R (2006) The genome of the sea
urchin Strongylocentrotus purpuratus. Science 314: 941-952
Tambutte R, Allemand D, Mueller E & Jaubert J (1996) A compartmental approach to the mechanism of calcification in hermatypic corals. J
Exp Biol 199: 1029-1041
Tambutte S, Tambutte E, Zoccola D, Caminiti N, Lotto S, Moya A, Allemand D & Adkins J (2007) Characterization and role of carbonic
anhydrase in the calcification process of the azooxanthellate coral Tubastrea aurea. Mar Biol 151: 71-83
Waldegger S, Fakler B, Bleich M, Barth P, Hopf A, Schulte U, Busch AE, Aller SG, Forrest JN, Jr., Greger R & Lang F (1999) Molecular and
functional characterization of s-KCNQ1 potassium channel from rectal gland of Squalus acanthias. Pflügers Arch 437: 298-304
Zoccola D, Tambutte E, Kulhanek E, Puverel S, Scimeca JC, Allemand D & Tambutte S (2004) Molecular cloning and localization of a
PMCA P-type calcium ATPase from the coral Stylophora pistillata. Biochim Biophys Acta 1663: 117-126
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Project 3.2: Calcification under pH-stress: Impacts on ecosystem and organismal
levels.
(PI: Ralph Tollrian)
i. Objectives
Anthropogenic CO2 emissions are reducing ocean pH-values and carbonate saturation state, with
strongest effects in high-latitude surface waters. In this project we will compare the responses of
organisms from pteropods, scleractinian corals and red coralline algae, which have been
identified as key groups for the understanding of community responses of ocean acidification
(Kleypas et al. 2006). These calcifying organisms are vulnerable to acidification and because of
their relevant role in their ecosystems these impacts have the potential to cause cascading effects
and to alter ecosystem stability and functioning. Pteropods dominate zooplankton communities in
polar regions of both hemispheres, scleractinian corals form the physical structure of the tropical
reef ecosystems and coralline red algae are cosmopolite in the ocean and found at all depths
within the photic zones. The thresholds and tipping-points where the different organisms and
systems respond to acidification, the magnitude of the response and the impact on the
communities will be determined for different ontogenetic stages.
Subsaturation with respect to aragonite, the metastable form of calcium carbonate, could begin in
the Southern Ocean and in sub-Arctic regions before the end of this century (Orr et al. 2005). In
these cold regions, the only pelagic aragonite-producers are shelled pteropods, e.g. the bipolar
species Limacina helicina. These shelled pteropods will be among the first organisms
experiencing carbonate saturation states <1 within their actual geographical ranges. We
hypothesize that ocean acidification will have pronounced effects on the calcification,
dissolution, and metabolism of pteropods. Despite their crucial role and high sensitivity to
changes in seawater chemistry, pteropods have been poorly studied so far. Shell dissolution was
observed to occur in live subantarctic pteropods exposed to water undersaturated in aragonite for
48 h (Orr et al. 2005). In the Subarctic Pacific pteropod Clio pyramidata net calcification rates
decreased progressively with time during a 48 h incubation in closed 1 litre jars, as the aragonite
saturation state of seawater declined owing to respiratory CO2 increase (Fabry et al. 2008). These
authors also report a decrease in oxygen consumption rates at elevated CO2 for Limacina helicina
antarctica Systematic studies on the calcification and other metabolic responses of pteropods
under conditions of well-controlled carbonate chemistry and in combination rising temperatures
have not been carried out as yet and are therefore urgently called for (Kleypas et al. 2006).
Beside the vulnerability of subpolar regions also tropical coral reef ecosystems are globally
threatened by a twofold effect of the increasing CO2 concentration in the atmosphere: Global
warming as a major cause of coral bleaching and ocean acidification. Elevated pCO2 is causing
the calcium carbonate saturation horizon to shoal, even in low-latitude and coastal seas with
suboxic zones (Fabry et al. 2008). In the deep semi-enclosed basins of South-East Asia, including
the Andaman Sea, Celebes Sea, and Sulu Sea, the pycnocline separating the aragonite
supersaturated surface waters from the suboxic low aragonite deep waters undergoes very large
oscillations exceeding 60 m amplitude – on a tidal basis (Jackson 2004). The occurrence of
solibores resulting from the breaking of these waves provides a unique and so far unexplored
opportunity to investigate the effect of spatio-temporal changes in pCO2 on coral calcification in
168
the field. Although it is generally assumed that corals are negatively affected at a certain level of
pCO2 (Hoegh-Guldberg et al. 2007), differential effects in corals (Lough and Barnes 2000) and
other calcifiers (Iglesias-Rodriguez et al. 2008, Fabry et al. 2008) indicate that the picture may be
much more complicated than previously thought. Of particular interest is how different
ontogenetic stages respond to the same stress factor. Possibly, recruitment will be influenced
before an effect on growth of adult colonies can be detected. In scleractinian corals, settling of
larvae and early colony formation are critical steps where a reduced calcification will threaten the
recruit because it lives in competition with overgrowing turf algae, is preyed on by predators and
is exposed to herbivores which feed on algae and scratch off the fragile young colonies. But also
coral community compositions may change as a consequence of increasing pCO2. Corals have
two main modes of larval production: “brooding”, where larvae develop within the colony and
are released at a relatively large size, or “broadcast spawning”, where eggs and sperm are
released simultaneously and larvae develop in the water column. The larger larvae of brooding
species possibly will have an advantage under decreased ph-conditions because they contain
more reserves, which allow faster initial growth.
The health of reefs critically depends on recruitment, especially when reefs are devastated by
coral bleaching or storms. Coral reefs have been shown to possess two alternative stable states
with coral- or algae-dominated climax conditions and with a limited ability to return into a coral
dominated state once the tipping point has been exceeded (Mumby et al. 2006). Ocean
acidification is supposed to influence corals negatively, while macroalgae might even benefit
from higher CO2-levels. Additionally coralline red algae may decline with a negative effect on
coral recruitment. Thus, ocean acidification has the potential to influence survival of early stages,
change community compositions and processes and on a higher level, lead to phase shifts in
whole ecosystems and alter ecosystem functioning and stability.
Despite the broad abundance of coralline red algae so far little is known about their calcification
mechanisms. Recently some studies focussed on the use of coralline red algae as environmental
recorders (Halfar et al. 2007, Kamenos et al. 2007), showing the strong response on
environmental changes, displayed in systematic variations in the chemical composition of the
precipitated carbonates (high magnesium calcite). This observation suggests that coralline red
algae are promising candidates to study the responses of important calcifiers on ocean
acidification in a wide range of habitats.
In a recent short (7 weeks) mesocosm experiment study (Kuffner et al. 2008) at low latitudes, the
recruitment rate and growth of red algae was shown to be severely inhibited at elevated pCO2. No
research was however undertaken to determine whether changes in the carbonate structure or
microchemistry or adaptations occurred over a longer period (i.e. over an annual growth cycle).
Such changes in the chemical composition would clearly affect the solubility of the carbonates
formed by those algae. So far no study was done addressing pH thresholds for reproduction and
growth/calcification of coralline red algae or possible physiological responses to deal with
decreased ambient pCO2 levels.
Finally, solubility changes due to chemical modifications of the carbonates directly affect the pH
buffering capacities of shelf regions where coralline red algae contribute significantly to the
carbonates deposited in the sediments.
3.2.1 The group of U. Riebesell has been among the first to study direct effects of CO2-induced
seawater acidification (Riebesell et al. 1993, Riebesell et al. 2000). Presently, research in his
169
group addresses a variety of OA-related aspects from the cellular to the community level
(Riebesell 2004, Riebesell et al. 2007). This includes studies on the CO2/pH sensitivity of a
variety of marine calcifying groups, ranging from coccolithophores and rhodoliths to bivalves,
and cold water corals. In preparation for the proposed work on pteropods calcification in
BIOACID, a pre-study is being carried out with Limacina helicina in Ny Alesund, Svalbard, in
May/June 2008. The group of I. Werner has expertise in cold-life culturing and experimental
approaches on measuring metabolic rates such as respiration, grazing, and predation of Arctic
marine invertebrates (Werner et al. 2002, Werner and Auel 2005). Of direct relevance to the
proposed project are previous studies on the seasonal dynamics of the Arctic shelled pteropod
species Limacina helicina in epipelagic, partly ice-covered waters in the Fram Strait area and
Kongsfjorden (Werner 2006).
3.2.2 The research team has long experience in raising coral larvae. R. Tollrian had successfully
established coral reef research systems in Munich and conducted settlement and growth
experiments with coral larvae (Petersen and Tollrian 1991) and population genetic studies in
corals (Maier et al. 2001, 2005). This work provided the tools for the SECORE (Sexual coral
reproduction) initiative. D. Petersen initiated the SECORE program and is a leading expert in,
and developed methods for, breeding and raising corals in closed aquaria systems (e.g., Petersen
et al. 2006). E. Grieshaber. and W. Schmahl are experts in crystallography. Their methods will be
applied to studies of the coral skeleton structure. A. Eisenhauer is the head of the isotope
geochemistry lab at the IFM-GEOMAR. The work proposed here is interdisciplinary and
involves biology, biodiversity, geo-chemistry, mineralogy, crystallography and material sciences.
3.2.3 The group of C. Richter has been working on coral reef ecology over the last 12 years,
with a regional focus on the Red Sea and South-East-Asia (Indonesia, Thailand and China).
Major findings using novel endoscopic tools were the discovery of biomass-rich and diverse
communities of sponges in Red Sea coral reef crevices fuelling a significant part of reef
production (Richter et al. 2001, Richter and Abu-Hilal 2006). The group also contributed to a
seminal paper on the interaction of currents and plankton explaining the enrichment of plankton
near coral reefs (Genin et al. 2005). Ongoing research addresses the effect of solibores on coral
communities of the Similan Islands in the Andaman Sea. The findings of significant differences
in coral cover and growth between soliton-exposed and soliton-protected communities, and the
development of a micro-CaveCam form the scientific and technological basis of the present
proposal. A complementary project on the supply and early recruitment of corals has recently
been approved within the EU-ITN CalMarO.
3.2.4 J. Fietzke (IFM-GEOMAR) has a strong research record in developing and applying mass
spectrometric techniques for trace and ultra-trace elements (e.g. Fietzke et al., 2004, 2006, in
press). His work focused recently on issues of biomineralisation with marine biogenic carbonates
(e.g. Heinemann et al. 2008, Rüggeberg et al. 2008). The research group has a strong background
in carrying out microanalytical studies in biogenic carbonates. T. Hansteen is a specialist in
microanalytical techniques such as Synchrotron X-ray Fluorescence microprobe (SYXRF;
Hansteen et al. 2000), electron microprobe (EMP), NanoSIMS (e.g. Zumholz et al. 2007 a, b),
and has co-supervised a PhD in Marine Ecology (K. Zumholz; Zumholz et al. 2006, 2007c).
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iv. Work Programmes, Schedules, and Milestones
3.2.1 Impact of ocean acidification and warming on sub-polar shelled pteropods
(U. Riebesell)
The aim of this project is to study the synergistic effects of ocean acidification and rising
seawater temperatures on a key component of polar and sub-polar epipelagic communities, the
shelled pteropods. In a second step, ecological consequences of these effects for different lifestages will be assessed. This will be achieved through a collaboration involving experimental
marine biology, physiology, biochemistry and marine ecology. Specifically, the project will
address the following aspects:
1. Calcification/dissolution response
2. Metabolic response (oxygen consumption, swimming performance)
3. Life cycle responses (egg and larval development, growth and reproduction)
4. Ecosystem response (feeding rates, predation, survival)
Pteropods of the dominant and bipolar genus Limacina (L. helicina or L. retroversa) will be
collected in Arctic waters (Kongsfjord, Svalbard) and will be reared in the Kings Bay Marine Lab
under ambient conditions. Pteropods will be held in specially designed, V-shaped aquaria
equipped with a circulating current system and filled with filtered, cold seawater. Food will be
provided in form of algal cultures. Ambient pH/pCO2 and temperature will be altered in
temperature-controlled incubation rooms equipped with a new gas-producing system.
Methodologies/approaches for aspects 1-4 include:
1. Calcification rate: 45Ca incubations, shell weight, fluorochroms; shell composition:
Mg/Ca, Sr/Ca, possibly boron isotopes; shell dissolution: SEM
2. Respirometer measurements, digital camera system
3./4. If culturing attempts turn out successful, life cycle experiments and grazing experiments
of low and high CO2 exposed Limacina will be started in year two of the project. Grazing
experiments will be conducted with a major predator, the gymnosomate pteropod Clione
limacina, which was successfully cultivated previously.
Links to other projects: The pteropod work in this sub-project will be closely linked to studies
on mollusc larvae (3.5.1) and mollusc phyiology (3.1.3). Results of this sub-project with
relevance to the ballast effect will also feed into sub-projects 1.2.5. Estimates of OA-induced
changes in pteropod calcification will be provided to sub-projects 1.3 and 5.2. Samples of
pteropod shells will be provided to sub-project 3.2.4 for SEM and electron backscatter diffraction
(EBSD) analysis of the skeletal architecture as well as high-resolution geochemical analyses
applying electron microprobe, NanoSIMS, LA-ICP-MS, SYXRF.
Schedule
3.2.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Design & construction of experimental
set-ups for field- and lab-based studies
Pteropod CO2 perturbation experiments
at field station Ny Alesund
171
3.2.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Attempts to culture pteropods in IFMGEOMAR culturing facilities
CO2 perturbation exps. in IFMGEOMAR culturing facilities
Combined CO2 and temperature
perturbation exps. at Ny Alesund
Follow-up exps. in IFM-GEOMAR
culturing facilities
data interpretation
Milestones (3.2.1)
- Experimental facilities implemented
month 06
- Data set on CO2/pH sensitivity of adult pteropods
month 12
- First practice in culturing of juvenile and adult pteropods
month 21
- Data set on synergistic effects of CO2 and temperature
month 24
month 33
3.2.2 Impact of ocean acidification on reproduction, recruitment and growth of
scleractinian corals (R. Tollrian).
Our main goals are: 1) Test survival of coral larvae, early colony establishment and growth of
different species from several geographic regions, to find species specific thresholds and
geographic differences. 2) Assess competitive ability against algae. 3) Test the hypotheses that
brooding species will be less influenced compared to broadcast spawning species. 4) Test for
differences in reproductive investment and growth in brooding corals under different ph
conditions to assess sublethal stress effects. 5) Compare amount and quality of calcite formation
of fragments under different ph-conditions and quantify changes in ultrastructure,
biogeochemistry and material properties.
Aim 1) The SECORE (sexual coral reproduction) working group has successfully
established a recruitment program for coral conservation where larvae are collected during mass
spawning events in the Caribbean, transported to public and research aquaria and raised in these
closed systems (Petersen et al. 2006). We will be able to obtain larvae from broadcast spawning
species from different locations during the highly predictable yearly spawning events: a)
Maldives (March), b) Okinawa/Japan (May), c) Curacao/Caribbean (August/September), d)
Mozambique (October), e) Great Barrier Reef (December). Experiments studying fertilisation,
larval survival and early colony formation will be conducted during field studies. For long-term
lab studies, larvae will be collected and transported to our aquaria facilities following an
improved method (Petersen et al. 2005a). Larvae will not settle in filtered seawater because they
172
recognize suitable substrates based on relevant biofilms. Specifically designed ceramic tiles will
be offered as settlement substrate (Petersen et al. 2005b). The larvae will be raised under five
different pH conditions in closed aquaria systems. Each treatment will be fully replicated. We
will measure larval survival, settlement, transformation and early colony growth and will identify
threshold conditions. A share of the larvae will be settled according to the SECORE protocol
under standard conditions and experiments with different pH treatments will be started after the
young colonies have been established. This experiment will allow separating OA effects on larval
survival from effects on juvenile growth. We will compare the species composition of successful
settlers in the different treatments to test whether species differ in their tolerance to low pH
conditions. By repeating the experiments with larvae from different locations we will be able to
cover a wide range of different species and to compare thresholds within and between geographic
regions.
Aim 2) The competition with algae can be tested with the same experimental design but
with different amounts of turf algae in the systems.
Aim 3) The brooding coral species Pocillopora damicornis and Favia fragum are
available in the aquaria facilities in Rotterdam and Munich where they reproduce in closed
systems. Colonies and larvae can be obtained from Rotterdam. The experiments above will be
repeated with larvae from brooding species to test for different thresholds and differences in
vulnerability to lower pH levels.
Aim 4) Sublethal effects on adult colonies may be measurable in reduced energy
availability for growth, reproduction or both. To test for sublethal effects, adult colonies of P.
damicornis and F. fragum will be raised in aquaria with different ph-levels to measure
investments into reproduction and growth. Colonies of F. fragum are ideal for this purpose,
because they reach maturity extremely fast, after just one year.
Aim 5) Another mode of recruitment in corals is fragmentation. Fragments from
individuals can be exposed to different treatments and will allow to test the effects of OA on
growth without confounding effects of genotypic variation. We will use fragments of fast
growing, branching Acropora species for the experiment (larvae of the same species will be used
in the experiments with sexually produced larvae). The experiment will run 12 month but can be
prolonged if necessary. Within 12 month the branches will grow up to 10 cm under good
conditions. This growth should provide enough new skeleton material, even under low pH
conditions, for isotope analysis and analysis of material properties. Additionally we will analyse
material of the corals raised from larvae. Microsensor technology will allow measuring Ca2+, CO2
and CO32- in the cell regions where calcification takes place. These analyses should reveal
whether corals sacrifice growth rate, skeleton stability or both under stress conditions
Links to other projects:
2.1.1: In this project responses of early life stages of different organisms to acidification will be
analysed. In collaboration we will study responses of coral eggs to trace mechanisms of
acidification effects on the earliest ontogenetic stages. Our study will provide additional
information for theme 2 (Performance characters: reproduction, growth and behaviours in animal
species). 3.1.2: We will compare calcification mechanisms and responses to acidification
between tropical and cold water corals. 3.2.3: Our project with a focus on early live stages of
corals directly complements project 3.2.3 where the focus is on responses of corals in nature.
3.2.4: The measurement of minor and trace elements (e.g. Sr, Mg, Ba, S) at micron scale
resolution using electron microprobe (EMP) and LA-ICP-MS provides a tool to analyze changes
173
in chemistry within carbonate material such as coral skeletons and will be done in conjunction
with project 3.2.4. We will deliver coralline red algae from our coral incubation experiments.
3.3.2: For the analysis of the biogenic carbon composition we will collaborate with A. Eisenhauer
and will apply his isotope techniques. 3.4.2: For the analyses of the cellular mechanisms of
calcification under different ph-levels we will collaborate with D. de Beer and will apply his
microsensor techniques. 4.1.2: this project will study effects on larvae of selected organisms and
results will be compared to our results on coral larvae. 4.1.3: Our study includes competition
experiments between early coral colonies and macroalgae. These results will be compared with
the results on tropical macroalgae of project 4.1.3 and will provide relevant information about
regime-shifts for theme 4 (Species interactions and community structure: will ocean acidification
cause regime shifts?).
Schedule
3.2.2
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Design & construction of experimental
set-ups for field- and lab-based studies
Experiments with brooding corals in
Rotterdam and transport to Bochum
Experiments with coral larvae from
spawning species
Competition experiments between
coral larvae and macroalgae
Field study during coral spawning
Analysis of reproductive investment of
adult brooding corals
Microsensor analysis
Comparative analysis of coral
skeletons
data interpretation
Milestones (3.2.2)
- Experimental facilities implemented
month 06
- Data set on CO2/pH sensitivity of larvae from brooding corals
month 12
- Data set on CO2/pH sensitivity of larvae from spawning corals
month 21
- Data set on CO2/pH sensitivity of adult brooding corals
month 24
- Microsensor analysis of calcification in corals grown at different CO2/pH
month 28
- Analysis of ultrastructure, crystallography and material properties of corals grown
at different CO2/pH
month 33
174
3.2.3 Coral calcification in marginal reefs (C. Richter)
The aim of this project is to study the effect of natural oscillations of the aragonite saturation
horizon, via the shoaling and breaking solitary internal waves, on coral calcification in marginal
reefs. Marginal reefs mark the transition between flourishing coral reefs on the one hand, and
rocky bottom devoid of macroskeletal organisms on the other. Such reefs occur under conditions
where calcium carbonate accretion balances corrosion/erosion. In the Similan Islands, in the
Andaman Sea of Thailand, the transition from reef to rock occurs over scales of only 100 m,
between the solibore-protected East and the solibore-exposed West sides of the island. Marginal
reefs at the flexion point provide a unique opportunity to assess tipping points in aragonite
saturation state with regard to net coral calcification, and to explore the synergies between
aragonite saturation, and other parameters (e.g. temperature, nutrients) associated with solibores.
Specifically, the project will address calcification/decalcification and photosynthesis/respiration
of corals using in situ microsensors and in situ chambers (Funke chamber). The work will involve
three Steps: (1) technological development, (2) testing/deployment instrumentation in the field
and (3) field measurements.
As aragonite saturation co-varies with temperature, the fine-scale vertical and horizontal
temperature field will be monitored with an array of Tidbit temperature loggers (Onset comp.
Corporation). Two moored multiparameter-loggers (T, S, O2, pH, Fluorescence, OBS), a Rapid
Access Sampler, RAS (McLane Inc.) and ZPS (McLane Inc.) available at AWI will allow
autonomous pre-programmed sampling of alkalinity and nutrients. A novel in situ respirometer
developed by the group (Funke chamber) combined with RAS will be used to measure timeseries of photosynthesis, respiration and calcification in situ, where calcification will be measured
using the alkalinity anomaly technique. Oxygen, calcium, pH and novel CO32- microsensortechniques developed at MPIMM (D. de Beer) will be combined with the micro-CaveCam
developed by the AWI group to measure photosynthesis and calcification in the immediate
vicinity of corals along a spatio- temporal gradient of aragonite saturation state.
Research will concentrate on the massive scleractinian coral Porites lutea, but to assess the
general validity of our findings we will investigate also other abundant genera, including
Acropora and Pocillopora, as well as purely heterotrophic forms such as Tubastraea micranthus.
To test the equipment and laboratory runs, coral specimens will be collected in Koh Racha island
off Phuket and reared in the Phuket Marine Biologial Laboratory (PMBC) under ambient
conditions in a through flow system. Variable pH/pCO2 and temperature will be supplied in
temperature-controlled incubators equipped with a CO2 bubbling system developed according to
the blue-print of the Bioacid consortium. The analytical techniques and calculations for the
incubation experiments follow Schneider and Erez (2006), for the microsensor set-up cf. Weber
et al. (2007).
Methodology of combining micro-CaveCam and microsensors will be carried out in close
cooperation with the microsensor group of D. de Beer (MPIMM) (project 3.4.2). Field work will
be coordinated with the other teams working in tropical waters, including R. Tollrian (project
3.2.2.) and K. Bischof (project 4.1.3.), using common field stations as a logistic basis. High
resolution records of pH (boron isotopes), and nutrients (P:Ca) in relation to linear growth and
175
skeletal density of Porites skeletons will be examined with J. Fietzke (project 3.2.4.) and A.
Eisenhauer (project 3.3.2.). Results of this project will have important ramifications for the
membrane studies in coldwater corals by A. Form (project 3.1.2), and provide baseline data for
theme 2 and theme 4.
Schedule
3.2.3
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Recruitment of staff, technological
development
Collection of corals and laboratory test
runs
In situ microsensor and incubation
experiments
Data analysis
Milestones (3.2.3)
- In situ instrumentation developed and tested
month 06
- Laboratory data set on CO2/pH sensitivity of corals
month 12
- First field data set on CO2/pH sensitivity of corals (end NE monsoon)
month 18
- Second field data set on CO2/pH sensitivity of corals (end SW monsoon)
month 24
- Data set on synergistic effects of CO2, temperature and nutrients
month 24
- Data analysis, submission of theses and manuscripts
month 36
3.2.4 Impact of ocean acidification on coralline red algae (J. Fietzke)
Coralline red algae are important carbonate forming species both at low and high latitudes. In the
former, they are important for the development and stability of the coral reefs. In the latter, they
are the dominant benthic marine carbonates. This project therefore will investigate the response
of different species of coralline red algae in these two contrasting environmental settings to ocean
acidification. In particular, this will include:
1. Culturing experiments with CO2 perturbations using a high-latitude species
2. Micro-scale skeletal architecture and geochemical analyses to compare cultured with insitu specimens.
3. Comparison of the response of coralline red algae and corals from the same location at
low latitudes, which has a high natural pH variability (linked to project 3.2.3).
4. Investigation of the responses in long-lived coralline red algae to the anthropogenic
driven CO2 increase since the beginning of the industrial revolution.
176
5. Comparison between different pH responses of macroalgae and macroorganisms (linked
to project 2.1.3; 4.1.1; 4.1.3).
To achieve these objectives, Lithothamnium topiphorme
will be cultured from existing specimens available at
IFM-GEOMAR. The samples will be grown for 12-14
months, using the five standard pH levels within the
BIOACID project. This will be done in conjunction with
U. Riebesell and A. Form (project 3.2.1 and 3.1.2
respectively). The light irradiance and temperature values
used would be similar to those occurring at the collection
site. In-situ analyses of the calcification process will be
carried out (in conjunction with project 3.4.2). The Electron microprobe map of sulphur
skeletal architecture of naturally grown sample material variation in coralline red algae
will be analysed using SEM, electron backscatter
diffraction (EBSD). This will be complemented by high-resolution geochemical analyses
applying electron microprobe, NanoSIMS, LA-ICP-MS, SYXRF. After the completion of the
culturing, this sample material will be subjected to the same analytical tools, to investigate the
effects of the pH treatment. The architectural and geochemical analyses will be available for
other cultured material (project 3.2.1 and 3.1.2) to understand how different calcifying organisms
react to different pH.
To further understand existing adaption to natural pH variability within coral and red algae
communities in tropical reefs, samples from both taxa, grown in close proximity, will be used for
geochemical comparison. This material and accompanying temperature and pH data will be made
available through collaboration with project 3.2.3. Architectural and geochemical analyses of
new coral and coralline red algae growth will be carried out (on material from project 3.2.2) to
determine how changes in pH are affecting newly establishing colonies.
Finally, using samples of long-lived species e.g. Clathromorphum nereostratum, we will study
whether ocean acidification since the industrial revolution has impacted on the micro-architecture
and geochemistry of coralline algae over decadal timescales. Thus, the rate of adaptation and
changes from short-term studies can be compared to changes over extended timescales in order to
predict the long-term effects of progressing ocean acidification.
3.2.3: Coral and coralline red algae samples from the same habitat will be compared with respect
to their architectural and microchemical responses on variable pH and temperature conditions. A
comparable approach focussing on cultured specimen from pH controlled coral larvae
recruitment experiments will be used in collaboration with 3.2.2. Both collaborations (3.2.2 and
3.2.3) will contribute to the understanding the pH influence on the initial growth of coral
colonies. Experience of U. Riebesell and A. Form (project 3.2.1 and 3.1.2 respectively) will be
utilised in culturing the coralline red algae. In return our experience of geochemical and
architectural analyses will be utilised to understand the effects of pH on pteropod and cold water
coral samples from this two projects. 4.1.1 and 4.1.3: Joint experiments on the calcification
processes in coralline red algae and other macroalgae will be undertaken focussing on the
competition between different algae groups. 3.4.2: Joint experiments to study the calcification
process on a cellular level will be carried out using microsensor techniques. T. Brey (project
2.1.3) will provide time-series data for the coralline red algae samples from Svalbard, Norway.
177
Schedule
3.2.4
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Set-up of culturing facility
CO2 perturbation experiment
Analyses of skeletal architecture from
in-situ material
Geochemical analyses of in-situ
material (incl. archive material &
corals)
Analyses of skeletal architecture from
cultured material
Geochemical analyses of cultured
material
data interpretation
Milestones (3.2.4)
- Experimental set-up for CO2/pH implemented
month 03
- Baseline skeletal architecture and microchemistry
month 09
- Geochemical comparison between coexisting red algae and corals
month 12
- Long-term data set on centennial archive
month 15
- Collection of cultured samples
month 15
- Skeletal architecture and microchemistry of cultured samples
month 21
- Data set on CO2/pH sensitivity of cultured samples
month 24
- Completion of manuscripts
month 30
First Year
Personnel costs
3.2.1
3.2.2
3.2.3
3.2.4
Subtotal
Consumables
178
Second Year
Third Year
Total
First Year
Second Year
Third Year
Total
3.2.1
3.2.2
3.2.3
3.2.4
Subtotal
Travel
3.2.1
3.2.2
3.2.3
3.2.4
Subtotal
Investments
3.2.1
3.2.2
3.2.3
3.2.4
Subtotal
Other costs
3.2.1
3.2.2
3.2.3
3.2.4
Subtotal
TOTAL
3.2.1
Personnel costs: 1 Ph.D. position (Jan Büdenbender, per year), student helpers (€ per year) to
assist with Svalbard experiments and lab culturing of pteropods and food algae
Consumables: synthetic gases, Ca isotopes, Mg,, Sr and Ca analyses, fluorochromes, SEM
analyses, reagents for artificial media, nutrient and Winkler analyses (€ in years 1 and 2)
179
Travel: 2 trips for 3 persons (Ph.D. student + 2 student helpers) to Ny Alesund, Svalbard each),
meals and accommodation /person/day) and lab fee ( /person/day) at field station Ny Alesund
for 2 visits of 28 days each for 3 persons: total , 2 trips for 2 persons (Ph.D. student + Co-PI) per
year to international conferences), attendance of BIOACID workshops and annual meetings.
Investments: Culturing facilities for pteropod maintenance (€), 12 experimental laminar-flow
incubation vessels for CO2/temperature controlled incubations (x)
Other costs: transport of equipment to Svalbard (€).
3.2.2
Personnel costs: 1 Ph.D. position (Sebastian Striewski, per year), student helpers (per year for
year 1 and 2, for year 3) to assist with field experiments, lab experiments, calibration and
control of the CO2-supply systems and chemical parameters.
Consumables: synthetic gases, isotope analyses, ultrastructure analyses, reagents for artificial
sea water, supply for culturing facility, nutrient and Winkler analyses, settling substrates for coral
larvae (€ in year 1, in year 2, in year 3)
Travel: 2 trips for 3 persons (PI, Ph.D. student + student helper) for field experiments to Lizard
Island (Australia) ( each + bench fees and accommodation for 35 days: AUD 113/person and
day), 3 short trips for 2 persons to field stations in different geographic regions for larvae
collections after mass spawning events, 2 trips for 2 persons (Ph.D. student + PI) for year 2 and 3
to international conferences ( each), attendance of BIOACID workshops and annual meetings (€
per year). Attendance of microsensor workshop, collaborative works in Rotterdam, Kiel and
Munich for Ph.D.. Total: € .
Investments: Culturing facilities for coral larvae and adult colonies. Separate systems for each
CO2-level, including tanks, pumps, lamps, water purification, systems for CO2- and temperaturecontrolled incubations (total € ).
Other costs: Coral larvae rearing tanks with circular flow, mobile CO2- and temperature-control
systems for the field (total € ). Air freight (year 1 and year 2; total € ) total €
3.2.3
Personnel costs: 1 Ph.D. position (N.N., € per year) is needed to carry out the combined microCaveCam-microsensor work in situ and in the lab, on natural and simulated in situ variations of
aragonite saturation state on coral calcification in marginal reefs. The student is to be trained in
the BIOACID consortium using the expertise available in the various institutions, e.g. in
methodology (CO2-experiments, IFM-GEOMAR) and instrumentation (microsensors, MPIMM).
Student helpers (6 mo. x 80 h, € per year) are needed to assist with Thailand experiments and
chemical analyses of samples.
Consumables: microsensor materials, reagents for chemical analyses, glass- and plastic ware for
coral maintenance, CO2 gases (€ in years 1 and 2; € in year 3)
Travel: 1 flight (year 1) and 2 flights (year 2), each for 3 persons (co-PI, Ph.D. student, 1 student
helper) to Phuket, Thailand (€ and € ); contribution to Accommodation/Living expenses (€
/person/day for 3 visits á 20 days for 3 persons); attendance of BIOACID workshops and annual
meetings (€ per year, total € );
180
Investments: Microsensor set-up (€ ; year 1); in-situ respirometer (custom-built, € , year 1)
Other costs: Transfers Phuket-Koh Racha/Similans (shared cost € /boat/day for 3 visits á 10
days; total € ), Air freight (year 1 and year 3; total € ).
3.2.4
Personnel costs: Postdoc for 30 months (Laura Foster, € per year). In-depth experience of
skeletal architecture and geochemical analyses is essential for this project. The named postdoc
has a strong background in this type of research (e.g. Foster et al. 2008a+b, Foster et al. in
review). She has extensive experience with a large suite of microchemical techniques e.g. SIMS,
LA-ICP-MS, EMP, XAFS as well as studying changes in architecture within biogenic carbonates
e.g. critical point drying and SEM.
Consumables: During the first year € is required for culturing equipment with €
for
analyses of pre-existing material: electron microprobe (EMP) ~€ /day, LA-ICP-MS ~€
/day
nd
(in house prices); additional costs for preparation and SEM analyses. 2 year costs will be for
analytics (LA-ICP-MS, SEM, EMP) with chemical analysis of the culturing water (quadrupole
ICP-MS, ~€
). 3rd year completion of LA-ICP-MS, SEM, EMP analyses.
Travel: International conference attendance; work at Bremen (NanoSIMS and microsensors);
Glasgow (for EBSD analyses and meetings); SYXRF at Hamburg and attendance at BIOACID
workshops.
vi. References
Cusack, M, Perez-Huerta, A Dalbeck, P (2007) Common crystallographic control in calcite biomineralization of bivalved shells. Crysteng
9:1215-1218
Fabry V et al. (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICESJ Mar Sci 65:414-432
Fietzke J, Liebetrau V, Günther D, Gürs K, Hametner K, Zumholz K, Hansteen TH and Eisenhauer A (accepted) An alternative data
acquisition and evaluation strategy for improved isotope ratio precision using LA-MC-ICP-MS applied for stable and radiogenic strontium
isotopes in carbonates. J An AtSpectrom
Fietzke J, Eisenhauer A (2006) Determination of temperature-dependent stable strontium isotope (88Sr/86Sr) fractionation via bracketing
standard MC-ICP-MS. Geochem Geophys Geosys doi:10.1029/2006GC001243
Fietzke J, Eisenhauer A, Gussone N, Bock B, Liebetrau V, Nägler Th.F, Spero HJ, Bijma J, and Dullo C (2004) Direct measurement of
44
Ca/40Ca ratios by MC-ICP-MS using the cool plasma technique. Chem Geol 206:11-20
Foster LC, Allison N, Finch AA, Andersson, C (in review) Strontium distribution in the shell of the aragonite bivalve Arctica islandica.
Geochem Geophys Geosys
Foster LC, Allison N, Finch AA, Andersson C, Clarke LJ (2008a) Mg in aragonitic bivalve shells: seasonal variations and mode of
incorporation in Arctica islandica. Chem Geol, accepted
Foster LC, Andersson C, Høie H, Allison N, Finch AA, Johansen T (2008b) Effects of micromilling on δ18O in biogenic aragonite.
Geochemistry Geophysics Geosystems doi:10.1029/2007GC001911.
Genin A, Jaffe JS, Reef R, Richter C, Franks PJS (2005) Swimming against the flow: a mechanism of zooplankton aggregation. Science 308:
860-862
Halfar J, Steneck R, Schöne B, Moore GWK, Joachimski M, Kronz A, Fietzke J, Estes J (2007) Coralline alga reveals first marine record of
subarctic North Pacific climate change. Geophysical Research Letters 34, doi:10.1029/2006GL028811
Hansteen TH, Sachs PM, Lechtenberg F (2000) Synchrotron-XRF microprobe analysis of silicate reference standards using fundamentalparameter quantification. Euro J Mineral, 12:25-31
Heinemann A, Fietzke J, Eisenhauer A, Zumholz K (2008) Modification of Ca isotope and trace metal composition of the major matrices
involved in shell formation of Mytilus edulis. Geochem Geophys Geosys doi:10.1029/2007GC001777
Hoegh-Guldberg, O et al. (2007) Coral reefs under rapid climate change and ocean acidification. Science 318:1737-1742
Jackson CR (2004) An Atlas of Internal Solitary-like Waves and their Properties, 2nd Edition. Office of Naval Research, Global Ocean
Associates, Alexandria, VA , USA
Kamenos NA, Cusack M, Moore PG (2008) Coralline algae are global paleothermometers with bi-weekly resolution. Geochim cosmochim
acta 72:771-779
Kleypas JA et al. (2006) Impacts of ocean acidification on coral reefs and other marine calcifiers: a guide for future research. 88 p. Boulder,
Colorado: Institute for the Study of Society and Environment (ISSE) of the University Corporation for Atmospheric Research (UCAR).
Maier E, Tollrian R, Nürnberger B (2001) Development of species-specific markers in an organism with endosymbionts: microsatellites in the
scleractinian coral Seriatopora hystrix. Molecular Ecology (Notes) 1:157-159
Maier E, Tollrian R, Rinkevich B, Nürnberger B (2005) Reproductive mode and isolation by distance in the scleractinian coral Seriatopora
hystrix. Marine Biology 147: 1109-1120
181
Morse, DE et al. (1994) Morphogen-based chemical flypaper for Agaricia humilis coral larvae. Biol Bull 186:172-181
Mumby, PJ et al. (2006) Fishing, trophic cascades, and the process of grazing on coral reefs. Science 311:98-101
Orr JC et al. (2005) Anthropogenic ocean acidification over the twenty-first century and its impact on calcifying organisms. Nature 437:681686
Petersen D, Tollrian R (2001) Methods to enhance sexual recruitment for restoration of damaged reefs. Bulletin of Marine Science 69:9891000
Petersen D, Hatta M, Laterveer M, van Bergen D (2005a). Ex situ transportation of coral larvae for research, conservation, and aquaculture.
Coral Reefs 24:510-513
Petersen D, Laterveer M, Schuhmacher H (2005b). Innovative substrate tiles to spatially control larval settlement in coral culture. Mar Biol
146:937-942
Petersen D et al. (2006). The application of sexual coral recruits for the sustainable management of ex situ populations in public aquariums to
promote coral reef conservation - SECORE Project. Aquatic conserv: Mar Freshw Ecosyst 16:167-179
Richter C, Abu-Hilal A (2006) Seas of the Arabian Region (29,S). In: Robinson AR, Brink KH (eds) The global coastal ocean Interdisciplinary regional studies and syntheses, Vol 14, Part B. Harvard University Press, Cambridge, MA, p 1373-1412
Richter C, Bon M, Fillinger L, Jantzen C, Roder C, Schmidt G, Phongsuwan N, Khokiattiwong S (2008) Ocean dynamics drive coral reef
processes in the Andaman Sea 11th International Coral Reef Symposium. International Society for Reef Studies, Ft. Lauderdale, Florida,
USA
Richter C, Wunsch M, Rasheed M, Koetter I, Badran MI (2001) Endoscopic exploration of Red Sea coral reefs reveals dense populations of
cavity-dwelling sponges. Nature 413,726-730
Riebesell U et al. (2007) Enhanced biological carbon consumption in a high CO2 ocean. Nature 450:545-549
Riebesell U (2004) Effects of CO2 enrichment on marine phytoplankton. J Oceanogr 60:719-729
Riebesell U et al. (2000) Reduced calcification in marine plankton in response to increased atmospheric CO2. Nature 407:634-637
Riebesell U et al. (1993) Phytoplankton growth and CO2. Nature 363:678
Rüggeberg A, Fietzke J, Liebetrau V, Eisenhauer A, Dullo W-Ch, Freiwald A (2008) Stable strontium isotopes (88Sr/86Sr) in cold-water corals
– a new proxy for reconstruction of intermediate ocean water temperatures. Earth Planet Sci Lett,accepted
Weber M, Faerber P, Meyer V, Lott C, Eickert G, Fabricius KE, de Beer D (2007) In Situ Applications of a New Diver-Operated Motorized
Microsensor Profiler. Environ Sci Technol 41:6210-6215
Werner I, et al. (2002) Carnivorous feeding and respiration of the Arctic under-ice amphipod Gammarus wilkitzkii. Polar Biol 25:523-530
Werner I, Auel H (2005) Seasonal variability in abundance, respiration and lipid composition of Arctic under-ice amphipods. Mar Ecol Prog
Ser 292:251-262
Werner I (2006) Seasonal dynamics of sub-ice fauna below pack ice in the Arctic (Fram Strait). Deep-Sea Res I 53:294-309
Zumholz K, Hansteen TH, Hillion F, Horreard F, Piatkowski U (2007a) Elemental distribution in cephalopod statholiths: NanoSIMS provides
new insights into nano-scale structure. Rev Fish Biol Fisheries DOI 10.1007/s11160-006-9036-4
Zumholz K, Klügel A, Hansteen TH, Piatkowski U (2007b) Statolith microchemistry traces environmental history of the boreoatlantic
armhook squid Gonatus fabricii. Mar Ecol Prog Ser 333:195-204
Zumholz K, Hansteen TH, Piatkowski U, Croot PL (2007c) Influence of temperature and salinity on the trace element incorporation into
statoliths of the common cuttlefish (Sepia officinalis). Mar Biol 151:1321–1330 DOI 10.1007/s00227-006-0564-1
Zumholz K, Hansteen TH, Klügel A, Piatkowski U (2006) Food effects on statolith composition of the common cuttlefish (Sepia officinalis).
Mar Biol DOI 10.1007/s00227-006-0342-0
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Project 3.3: Ultra-structural changes and trace element / isotope partitioning in
calcifying organisms (foraminifera, corals)
(Jelle Bijma (PI und co-PI 3.3.1), Anton Eisenhauer (co-PI 3.3.2))
i. Objectives
For uni-cellular organisms like foraminifera, it has been shown that the carbonate chemistry of
the ocean not only affects the shell weight (Bijma et al., 2002) but also shell chemistry (e.g.
Bijma et al., 1999). Similar observations have been made for multi-cellular organisms like reef
building corals. The similarity in the ultra-structures of different calcifying organisms suggests a
common ancestral calcification mechanism. Here we propose to investigate the links between the
calcifying mechanism, the ultra-structure and the chemical composition. One group of the project
(3.3.1: co-PI J. Bijma) will investigate the ultra-structure of foraminifera and corals cultured
under different pCO2 conditions, whereas the second group (3.3.2: co-PI A. Eisenhauer) will
focus on trace element and isotope partitioning. In addition, the physico-chemical environment
within the cell will be determined to identify physiological processes that control the ultra
structure as well as the chemical and isotopical composition of the biogenic carbonate. The
overall aim of this study is to further develop a process based understanding of biocalcification
for uni- (foraminifer) and multicellular (corals) organisms, which would allow to predict the
consequences of ocean acidification on marine calcifying organisms and how this affects the
structural and chemical properties of their skeletons.
To understand the trace element partitioning and isotope fractionation in foraminiferal tests and
corals, used for paleo-climate reconstruction, extensive research on the calcifying mechanism
itself was performed during the last decades. For foraminifers this investigation led to a model
which builds on the following major mechanisms. Seawater is taken up by the organism
(endocytosis) and transported to the site of calcification. During this transport the composition of
the solution inside the vesicle is modified (concentrating Ca2+ while possibly removing other
divalent cations (notably Mg2+) and at the same time pumping H+ to “acidic” vesicles, thereby
increasing pH and CO32- in the “calcification” vesicles). This explains, on the one hand, why
foraminifers are among the best recorders of paleo-proxies (because calcification is based on
ambient seawater). On the other hand, but by the same token, it also explains why foraminifera
are so sensitive to ocean acidification. A similar model has been developed for corals (Erez,
personal com.). Physiological responses within the foraminiferal and coral calcification pathways
will determine their success as calcifiers in an acidified ocean. These responses, however, are
completely unknown and subject of the current proposal.
Uni- as well as multi-cellular marine calcifying organisms maintain their own distinct trace metal
homeostasis which results in characteristic and species specific elemental ratios as well as in
peculiar isotope fractionation (“vital effect”) which is significantly different from any inorganicthermodynamic expectations. This “vital effect” is most likely due to the development of
biochemical mechanisms to keep the trace metal homeostasis of the most important divalent
cations, magnesium (Mg) and calcium (Ca), and other trace elements, e.g. strontium (Sr) in
narrow limits in order to meet certain physiological needs. In particular, the uptake of Ca into the
cytoplasm is strongly limited due to its cell-poisoning effect. Furthermore, uni- and multi-cellular
calcifying organisms apply biochemical mechanisms in order to actively lower their Mg
concentrations and the Mg/Ca ratio in cell vesicles below a certain threshold before calcification
starts in order to precipitate calcite instead of aragonite.
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In this regard, the function of Ca2+-selective channels and Ca2+-ATPases embedded in the cellular
membranes have been recognized to be very important gateways to control trace metal fluxes
from an outside solution via the cytoplasm to the site of calcification. However, information
about their function for the transport of trace metals from seawater to the site of calcification is
limited. In particular, the partitioning of trace metals between seawater and biogenic CaCO3 of
the major divalent cation fluxes (e.g. Ca, Mg, Sr) and their dependency on external
environmental factors like seawater pH, salinity and water temperature has remained unexplored.
In order to overcome this lack of knowledge on the function of channels and ATPases we
propose to study divalent cation (Ca, Mg, Sr) partitioning between seawater and coral aragonite
as well as foraminiferal calcite as a function of those important environmental parameters in the
marine environment expected to change as a function of global increase in pCO2. The results of
this study will provide a better understanding of the calcification mechanisms as well as provide
quantitative information on trace metal ratios and isotope fractionation as a function of pH,
salinity and temperature. This will put distinct quantitative constraints on the modeling of the role
of ion selective channels and ATPases for the trace metal fluxes across membranes and the
marine calcification mechanisms.
Jelle Bijma established the interdisciplinary section Marine Biogeosciences at the AWI. He
holds a PhD degree in Marine Biology and Actuo-micropaleontology. Since almost 25 years he
and his group work on biological and paleoceanographic aspects of calcifying organisms. He is a
specialist in foraminiferal ecology, stable isotope and trace element geochemistry and
paleoceanography. His particular strengths for this project are his experience in interdisciplinary
work and expertise in foraminiferal calcification and its stable isotope and trace element
geochemistry.
Anton Eisenhauer is a physicist and has a long scientific track record in the field of lowtemperature and isotope geochemistry and their application to reconstruct past and present
environmental conditions in the marine realm. The proponent is the head of the isotope
geochemistry laboratory of the IFM-GEOMAR which is among the leading facilities for the
analysis of traditional and non-traditional isotope systems (e.g. δ25Mg, δ44/40Ca, δ88/86Sr, δ11B).
The proponent manages the IFM-GEOMAR laboratories which are equipped with state-of-the-art
mass-spectrometers and laser-ablation systems. The proponent has initiated studies on inorganic
precipitation experiments and biomineralization studies to examine the partitioning between the
bulk solution and the CaCO3.
The subproject 3.3.1 addresses the ultra-structure of the shells (foraminifera, pteropods,
ostracods) or skeletons (corals). Specifically, it focuses on the impact of the carbonate chemistry
on the calcifying mechanisms by investigating the resulting changes in the ultra-structure as well
as by following the fate of the “calcifying vesicles” using fluorescent probes.
The subproject 3.3.2 focuses on the function of ion selective channels and pumps by measuring
trace element and isotope partitioning between artificially altered seawater and the CaCO3
skeleton precipitated by selected marine calcifying organisms. During the course of the project
trace element partitioning and isotope fractionation will be studied by applying Ussing chamber
experiments. These kinds of experiments allow to study the role of Ca-activated channels and
pumps for trace element partitioning and isotope fractionation.
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Joint culture experiments (3.3.1 & 3.3.2)
Planktonic foraminifera
The basis for our investigations are culture experiments under controlled laboratory conditions.
Our groups have routinely carried out such experiments for many years. We use established
procedures for maintaining planktonic foraminifera in laboratory culture. Scuba divers
hand-collect live specimens of planktonic foraminifera from water depths of 2 to 6 m. Specimens
are brought back to the laboratory, where they were identified, measured with an inverted
microscope, and transferred to 0.8 µm filtered sea water in 115 ml culture jars. The jars are
sealed without an air space and placed into thermostated water tanks maintained at constant
temperatures (three groups: 23, 26 and 29°C).
Illumination is provided by F24T12/CW/HO fluorescent bulbs on a 12:12 hr light:dark cycle.
Symbiont bearing species are maintained under high light (“HL”, >380 µmol m-2 s-1), which
corresponds to maximum symbiont photosynthetic rates (Pmax) , and low light (“LL”, 20-30
µmol m-2 s-1), which is below the compensation light level . For comparison, ambient field light
levels are >2000 µmol m-2 s-1. The foraminifera are fed one brine shrimp nauplius every other day
until gametogenesis. Empty shells are then rinsed in purified water and archived in covered slides
for later analysis. The carbonate chemistry of the culture water is changed by keeping total
alkalinity, DIC or pH constant.
Corals and benthic foraminifera
Scleractinian and soft corals as well as benthic foraminifera will be cultured under controlled
laboratory conditions. In order to simulate global change on laboratory scale scleractinian and
soft corals (e.g. Pavona clavus) as well as benthic foraminifera will be cultured as a function of
selected pH (equivalent to atmospheric pCO2 of 180, 280, 380 and 700 ppmv), salinity (between
33 to 40) and temperature (benthic: 5, 10 and 15°C; corals: 23, 26 and 29°C ). To provide enough
material for analysis two field seasons (each lasting 3 to 6 months) of culturing at the Hebrew
University of Jerusalem (corals) and the Inter-university Institute (IUI) at Elat, Israel (e.g.
planktonic foraminfera and corals) are scheduled. In addition, the culturing facilities of IFMGEOMAR (e.g. soft corals and benthic foraminifera) and of the AWI (ostracods and benthic
foraminifera) will also be used in the framework of this study.
185
3.3.1
Impact of ocean acidification on the calcification mechanisms in marine calcifying
organisms and on ultra structural changes of biogenic calcite
Ultra structure
We will investigate the impact of ocean acidification on biogenic calcification, where nucleation
on organic membranes is a key process (Fig. 3.7).
Fig. 3.7: SEM micrographs. The left micrograph is showing the polished cross section of a foraminifer embedded into
a resin. The right micrograph is showing the close-up of the test wall. A special etching technique reveals structural
details, like the presence of layers, interpreted as organic linings.
By using AFM (Fig. 3.8) we will investigate how the test wall ultra-structure of (hyaline)
foraminifera (build up of units only a few nm in size, surrounded by organic layers) is affected by
changes in the carbonate chemistry of the culture medium. Such structures have also been
demonstrated in bivalves and corals. This consistency may point towards a universal mechanism
of calcification among these groups. Therefore we will investigate the ultra-structure in other
groups of calcifying organisms (pteropods, ostracods). High resolution AFM (i.e. below 1 nm)
will also be used to investigate ultra-structural changes of the calcifying units due to growth in
different carbonate chemistries.
Fig. 3.8: left: AFM scan of the three prominent lines shown in Figure 3.8 (right micrograph). Right: 500 nm x 500 nm
AFM scan, done between two of the lines shown in the left figure.
186
In addition, AFM force curves on the “organic membranes” will be performed to determine
functional groups active in biomineralisation. This will be complemented by inorganic calcite
growth experiments in AFM fluid cells which will allow direct monitoring of crystal growth in
the presence of organic molecules identified in foraminiferal tests. In this way, mimicking
processes assumed during the growth of biogenic CaCO3 will provide new insights into controls
on calcification and its relation to sea water chemistry that cannot be observed in vivo.
Vacuolisation and calcification
High-resolution fluorescence microscopy and con-focal laser microscopy will allow us to follow
the complete pathway from sea water vacuolization until calcification (Fig. 3.9). The incubations
will be extended by combining HPTS-incubations with other fluorescent probes to investigate the
role of organelles and analyze the contents of the high-pH vesicles (like Ca2+).
Fig. 3.9: Recently, the fluorescent probes
HPTS and Fluo-3AM have been applied to
foraminifera and can now be used to visualize
foraminiferal
intracellular
pH
and
Ca2+
respectively. Results with HPTS dissolved in
natural seawater (pH=8.2) indicate that before
a new chamber is formed in C. lobatulus,
vesicles with an elevated pH are formed in the
outer chambers and migrate towards the site of
calcification, where they produce a zone of
high (>9.0) pH in which CaCO3 precipitates.
Investigating the pH change along the calcification pathway and at the site of calcification is of
vital importance to understand the physiological response of foraminifera to ocean acidification.
To do so, foraminifera will be cultured under a range of different sea water pH's with dissolved
fluorescent probes. At different stages during calcification, the intracellular pH will be
determined.
3.3.2 The effect of decreasing pH, salinity and temperature on the trace element
partitioning between marine calcifying organisms and seawater.
Based on existing models on the function of ion selective channels (c.f (Gussone, Eisenhauer, et
al., 2003)) and pumps we will approach the problem by the measurement of trace element (Mg,
Ca, Sr) and isotope (δ25Mg, δ44/40Ca, δ88/86Sr, δ11B, δ13C, δ18O) partitioning between artificially
altered seawater and the CaCO3 skeleton precipitated by selected marine calcifying organisms
(scleractinian and soft corals, benthic foraminifera). According to existing models pH variations
should be reflected by the strength of the isotope fractionation process. These variations will then
help to improve our quantitative understanding of biocalcification as well as the impact of pH
variations on the rate of calcification.
Trace element and isotope analysis
All trace element (Mg/Ca, Sr/Ca, B/Ca, etc.) and isotope (δ25Mg, δ44/40Ca, δ88/86Sr, δ11B, δ13C,
δ18O) measurements will be performed at the mass-spectrometer facilities of the IFM-GEOMAR,
Kiel, Germany, using state-of-the-art mass-spectrometer technique (c.f. Heuser, Eisenhauer et al,
187
2002). Beside the Mg-, Ca-, and Sr-isotopes the boron isotope systematic (δ11B and B/Ca) will
also be monitored due to its sensitivity to pH- and CO32--concentration.
Spatial resolution and shell inhomogenities
In order to test spatial trace element and isotope inhomogenities we will apply the Laser-Ablation
system at the IFM-GEOMAR which allows a spatial resolution of about 10 µm and even better.
The Laser-Ablation system is designed to directly inject the sputtered material in the MC-ICPMS
for in situ determination of trace element concentrations and isotope fractionation.
Identification of transport enzymes
The identification of the Ca2+-binding enzymes (cf. Fig. 3.10) will be performed in close
collaboration with Prof. M. Bleich (Physiologisches Institut, CAU, Kiel). Furthermore, in order
to test the influence of Ca-selected channels and pumps on Ca isotope fractionation we will
perform experiments by using Ussing chambers. These kinds of experiments will be done in
close collaboration with M. Bleich and his staff members.
Links to other subprojects
There are several links to other BIOACID projects. A major link will be to the subproject 0.4
concerning “Training and transfer of know-how”. Together with U. Riebesell and M. Meyerhöfer
(subproject 0.4) and J. Fietzke (3.2.4) we will organize a workshop on MC-ICP-MS-, TIMSmeasurements and isotope fractionation during biomineralization. Such an event is important in
order to inform interested BIOACID scientists about instrumental and analytical progress.
There are close links concerning the study of the calcification mechanisms to subproject 2.1.3
which is focused on calcifying processes in macroorganisms and their relationship to changing
environmental conditions in the ocean. Further links exist to project 2.3.1 which will examine the
effects of changing ocean conditions on the otolith/statolith formation. Latter approach is also
directly relevant to our study.
Direct links exist to the projects 3.1.2, 3.1.3 and 3.1.4 which focus on calcification processes in
cold water corals, molluscs as well as on the membrane transport mechanisms for calcification
and pH-regulation.
Further links concerning the application of trace element and isotope partitioning effects exist
with subproject 3.2.1 (U. Riebesell). In collaboration with this subproject we will examine the
effect of ocean acidification and warming on pteropods from subpolar regions.
188
Fig. 3.10: ‘‘Epithelial’’ model of Mg2+ removal from the privileged space in perforate foraminifera. The parent
solution is based on seawater which arrives to the privileged space via vacuoles from the apical side. From the
privileged space Mg2+ diffuses into the cell via Mg2+ channels (bold arrow); the diffusion is favored by both the
concentration gradient and the membrane potential (_40 mV). In the cell, the free Mg2+ is buffered by binding to
negatively charged cytosolic molecules, mainly ATP, and by sequestration into cellular compartments such as
mitochondria and endoplasmic reticulum (ER). Simultaneously, active extrusion of the excess Mg2+ by ion exchangers
and pumps is exerted at the apical side. Note that all the above Mg2+ transport processes may also be employed on the
seawater vacuoles during their intracellular pathway to the calcification site (from Bentov and Erez, 2006).
Schedules 3.3.1 and 3.3.2
3.3.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Literature study, preparation field
season
Set-up of culturing facility; first CO2
perturbation experiment
Sample processing and
measurements: AFM +
(convocal)microscopy
Set-up of culturing facility; second
Sample processing and
measurements: AFM +
(convocal)microscopy
data interpretation
Manuscript preparation, presentation
of results at conferences
189
Milestones (3.3.1)
- 1st experiment successfully carried out
month 09
- 1st experimental data set on CO2/pH sensitivity (180 and 380 ppmv)
month 18
- 2nd experiment successfully carried out
month 21
- 2nd experimental data set on CO2/pH sensitivity (280 and 700 ppmv)
month 30
- Evaluation of combined data set; Sensitivities and uncertainties
month 33
3.3.2
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Literature study, preparation field
season
Set-up of culturing facility; first CO2
perturbation experiment
Sample processing and measurements:
TIMS MC-ICPMS
Set-up of culturing facility; second
Sample processing and measurements:
TIMS MC-ICPMS
data interpretation
Milestones (3.3.2)
- 1st experiment successfully carried out
month 09
- 1st experimental data set on CO2/pH sensitivity (180 and 380 ppmv)
month 18
- 2nd experiment successfully carried out
month 21
- 2nd experimental data set on CO2/pH sensitivity (280 and 700 ppmv)
month 30
- Evaluation of combined data set; Sensitivities and uncertainties
month 33
First Year
Personnel costs
3.3.1
3.3.2
Subtotal
190
Second Year
Third Year
Total
First Year
Second Year
Third Year
Total
Consumables
3.3.1
3.3.2
Subtotal
Travel
3.3.1
3.3.2
Subtotal
Other costs
3.3.1
3.3.2
Subtotal
Investments
3.3.1
3.3.2
Subtotal
TOTAL
3.3.1
Personnel costs: A post doctoral position for 30 months (6 months financed via different
funding). We request a post doc for this project because the scheduled scientific program is much
too ambitious for a Ph.D.-student.
Consumables: microelectrodes, fluorescent probes, AFM tips, chemicals, glasware, etc. for
experiments
Travel: travel for PI and post doc to conferences and to Elat for fieldwork.
Other costs: The total costs of carrying out fieldwork in Elat will amount to about k€ per field
season (bench-fees, use of boat and divers for collection, accommodation and subsistence). The
AWI contribution to this is € k€.
3.3.2
Personnel costs: The personnel costs for the project comprise 1.5 post-doc years. We request a
post doc for this project because the scheduled scientific program is much too ambitious for a
Ph.D.-student and request an experienced post-doc. We intend to hire Dr. Isabell Taubner who
has already started to perform culturing of the requested sample material in the frame of an
191
excellence cluster project in Israel. Part of the sample material for the intended Bioacid study will
be available already end of 2008 when the cluster project is terminated. Further material will then
be cultured in the frame of this project Although the intended 18 post-doc months in the
subproject 3.3.2 will not cover the whole project time the continuity in data interpretation and
networking with other Bioacid projects is further guaranteed by the project Co-PI A. Eisenhauer.
Consumables: The costs for consumables cover the expenses for lab rent in Eilat (in
collaboration with Prof J. Erez, HUJI) and Jerusalem as well as the expenses for chemicals, lab
ware for both laboratories and transportation from Israel to Germany. Furthermore, costs for
chemical preparation of samples and for the mass-spectrometer measurements at the massspectrometer facilities of IFM-GEOMAR by applying state-of-the-art equipment (Laser-MCICP-MS, TIMS, MC-ICP-MS, etc) will also be covered by the consumables.
Travel: Culturing experiments will be performed in the first project year in Israel. The requested
travel costs cover the transfer of personal from Germany to Israel and will also cover the daily
expenses for the post-doc during his stay in Israel. The travel costs will also cover expenses for
renting a flat to save hotel costs. Furthermore, the costs will also cover the costs for the
occasional renting of a car to transport Gulf of Elat water from Elat to Jerusalem for the culturing
experiments.
vi. References
Bijma JH, Spero J, et al. (1999) Reassessing foraminiferal stable isotope geochemistry: Impact of the oceanic carbonate system (experimental
results). Use of Proxies in Paleoceanography: Examples from the South Atlantic. G. Fischer and G. Wefer. Berlin, Heidelberg, SpringerVerlag: 489-512.
Bijma JH, Hönisch B et al. (2002) Impact of the ocean carbonate chemistry on living foraminiferal shell weight: Comment on "Carbonate ion
concentration in glacial-age deep waters of the Caribbean Sea" by W. S. Broecker and E. Clark - art. no. 1064." Geochemistry Geophysics
Geosystems 3: 1064-1064.
Bentov S, Erez J (2006) Impact of biomineralization processes on the Mg content of foraminiferal shells: A biological perspective. Geochem.
Geophys. Geosyst. 7(Q01P08).
Griffith EM, Paytan A, Kozdon R, Eisenhauer A, Ravelo AC (2008) Influences on the fractionation of calcium isotopes in planktonic
foraminifera, Earth Planet Sci Lett., 268, 124-136.
Gussone N, Eisenhauer A et al. (2003) Model for Kinetic Effects on Calcium Isotope Fractionation (δ44Ca) in Inorganic Aragonite and
Cultured Planktonic Foraminifera. Geochim Cosmochim Acta 67(7), 1375-1382.
Heuser A, Eisenhauer A et al. (2002) Measurement of Calcium Isotopes (δ44Ca) Using a Multicollector TIMS Technique. International Journal
of Mass Spectrometry 220, 385-397.
192
Project 3.4 Microenvironmentally controlled (de-)calcification mechanisms
(PI: Michael Böttcher,)
i. Objectives:
Our plan is to determine the effect of changing seawater pH and the dissolved carbonate species
on calcification and decalcification in marine sediments and microbial mats. In these matrices
microbial activity and mass transfer resistances control the pore water chemistry and thus the
microenvironments where calcification and decalcification occurs. One hypothesis we will test is
that the microenvironment in the top mm of such systems is buffered against pH and CO2
changes, due to mass transfer resistances. Secondly, we will investigate in how far sediment
processes can buffer the pH in the water column, depending on exchange rates and the reactivity
of the biogenic carbonates. We will further investigate how the water column chemistry of the
Wadden Sea reflects exchange with the North Sea and surface sediments.
ii. State of the Art:
Effect of ocean acidification on benthic calcification
Whereas much research is done towards calcium cycling in ‘classical’ calcifying marine systems
such as corals, foraminifera and coccolithophores, and much progress towards its understanding
has been obtained, much less is done on calcification driven by photosynthesising
microorganisms. Examples of the latter are calcification in stromatolites and tufas, calcareous
sediments, and beachrock (Krumbein, 1979). The fundamental difference with the main marine
calcifiers is that calcification is a side process of metabolic activity, rather then a well controlled
process leading to species-specific structures, such as shells and skeletons. Essentially,
photosynthetic CO2 fixation leads to a shift in the pH, increasing the oversaturation of CaCO3.
This may be a globally important process for all shallow coastal areas with sediments in the
photic zone. Microbially driven calcification in reef sediments was estimated to be of the same
order of magnitude as by corals (Werner et al., 2007), calcified structures developing in hardwater creeks, tufas, were shown to be formed by photosynthesis (Bissett et al., 2007), and also in
stromatolites and hypersaline mats microbial photosynthesis was shown the driving force for
calcification (Ludwig et al., 2005). Degradation of EPS by sulphate reduction is thought to play a
role in calcification in stromatolites and cyanobacterial mats (Arp, Reimer & Reitner, 1999;
Visscher et al., 1998). It is thought that microbial degradation of these calcium ion binding
polymers plays an initiating and structuring role in calcification. Calcification in benthic
communities can be studied by direct budgeting methods, including tracer studies, using
radioactive 45Ca2+ or 14CO32- (Al-Horani et al., 2005), alkalinity shifts (Gattuso, Allemand &
Michel, 1999; Gattuso et al., 1996), Ca2+ fluxes, the general characterization of the calcifying
sites and driving microbial processes can be measured by microsensors (Bissett et al., 2007; de
Beer et al., 2000; Werner et al., 2007; Wieland et al., 2001).
Dissolution of carbonates in sediments of the North Sea and the Southern Ocean
Decalcification leads to a pH increase and thus can buffer the effects of increasing CO2. The
marine biogeochemical carbon cycle is controlled by biological processes that influence the
distribution of carbon in the surface sediments of the Ocean and exchange processes between
sediments and water column and by exchange processes with the tidal coastal areas (Brasse et
al.1999; Kempe & Pegler, 1991; Thomas et al., 2007). After the death of organisms with biogenic
193
carbonate shells, part of this material arrives on the sea floor where it is decayed. Biogenic shells
are an important reservoir of bound oxidized carbon. The periostracum layer of shells enhances
the availability of electron donors for remineralizing epibiontes that may enhance carbonate
dissolution (Knauth-Köhler et al. 1996). The geographic distribution of biogenic carbonates is the
result of synergistic thermodynamic and kinetic interaction processes with aqueous solutions
(Wollast 1994), controlling the degree of carbonate preservation in surface sediments. Higher
solubility of CaCO3 in cold water regions leads to a low preservation potential in temperate and
cold climate zones (Morse and Mackenzie, 1990; Wefer et al., 1987). Alexandersson (1979) was
able to demonstrate the decay of carbonate shells in Skagerrak surface sediments. Most of the
North Sea regions are living grounds for a large number of carbonate-forming species, mainly
bivalve molluscs with an epibiontic (Mytilus edulis, Crassostrea gigas) or endobiontic life mode
(Cerastoderma edule, Mya arenaria, Ensis ensis and Macoma baltica). In addition, eroded
terrestrial carbonates form part of the carbonate pool in North Sea surface sediments that may
interact with the pelagic dissolved carbonate system. Decreasing pH and increasing carbon
dioxide partial pressure will increase the solubility and hence will affect the formation and
dissolution rates of the different CaCO3 fractions. Although the influence of bacterial activity on
carbonate precipitation has been demonstrated in tropical areas, less is known on the influence of
biological (microbial) activity on the degradation processes in shallow water areas of the
temperate climate zone (e.g. Aller 1982; Golubic & Schneider, 1979; Green and Aller 1998;
Krumbein, 1979). Therefore, these processes are an as yet quantitatively unknown factor in the
carbon cycle of coastal areas. The North Sea carbonate system is closely related via exchange to
the processes taking place in the intertidal areas and is already under influence of human activity
and global climate change (Brasse et al., 1999; Thomas et al., 2007).
In the high-latitude oceans the carbonate concentration is the lowest in the world because of the
very low water temperatures which shift the chemical equilibria of the carbonate system towards
low carbonate concentration. The saturation state with respect to aragonite and calcite in these
waters is, therefore, the lowest, although oversaturation is still prevailing. Changes in the
carbonate system due to acidification will have the highest impact here. These waters will in the
future turn out to be most corrosive to aragonite and calcite (e,g. Orr et al., 2005). Ocean
acidification will first affect the surface ocean, but the signal will subsequently be transferred to
the subsurface and deep oceans. This will largely occur in the polar oceans, where new deep and
bottom waters are formed, mainly at and near the continental shelves. The high-latitude oceans
are also the conduit for the transfer of anthropogenic CO2 from the surface layer to the deep
oceans. Low carbonate concentration and high uptake of anthropogenic CO2 constitute the most
destructive combination with respect to ocean acidification. Calcareous sediments in the polar
regions represent the initial spatial and temporal buffering of global ocean acidification – further
buffering will occur on centennial time scales on the abyssal sea floor. It has long been known
that Antarctic Bottom Water plays a substantial role in the dissolution of calcareous sediments
(Berger, 1970). Although the Southern Ocean is more known for its arenaceous sediments, also
significant portions of carbonate can be found (Hulth et al., 1997). Foraminiferal assemblages are
found in the surface sediments of the Weddell Sea (e.g. Anderson, 1975). Also pteropods shells
are preserved in the surface sediment muds. Pteropods have high abundances in the Southern
Ocean, both in the open ocean and on the continental shelves (e.g. Accornero et al., 2003; Hunt
et al., 2007). Pteropods produce aragonite, a metastable form of CaCO3 that is more soluble than
calcite, making aragonite the more important buffering agent of ocean acidification. Berner and
Honjo (1981) estimated that, as an example, aragonite constitutes at least 12% of the global
CaCO3 flux.
194
3.4.1 Michael E. Böttcher (IOW) has studied the biogeochemistry and stable isotope
geochemistry of carbon, sulfur and metals in the water column, in modern and ancient marine
sediments, and ground water systems at different locations, including the intertidal and the
pelagial of the open North Sea, the Baltic Sea, and carbonate karst systems (e.g., Böttcher, 1999;
Böttcher et al., 2000, 2007; Dellwig et al., 2007). Additionally, a number of studies were carried
out on the geochemistry of the experimental formation and destruction kinetics in temperate
carbonate systems (Böttcher, 1997, a,b, 1999), and the structural characterization of biogenic
carbonates (e.g. Böttcher et al., 1997). Since 2006 he is heading the Marine Geochemistry group
at IOW.
Gerd Liebezeit is working since 1977 on different chemical aspects of coastal regions and landocean interactions, since 1991 with emphasis on the the Wadden Sea. Research topics include
a.o. dating of biogenic carbonates (Behrends et al., 2003, 2005), geochemistry of intertidal
deposits (Hertweck and Liebezeit, 1996; Hertweck and Liebezeit, 2002; Hertweck et al., 2006)
and phosphate uptake by intertidal biogenic carbonates, dynamics of the nutrient cycles as well as
characterization and transformation of organic compounds.
3.4.2 Dirk de Beer (MPI-MM, Microsensor Group) studies how transport controls microbial
processes in sediments, biofilms and microbial mats. Microbial processes studied include
photosynthesis, aerobic respiration, denitrification, nitrification, sulphate reduction and sulphide
oxidation. He has over 130 peer reviewed publications. His special expertise is use and
development of micro-environmental analyses by microsensors, combined with geochemistry and
community studies. Calcification studies were done on calcification in corals, foraminifera,
cyanobacterial mats, Halimeda, tufas, and reef sediments.
Marcel Kuypers (MPI-MM, Nutrient Group) is a marine chemist who studies microbial controls
of the global element cycling using stable isotopes. He has studied mechanisms and
biogeochemical implications of global organic carbon burial. He has made break-throughs in the
research of the global nitrogen cycles, particularly on the importance of the recently discovered
ANAMMOX process. He has over 30 peer reviewed publications in the last 5 years. He is
responsible for the nanoSIMS, that is recently installed in our institute.
3.4.3 Mario Hoppema (AWI) has been active in Antarctic research since 1992, where the
biogeochemical cycle has been his main research interest (e.g. Hoppema, 2004; Hoppema and
Anderson, 2007). Chemical data have been used to obtain biological parameters of the Weddell
Sea (e.g. Hoppema et al., 2002; 2007). Within the framework of the EU project CarboOcean a
carbon data synthesis is underway, scheduled to be finished in 2008. Hoppema is the leader for
the Southern Ocean part of this synthesis
Christoph Völker has been modelling the physics of the Southern Ocean (Olbers et al., 2004)
and has developed a biogeochemical model, based on the MIT circulation model (Marshall et al.,
1997). He has studied the influence of phytoplankton community composition (Denman et al.,
2006) and phytoplankton physiology (Hohn et al., 2008) on elemental fluxes using global
planktonic ecosystem models. He is currently involved in EU project CarboOcean dealing with
physical and biological feedbacks of Southern Ocean carbon chemistry to climate change.
Dieter Wolf-Gladrow has been research scientist at AWI since 1987 and since 1999 Professor of
Theoretical Marine Ecology at the University of Bremen. He is an internationally recognized
expert on all aspects of the marine carbonate system (e.g. Jansen et al., 2002) and wrote a much
195
used textbook on the CO2 system (Zeebe and Wolf-Gladrow, 2001). Ocean acidification already
had drawn his interest at an early stage (Wolf-Gladrow et al., 1999).
Subproject 3.4.1 Impact of biogenic carbonates on pH buffering in an acidifying coastal sea
(North Sea)
(M.E. Böttcher)
We propose to investigate the influence of changing pH and carbon dioxide partial pressure, and
alkalinity on carbonate dissolution (and production) in surface sediments of the Wadden Sea and
the consequences for and relation to the carbonate system of the North Sea. Our hypothesis is that
biogenic carbonates at and just below the surface of intertidal sediments play a role in modifying
tidal waters that exchange with the shallow North Sea. This adds to carbonate fluxes caused by
benthic organic matter degradation. The absolute and relative importances will change as the
North Sea carbonate system will acidify in the future.
One important aspect will be the reactivity (reaction rates and thermodynamic stability) of
different biogenic carbonates (e.g., Mytilus, Crassostrea, Mya, Hydrobia, etc.) and the
experimental quantification of the different dissolution rates in pure and seawater media under
different controlled laboratory conditions, i.e. pH, partial pressure of carbon dioxide (pCO2-stat
and free-drift), and carbonate undersaturation. The reactive surface of the carbonate will be
characterized to extract specific dissolution rates from experimental data on the development of
liberated major minor and trace cations and the dissolved carbonate species. Experiments will be
carried out at different fixed CO2 partial pressures (as defined by the BIOACID consortium). This
approach will include in parallel experiments geomicrobiological effects on microbial
degradation of the organic matrix compared to purely abiotic reactions.
Experimental laboratory-based approaches will be compared to field in-situ transformation
experiments and will be followed by microscopic (e.g., SEM-EDX), inorganic geochemical (ICPOES, photometry, microsensors) and stable isotope (C, O; irmMS) approaches. Experiments will
also be carried out in collaboration with Dr. Hoppema (3.4.3) regarding benthic mesocosm
experiments at AWI. Field work will additionally characterize the sources and surface textures of
different carbonate fractions in surface sediments with geochemical methods and SEM-EDX.
Thus we will evaluate the time-dependent corrosion of biogenic carbonates as a function of burial
time. Application of microsensors to characterize the chemical gradients will be carried out in
collaboration with Dr. D. de Beer (MPI-MM; 3.4.2). From the side of carbonate formation, the
influence of a changed carbonate system on growth of Mytilus and Crassostrea is planned to be
eventually assessed in a later phase of the project.
Finally, pelagic measurements and experimental results will link the alkalinity and DIC exchange
with the coastal North Sea and the possible ecosystem consequences via biogeochemical
modelling. Parameters of the dissolved carbonate system (TA, CA, DIC, PCO2, pH) will be
measured in collaboration with PD Dr. Bernd Schneider (IOW) while δ13C(DIC) will be
determined via irmMS. These results will be provided for a collaborative integration in Theme 5
(Pätsch et al.) and the inclusion in the modelling of the North Sea carbonate system. The
approach provides the base for a quantitative understanding of the role of the carbonate system in
the water column on preservation, destruction and temporal authigenesis in the intertidal surface
sediments, and the role of benthic metabolisms.
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Links to other subprojects
There are links to subproject 1.2.5 regarding benthic carbonate-relevant processes and 2.1.3 and
3.1.3 which will provide TP 3.4.1 with shell material grown under well-defined conditions for
dissolution rate studies. Microsensor approaches in experimental dissolution studies will be used
in collaboration with TP 3.4.2, and an experimental collaboration with TP 3.4.3 will make use of
field aragonite in dissolution studies at IOW and benthic mesocosm experiments at AWI.
A direct link exists to project 5.1.which focuses on the modelling of the carbonate system of the
North Sea. TP 3.4.1 will provide field data for the carbonate system at the tidal-open North Sea
boundary leading to a close link between experimental and modelling approaches.
Schedule
3.4.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Measurements in the pelagic water
column of the intertidal, in-situ
experiments
Carbonate system of the intertidal,
transects and cycles
Experimental dissolution of shells
Evaluation of field data and calibration
of future campaigns
Mesocosm experiments with AWI
(3.4.3)
Growth experiments, Geomicrobiology
and Modelling
Writing of manuscripts presentation on
conferences
Milestones (3.4.1)
- Implementation of carbonate system data in model environment of theme 5
(Pätsch et al.)
month 9 and 24
- Implementation of experimental setup
month 12
- Field cruises (transects) on the pelagic Intertidal
month 27
- Mesocosm experiments at low temperatures
month 27
- Reactivity of carbonates in the North Sea Intertidal
month 33
- PhD Thesis and publications
month 36
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Subproject 3.4.2 Benthic (de-)calcification driven by microbial processes
(D. de Beer)
Hypothesis: Whereas corals and coccolithophores are very sensitive to pH, microbially driven
calcification is rather resilient against pH variations in the water column. This difference can be
understood from the essential differences in calcification mechanism. Calcification by skeleton
and shell building organisms is actively controlled by energy demanding active transport of H+
and Ca+2. A lower pH will make calcification more energy demanding, and thus make these
calcifiers less competitive than non-calcifiers. Calcification in mats and sediments is a sideproces of photosynthesis, inducing a microenvironment controlled by the microbial activities and
the transport resistance in the sediments. The transport barrier separates the benthos largely from
seawater, and buffers the sediments against changes in the seawater chemistry.
WP1
Experimental studies of calcification and decalcification
We will investigate calcification in phototrophic communities by (1) budgeting the exchange of
calcium between (de-)calcifiying sites and seawater and quantify the driving metabolic process
rates, and by (2) studying incorporation of calcium carbonate in the calcite matrices, and (3)
determine the effect of pH stress on the community structure and spatial distribution.
1) We will determine the microenvironment in the benthic communities and the relevant fluxes
by microsensors for pH, O2, Ca2+, CO2 and CO32-, and the rates of photosynthesis and respiration,
by membrane inlet mass spectrometry (MIMS), photopigment fluorescence (PAM) and
microsensor techniques. Calcification measurements in the field will be assessed using the eddy
correlation technique (Berg et al., 2003), which is recently adapted to measure Ca2+ fluxes.
2) The incorporation of calcium and carbonate into the solid matrices will be studied by radioand stable isotope techniques (by ß-imaging and nanoSIMS), to assess how closely calcification
is associated with microbial cells (with sub-µm resolution). We will investigate, with Dr. A.
Eisenhauer, trace element incorporation in response to environmental factors and calcification
rates.
3) We will determine the dynamics of phototrophic benthic communities in response to pH shifts
by hyperspectral (HS) imaging, which allows quantitative assessment of the 2-D distribution of
photopigments. By imaging their variable fluorescence we will assess whether they are part of an
intact photosystem. HS-imaging is a non-destructive community analysis, and thus we can follow
the short-term community dynamics in response to an environmental stress.
Experiments will be done under a wide range of artificially imposed pH values, adjusted by CO2
levels, in agreement with the strategy defined by the consortium. We will install mesocosms for
incubations and measurements in the institute. Samples will be obtained from different habitats,
from freshwater stromatolites (tufas), marine sediments and hypersaline mats. The tufas can be
obtained from several creeks in the Harz (ca 200 km from the MPI), marine carbonate sediments
from the Mediterranean and hypersaline mats are currently cultivated in the institute. The
comparison of different salinity is interesting as the pK2 strongly decreases with salinity. Field
measurements can well be done using a field station on Elba, from where we will also obtain
carbonate sediments. The experimental work will be done by a PhD student. The student will also
be active in teaching the microsensor course, with help of staff members from the microsensor
group.
198
WP2
Biogeochemical modelling of calcification and decalcification
The data will be used to construct and refine a transport-conversion model, that includes acidbase equilibria and calculates the pH and DIC profiles. An approach to build such a model was
described by colleagues from the NIOO (Ierseke, Nl) and the Free University of Brussels (Dr. P.
Meysman), with who we will collaborate. A second step will be to expand this model with
calcium carbonate dissolution and precipitation reactions. Conceptually, how microbial and
biogeochemical processes control calcification and decalcification are well known. Processes
leading to acidification lead to dissolution and increased alkalinity can enhance precipitation. As
for any heterogeneous (involving a water and solid phase) process, modelling of calcification
needs to address the quantitative description of the microenvironment where the reaction
(calcium carbonate precipitation or dissolution) occurs. A second step is to incorporate the local
solid-liquid exchange, so between calcite matrices and porewater, depending on the local
microenvironment. Both steps are highly challenging. The relevant microenvironmental
parameters for calcium carbonate dissolution and precipitation include the calcium and carbonate
concentrations, and thus the DIC and pH. Many biogeochemical processes influence this
seemingly simple set of parameters, complicating the modeling. Moreover, the kinetics of
calcium and carbonate exchange between the solid and water phase depends on a variety of
poorly understood parameters, such as organic matrices associated with the precipitate, and the
anion composition of the water phase. Thus meaningful modelling must be accompanied by
measurements.
A model of the carbonate system near photosynthesizing calcifying foraminifera was build
several years ago (Wolf-Gladrow, Bijma & Zeebe, 1999), which calculated the local pH from
respiration and photosynthesis in a spherical geometry. A comprehensive geochemical model
should include a range of geochemical processes (Soetaert et al., 2007). We aim to construct such
a more comprehensive biogeochemical model to calculate the local carbonate system and pH
profiles. The problems to solve a system with many variables and an equal number of equations
on processes with a wide variety of timescales are described in detail and solution methods are
offered (Hofmann et al., 2008). Their approach is a series of local mass balances, and includes
diffusional and/or advectional mass transfer. Such a model describes the gradients of the
carbonate system in sediments and microbial mats, including the pH, but not yet the effects of the
local microenvironment on the precipitation and dissolution of calcium carbonate. Parameters
controlling these processes must be experimentally determined for each studied system.
Obviously, calcification and decalcification again influence the local pH, and once known must
be integrated into the model. The determination of these processes under different
microenvironmental conditions is an important aim of our studies with microsensors, ß-imaging
and nanoSIMS. The experiments can be done on a range of conditions, using the modelling a
wide generalization of the phenomena observed will be achieved. Such a model will help
defining the threshold values for net calcium dissolution versus precipitation in various benthic
systems.
Microsensor course
We will offer a microsensor course for the participants of the Verbundprojekt. The course will be
in two weeks. The first week making of microsensors for oxygen and various ions will be
learned. The most practical teaching will be done by our expert team of technicians. The second
week will be a training in various microsensor applications, under guidance of scientists. Here the
participants are encouraged to bring their own samples, however, we have interesting sample
material available. Hopefully, microsensors that are build in the first week may be used, whereas
199
microsensors build by our experts will be needed in addition. There will be daily seminars on the
theory behind microsensor use and examples of complete studies, and also participants are
invited to present their work. Each course will be for 8 participants maximal, we foresee the need
for 2 courses 1 year separate.
Collaborative microsensor experiments are planned with project 4.1.3 (Bischof )
Schedule
3.4.2
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Mesocosm experiments
Tracerstudies incl. nanoSIMS
ß-imaging
modeling
Field measurements
Microsensor course
Collaborative work
Milestones (3.4.2)
- Implementation of experimental facility, and learning of basic experimental
skills
- Experimental data set on CO2/pH sensitivity
- Field data set on diel calcium fluxes
- Model for benthic microprofiles of solutes and activities
- Expansion of model for calcium exchange
- Microsensor courses
- Experimental data sets on effect of salinity on CO2/pH sensitivity
- Thesis defense
month 6
month 9
month 15
month 24
month 36
month 12 and 24
month 30
month 36
3.4.3 Buffering ocean acidification: Dissolution of carbonate sediments in the Southern
Ocean (M. Hoppema)
We propose to examine the buffering capacity of Antarctic sediments, where our first focus is on
the continental shelves. In particular, the role of pteropod aragonite in surface sediments in the
buffering will be investigated. This shall be done in a two-pronged study, addressing issues that
are related within the biogeochemical cycle of the ocean and the seafloor. The projection of the
(future) decadal trend of acidification of Southern Ocean waters constitutes one thrust of our
study. We shall determine the development of pH, pCO2 and the carbonate saturation index
Omega based on both observations and modeling. Data both of ourselves and from the EU IP
CarboOcean carbon synthesis will be used to obtain pH and pCO2 in waters from the Weddell
Sea and environs, with emphasis on the shelves. In some regions, we may get a time series of
200
repeat occupations. This will allow us to calculate the degree of saturation for aragonite and
calcite for the time series period. We will extend the observed time-series of anthropogenic
carbon over the next 100 years, using a state-of-the-art global biogeochemical model that has
been especially tuned to reproduce present-day observations of nutrients, biological activity and
the carbonate system in the Southern Ocean, using our observations of acidification as additional
initial constraints. We will run the model a) with a prescribed atmospheric pCO2 increase keeping
the physical forcing (wind stress, heat flux) fixed, and b) also with changes in the physical
forcing as obtained from coupled climate prediction runs, to separate the effect of changing
climate from those of increasing atmospheric pCO2 on the CaCO3 saturation on the Antarctic
shelf. This work will be done by a PhD student, strongly assisted by the PIs. As to the
parameterization of calcification, pCO2 sensitivities and DOC cycling, experience and data will
be shared with projects 1.3, 5.1 and 5.2.
As to the second prong, we will address the rate of dissolution of carbonate on shelves in the
Weddell Sea and Antarctic Ocean. The potential future acidification, determined from the other
prong of our research, will serve as a realistic scenario, against which the dissolution of
sedimentary carbonate will be assessed. Sediment samples of the Antarctic Ocean from the AWI
core repository will be investigated with respect to composition. Of particular interest is the
fraction of pteropod aragonite. The composition will be studied as a function of small-scale and
large-scale environmental factors, like grain size, water depth, sedimentation rate, characteristics
of the overlying water mass and current velocity. The maximum buffer capacity is obtained by
the absolute content of carbonate in the sediment. Analyses will be done with Isotope Ratio Mass
Spectrometry. As a further step, dissolution experiments with sediment samples and pteropod
shells under various conditions (pH, pCO2, CaCO3 saturation state, water current) will be
performed. Samples will be collected during a Polarstern cruise to the Southern Ocean using
large box corer (GKG) or multiple corer (MUC). We will use Rhizon sampling of pore waters
(Seeberg-Elverfeldt et al., 2005) and benthic flux chambers. Both microscopic and chemical
methods will be applied, where part of the experiments will be performed in joint work with
other sub-projects - a). Using micro-sensors of the MPI Bremen; Dr. de Beer, subproject 3.4.2,
and b) With subproject 3.4.1 (Dr. Böttcher), aragonite dissolution experiments at IOW and
benthic mesocosm work at AWI. This will render information on the dissolution rates.
Additionally, with subproject 1.2.5 we envision cooperation with regard to pressure-adapted flux
chambers. The pressure laboratories utilized for identification of turnover rates and fate of
nutrients and carbon from aggregates in the benthic boundary layer permit to actively
control/select erosion or deposition of sediments. This is achieved by pressure-adapted flux
chambers where bottom stress together with water column turbulence mimics the phases of tidal
cycles. Depending on the composition of the sediments and fluid introduced into the chamber, the
buffering capacity of the sediment/pore water region can be obtained from fluid samples drawn in
addition to the aggregate dynamics. Finally, within the Alfred Wegener Institute there is
extensive experience with sediment work, while the core repository contains numerous cores
from the Weddell Sea and Antarctic Ocean. Several sampling and measurement techniques are
available within the geochemistry and geology departments.
Eventually, we will estimate the spatio-temporal distribution of CaCO3 dissolution rates around
Antarctica from the combination of the laboratory results with sediment distribution. This
distribution will be implemented as bottom boundary condition for dissolved inorganic carbon
and alkalinity in the biogeochemical model to investigate the propagation of the buffering signal
into the ocean interior. Through the combination of observations, experiments, and simulations
we will thus estimate the buffering effect of CaCO3 and its kinetics in Antarctic shelf sediments
on a decadal to centennial time scale.
201
Schedule
3.4.3
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Reading, modelling of water column
data
Set up, preparation of experiments,
calibrations
Dissolution experiments on cores
Polarstern cruise to Southern Ocean, on
board experiments
Analysis of results, computing
Writing of manuscripts presentation on
conferences
Milestones (3.4.3)
- Future projection of acidification of Antarctic water column
- Implementation of experimental setup
- Data set of dissolution of sediments cores plus preliminary interpretation
- Polarstern cruise
- Data set of field data
- Thesis
month 09
month 12
month 21
month 27
month 30
month 36
First Year
Personnel 3.4.1costs
3.4.1
3.4.2
3.4.3
Subtotal
Consumables
3.4.1
3.4.2
3.4.3
Subtotal
Travel
3.4.1
3.4.2
3.4.3
202
Second Year
Third Year
Total
First Year
Second Year
Third Year
Total
Subtotal
Investments
3.4.1 Gas-mixing device for
chemostat chamber
pH-meter
3.4.2 Microsensor
equipment
(micromanipulator, motor,
data acquisition, cables)
Subtotal
Other costs
3.4.1 Contract
measurements,
geomicrobiology and ship
time for sampling the
intertidal pelagial
3.4.2 Microsensor course
Subtotal
TOTAL
Budget Justification
3.4.1
Personnel costs: The experimental studies on the reactivity of biogenic carbonates and the
carbonate system of the intertidal pelagial will be performed by a PhD student, paid according to
German TVöD system.
Consumables/p.a.:
Filtration material, disposables
€
Pure gases and gas mixtures
€
p.a. chemicals, ultrapure acids, standards
€
Electrodes, cable
€
Glass ware, fittings, vales, plastic tubings
€
Pipettes
€
Nylon net (incubations)
€
Travel: BIOACID-meetings in year 1, 2 and 3 (
€ each), 2 international scientific meetings
in the last 2 years (
each), 2 cruises in years 2 and 3 (
each), regular visits with
partners
203
Investment: Installation of a gas mixing (CO2-N2) device with fine-316ss scaling valves to
ensure proper CO2-partial pressure equilibration.
Other costs: For geomicrobiological work done by Prof. Dr. Wolfgang E. Krumbein (Biogeoma)
and µ-CT measurements (Prof. Freiwald, Erlangen) we apply for financial support for analyses
on the heterothrophic degradation of OM leading to CO2 production and in-situ destruction of
carbonate shells and µ-CT measurements of initial and degraded carbonates. In addition, ship
time for sampling the intertidal pelagial has to be paid to the University of Oldenburg with €
per day. Prof. Krumbein will also be involved in the interpretation of geomicrobiological
consequences of experimental results.
3.4.2
Personnel costs: The laboratory and field measurements will be performed by a PhD student,
and the modelling by a PostDoc, both according to German tariff system (TVöD)
Consumables: Disposables
glass ware
chemicals
(incl. for microsensors)
stable isotope tracers
radiotracers
Travel: Verbundmeeting in year 1, 2 and 3 (
), 2 international scientific meetings in last 2
years (
each), 2 field studies in year 2 and 3 (
each), regular visits with partners
Investment: The student will need a dedicated equipment for microsensor measurements,
consisting of a micromanipulator (Marzhäuser), a motorized stage (Faulhaber), a data-acquisition
board (National Instruments), a variety of shielded cables and a laptop with special software
(Labview, National Instruments), and special amplifiers for amperometric and potentiometric
microsensors (custom build in MPI).
Other costs: For the microsensor course we will have to supply additional microsensors, as
many will be broken in the learning phase. As more sensors than normal will be broken, and the
normal sensor production is interrupted, we will need to purchase sensors from a commercial
company to maintain the research in the group (over 20 scientists depend on microsensors). One
microsensor costs
Euro, so we allow 2 sensors to be broken per participant, as conservative
estimate.
3.4.3
Personal costs: Experimental work will be performed by a PhD student for three years, costs
according to AWI table. Additional modelling and data work by PIs and AWI staff at no cost.
Consumables: Laboratory measurements, glassware, disposables and chemicals. In the second
year the Polarstern cruise brings more costs.
Travel: BIOACID project meeting every year (
€). Two international conferences and
workshops each year (
€ each); in the second and third year more travelling to conferences
for presenting results. Field studies, Polarstern cruise; meeting with project participants at
laboratories.
204
vi. References
Accornero A, Manno C, Esposito F, Gambi MC (2003) The vertical flux of particulate matter in the polynya of Terra Nova Bay. Part II
Biological components. Antarct Sci 15:175-188
Alexandersson T (1979) Marine maceration of skeletal carbonates in the Skagerrak, North Sea. Sedimentology 26: 845-852.
Aller RC (1982) Carbonate dissolution in nearshore terrigeneous muds: the role of physical and biological reworking. J Geol 90: 79-95.
Al-Horani F, Ferdelman TG, Al-Moghrabi SM, and de Beer D (2005) Spatial distribution of calcification and photosynthesis in the
scleractinian coral Galaxea fascicularis. Coral Reefs 24:173-180.
Anderson JB (1975) Factors controlling CaCO3 dissolution in the Weddell Sea from foraminiferal distribution patterns. Mar Geol 19:315-332
Arp G, Reimer A, and Reitner J (1999) Calcification in cyanobacterial biofilms of alkaline salt lakes. Eur.J.Phycol. 34:393-403.
Arp G, Wedemeyer N, and Reitner J (2001) Fluvial tufa formation in a hard-water creek (Deinschwanger Bach, Franconian Alb, Germany).
Facies 44:1-22.
Behrends B, GA Goodfriend, Liebezeit G (2003). Amino acid dating of recent intertidal sediments in the Wadden Sea, Germany. Senckenberg.
marit. 32: 155-164.
Behrends B, G Hertweck, G Liebezeit, G Goodfriend (2005) Earliest Holocene occurrence of the soft-shell clam, Mya arenaria, in the
Greifswalder Bodden, southern Baltic. Mar. Geol. 216: 79-82.
Berg P, Roey H, Janssen F, Meyer V, Jørgensen BB, Huettel M, and de Beer D (2003) Oxygen uptake by aquatic sediments measured with a
novel non-invasive eddy correlation technique. Mar. Ecol. Prog. Ser. 261:75-83.
Berger WH (1970) Biogenous deep-sea sediments: fractionation by deep-sea circulation. Geol Soc Am Bull 81:1385-1402
Berner RA, Honjo S (1981) Pelagic sedimentation of aragonite: Its geochemical significance. Science 211:940-942
Bissett A, de Beer D, Schoon R, Shaaishi F, Reimer A, and Arp G (2007) Microbial mediation of tufa formation in karst-water creeks. Limnol
Oceanogr:Accepted.
Böttcher ME (1997) The transformation of aragonite to MnXCa(1-X)CO3 solid-solutions at 20°C: An experimental study. Mar Chem 57: 97106.
Böttcher ME (1997) Experimental dissolution of CaCO3-MnCO3 solid-solutions in CO2-H2O solutions at 20°C. I. Synthetic low-temperature
carbonates. Solid State Ionics 101-103: 1263-1266.
Böttcher ME (1999) The stable isotopic geochemistry of the sulfur and carbon cycles in a modern karst environment. Isotopes Environ. Health
Stud. 35: 39-61
Böttcher ME, Gehlken PL, Steele DF (1997) Characterization of inorganic and biogenic magnesian calcites by Fourier Transform infrared
spectroscopy. Solid State Ionics 101-103: 1379-1385.
Böttcher ME, Brumsack H-J, Dürselen C-D (2007) The isotopic composition of modern seawater sulfate: I. Coastal waters with special regard
to the North Sea. J. Mar. Sys. 67: 73-82.
Böttcher ME, Hespenheide B, and 8 co-authors (2000) Biogeochemistry, stable isotope geochemistry, and microbial community structure of a
temperate intertidal mudflat. Cont. Shelf Res. 20: 1749-1769.
Brasse S, Reimer A, Seifert R, Michaelis W (1999) The influence of intertidal mudflats on the dissolved inorganic carbon and total alkalinity
distribution in the German Bight, southeastern North Sea. J. Sea Res. 42: 93-103
Canfield DE, Thamdrup B, Kristensen E (2005) Aquatic Geomicrobiology. Elsevier Academic Press.
Cooper DJ, Watson AJ, and Nightingale PD (1996) Large decrease in ocean-surface CO-2 fugacity in response to in situ iron fertilization.
Nature 383:511-513.
de Beer D, Kühl M, Stambler N, and Vaki L (2000) A microsensor study of light enhanced Ca2+ uptake and photosynthesis in the reef-building
hermatypic coral Favia sp. Mar. Ecol. Prog.Ser. 194:75-85.
Dellwig O, Bosselmann K, Kölsch S, Hentscher M, Hinrichs J, Böttcher ME, Reuter R, Brumsack HJ (2007) Sources and fate of manganese in
a tidal basin of the German Wadden Sea. J. Sea Res. 57: 1-18
Denman KL, Völker C, Pena MA, Rivkin RB (2006) Modelling the ecosystem response to iron fertilization in the subarctic NE Pacific: The
influence of grazing, and Si and N cycling on CO2 drawdown. Deep-Sea Res II 53:2327-2352
Gattuso JP, Allemand D, and Michel F (1999) Photosynthesis and calcification at cellular, organismal and community levels in coral reefs: a
review on interactions and control by carbonate chemistry. Amer. Zool. 39:160-183.
Gattuso J-P, Pichon M, Delesalle B, Cqanon C, and Frankinouille M (1996) Carbon fluxes in coral reefs. I Lagrangian measurement of
community metabolism and resulting air-sea CO2 disequilibrium. Mar. Ecol. Prog. Ser. 145:109-121.
Golubic S, Schneider J (1979) Carbonate Dissolution. In Biogeochemical Cycling of Mineral-Forming Elements. In Trudinger PA und Swaine
DJ (eds.), 107-125. Elsevier.
Green MA and Aller RC (1998) Seasonal patterns of carbonate diagenesis in nearshore terrigenous muds: Relationship to spring phytoplankton
bloom and temperature. J Mar Res 56: 1097-1123.
Hertweck, G. and G. Liebezeit (1996). Biogenic and geochemical properties in intertidal biosedimentary deposits related to Mytilus beds.
P.S.Z.N. I: Mar. Ecol. 17:131-144.
Hertweck. G. and G. Liebezeit (2002). Historic mussel beds (Mytilus edulis) in the sedimentary record of a back-barrier tidal flat near
Spiekeroog Island, southern North Sea. Helgoland Mar. Res. 56: 51-58.
Hertweck, G., A. Wehrmann and G. Liebezeit (2006). Bioturbation structures of polychaetes in modern shallow marine environments and their
analogues to Chondrites group traces. Paleogeogr. Paleoceanogr. Paleoclimatol. 245: 382-389.
Hofmann AF, Meysman FJR, Soetaert K, and Middelburg JJ (2008) A step-by-step procedure for pH model construction in aquatic systems.
Biogeosciences 5:227-251.
Hohn S, Völker C, Wolf-Gladrow D (2008) A model of the carbon:nitrogen:silicon stoichiometry of diatoms based on metabolic processes.
Submitted to Mar Ecol Prog Ser
Hoppema M (2004) Weddell Sea is a globally significant contributor to deep-sea sequestration of natural carbon dioxide. Deep-Sea Res I
51:1169-1177
Hoppema M, Anderson LG (2007) Biogeochemistry of polynyas and their role in sequestration of anthropogenic constituents. In: Polynyas:
Windows to the World. Smith WO Jr, Barber DG (Eds.), Elsevier Oceanogr Ser, Amsterdam:193-221
Hoppema M, De Baar HJW, Bellerby RGJ, Fahrbach E, Bakker K (2002) Annual export production in the interior Weddell Gyre estimated
from a chemical mass balance of nutrients. Deep-Sea Res II 49:1675-1689
205
Hoppema M, Middag R, De Baar HJW, Fahrbach E, Van Weerlee EM, Thomas H (2007) Whole season net community production in the
Weddell Sea. Polar Biol 31:101–111
Hulth S, Tengberg A, Landén A, Hall POJ (1997) Mineralization and burial of organic carbon in sediments of the southern Weddell Sea
(Antarctica). Deep-Sea Res I 44:955-981
Hunt BPV, Pakhomov EA, Trotsenko BG (2007) The macrozooplankton of the Cosmonaut Sea, east Antarctica (30°E-60°E), 1987-1990.
Deep-Sea Res I 54:1042-1069.
Jansen H, Zeebe RE, Wolf-Gladrow DA (2002) Modelling the dissolution of settling CaCO3 in the ocean. Global Biogeochem Cycles 16:1-16
Kempe, S.; Pegler, K. (1991) Sinks and sources of CO2 in coastal sea: the North Sea. Tellus 43B, 224-235
Knauth-Köhler K, Albers BP, Krumbein WE (1996) Microbial mineralization of organic carbon and the dissolution of inorganic carbon from
mussel shells (Mytilus edulis). Senckenbergiana marit 26: 157-165.
Krumbein WE (1979) Photo lithotrophic and chemo organotrophic activity of bacteria and algae as related to beachrock formation and
degradation, Gulf of Aqaba, Sinai, Egypt. Geomicrobiol. J. 1:139-203.
Larkum AWD (1999) The cyanobacteria of coral reefs. Bulletin de l'Institute Oceanographique (Monaco) 0:149-167.
Ludwig R, Al-Horani FA, de Beer D, and Jonkers HM (2005) Photosynthesis controlled calcification in a hypersaline microbial mat.
Limnology & Oceanography 50:1836-1843.
Marshall J, Adcroft A, Hill C, Perelman L, Heisey C (1997) A finite-volume, incompressible Navier Stokes model for studies of the ocean on
parallel computers. J Geophys Res 102:5753-5766.
Morse JW & Mackenzie FT (1990) Geochemistry of sedimentary carbonates. Developments Sedimentol 48: 707 pp.
Olbers D, Borowski D, Völker C, Wolff JO (2004) The dynamical balance, transport and circulation of the Antarctic Circumpolar Current.
Antarct Sci 16:439-470
Orr JC et al (2005) Anthropogenic ocean acidification over the twenty-first century and its impact on calcifying organisms. Nature 437:681686.
Reise K (1998) Pacific oysters invade mussel beds in the European Wadden Sea. Senckenbergiana marit 28: 167-175.
Seeberg-Elverfeldt J, Schlüter M, Feseker T, Kölling M. (2005) Rhizon sampling of porewaters near the sediment-water interface of aquatic
systems. Limnol Oceanogr: Methods 3:361-371.
Shiraishi F, Reimer A, Bisset A, de Beer D, and Arp G (2008) Microbial effects on biofilm formation, ambient water chemistry and stable
sotope records in a highly supersaturated setting (Westerhöfer Bach, Germany). Palaeogeography Palaeoclimatology
Palaeoecology:accepted.
Soetaert K, Hofmann AF, Middelburg JJ, Meysman FJR, and Greenwood J (2007) The effect of biogeochemical processes on pH. Mar. Chem.
105:30-51.
Thomas, H, and 10 co-authors (2007) Rapid decline of the CO2 buffering capacity in the North Sea and implications for the North Atlantic
Ocean. Global Biogeochemical Cycles, 21
Visscher PT, Reid RP, Bebout BM, Hoeft SE, Macintyre IG, and Thompson JA (1998) Formation of lithified micritic laminae in modern
marine stromatolites (Bahamas): The role of sulfur cycling. American Mineralogist 83:1482-1493.
Wefer G, Balzer W, Bodungen B v, Suess E (1987) Biogenic carbonates in temperate and subtropical environments: Production and
accumulation, saturation state and stable isotopic composition. In: Rumohr J, Walger E, Zeitzschel B. (Eds.): Seawater-sediment
interactions in coastal waters. Berlin: Springer: 264-301.
Werner U, Blazejak A, Bird P, Eickert G, Schoon R, Abed R, Bissett A, and de Beer D (2007) Microbial photosynthesis in coral reef
sediments (Heron Reef, Australia). Estuar.Coast.Shelf Sci 76:876-888.
Wieland A, de Beer D, van Dusschoten D, Damgaard LR, Kuehl M, and van As H (2001) Fine-scale measurement of diffusivity in a microbial
mat with NMR imaging. Limnol. Oceanogr. 44:248-259.
Wolf-Gladrow DA, Bijma J, and Zeebe RE (1999) Model simulation of the carbonate chemistry in the microenvironment of symbiont bearing
foraminifera. Marine Chemistry 64:181-198.
Wollast R (1994) The relative importance of biomineralization and dissolution of CaCO3 in the global carbon cycle. Bull Inst Ocean, Monaco
13: 13-35.
Zeebe RE, Wolf-Gladrow DA (2001) CO2 in Seawater: Equilibrium, Kinetics, Isotopes. Elsevier Oceanography Book Series 65, Amsterdam,
346 pp
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Project 3.5: Impact of present and past ocean acidification on metabolism,
biomineralization and biodiversity of pelagic and neritic calcifiers
(PI: Adrian Immenhauser)
i. Objectives
The main aim of this collaborative research project is a detailed assessment of the past and future
performance (metabolism, biomineralization and biodiversity) of coastal/sessile and
oceanic/planktonic calcifyers exposed to CO2-induced ocean acidification. For this purpose, we
intend to combine observational (real-world) data from Cenozoic acidification events with such
obtained from experimental (culturing) work. We will study the effects of decreasing seawater
pH on groups that might potentially benefit (nannoplankton) and such that will most likely suffer
(bivalves) from acidification (see discussion in Iglesias-Rodriguez et al., 2008). With the
interdisciplinary approach applied here we will address the following six key questions: (1) How
sensitive are coastal and planktonic calcifyers to changing seawater pH? (2) What are the pH
threshold limits and are these limits universal or species dependent? (3) What is the impact of
changing seawater pH on biodiversity? (4) What is the adaptational potential of coastal and open
oceanic ecosystems in the time-scale of decennia to few centuries? (5) Are short-term (culturing)
experiments with calcifying organisms suitable analogues for pH/CO2 conditions predicted for
the end of the 21st century? (6) To which degree are Cenozoic acidification events suitable
analogues for predicted anthropogenic CO2 rise by 2100?
The present knowledge of CO2-induced ocean acidification (OA) is mainly based on modelling
and experimental (culturing) work providing valuable information on the response of specific,
short-lived organism exposed to pH/CO2 conditions predicted for the end of the 21st century (see
discussion in Fabry et al., 2008; Caldeira and Wickett, 2005 and references therein). This
approach, however, suffers from the inherent disadvantage related to the limited observational
time window. This shortcoming seriously compromises a prediction of the longer-term effects on
OA sensitive marine ecosystems and neglects the adaptational potential of natural systems. This
becomes particularly important as recent work controversially suggests that some groups of
calcifiers (mainly phytoplankton) might in fact benefit from increased CO2 partial pressure
(Iglesias-Rodriguez et al., 2008). An attempt to solve some of these controversies is the
investigation of past acidification events (Röhl et al., 2007; Pancost et al., 2007; and references
therein). The degree to which these pre-anthropogenic OA events are suitable analogues for
predicted scenarios in this century are, however, insufficiently understood (Zachos et al., 2001,
Iglesias-Rodriguez et al., 2008). It thus comes as a surprise that previous work combining the
strengths of past (observational) and future (experimental) effects of changing pH values on
marine calcifyers is at best scarce.
Perhaps the most prominent Cenozoic examples of transient climate perturbations are known
from the Palaeocene and Eocene intervals. This period of Earth’s history is punctuated by at least
four hyperthermals: the early Late Paleocene Event (58.4 Ma), the Paleocene-Eocene Thermal
Maximum (PETM; 55 Ma), the Early Eocene ELMO Event (53.3 Ma), and the Early Eocene XEvent (52.5 Ma; e.g., Nicolo et al., 2007). Negative δ13C excursions of 1-5‰ imply a massive
(ca. 2000 x 109 metric tons) input of 12C enriched carbon into the ocean-atmosphere system
probably in the form of methane. These events were marked by a general warming, at least 4x
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current CO2 concentrations, a low seawater pH and a shallow CCD (e.g., Zachos et al., 2005).
The rapid environmental changes are related to substantial changes in marine and continental
biota. But what was the impact of these past events on oceanic calcifyers and what is the
expected impact of anthropogenic CO2-induced OA by the end of the 21th century?
Nannoplankton (coccolithophores and some calcareous dinoflagellates) are among the most
abundant plankton organisms in the world’s oceans, and due to their capability of carrying out
both, photosynthesis and calcification, they have an impact on the organic and inorganic carbon
pump. They have been identified as key organisms for studying the impact of ocean acidification
on ocean ecosystems due to their debated response to elevated pCO2 concentrations (IglesiasRodriguez et al., 2008). Malformation and reduced calcite production per cell has been observed
for coccolithophores in culture and mesocosm experiments (Riebesell et al., 2000; Engel et al.,
2006). However, so far a direct proof for a response of nannoplankton to ocean acidification in
natural environments is lacking and short term disruptions of the carbonate system might not
reflect the observed natural changes in an adequate manner. Calcification in coccolithophores
may also be influenced by nutrient supply, temperature or salinity, and the effects of ocean
acidification could have positive or negative feedback effects on the natural variability. During
past ocean acidification events, coccolithophores as a group were capable to survive dramatic
changes in the carbonate system (Raffi et al. 2005; Gibbs et al. 2006, Medlin et al. 2008). In
comparison to other well studied plankton groups (e.g., foraminifera) research on nannoplankton
ecology, morphometry and geochemistry was previously limited by time consuming counting,
measuring and processing techniques. New technological developments are now available that
allow to overcome these problems.
Reef communities (corals, calcifying bivalves, macroalgae etc.) are one of the most OA sensitive
ecosystems. Due to their diagenesis-sensitive aragonitic skeletons, however, corals are poor
recorders of past acidification events. In contrast, sessile neritic bivalves (Mytilus, Artica etc.),
and particularly calcitic ones, are well-established, high-resolution archives of past and present
environmental parameters (Khim et al., 2000; Buick and Ivany, 2004; Immenhauser et al., 2005,
Rexfort & Mutterlose, 2006). Previous work, however, was severely limited by the lack of
stratigraphically continuous onshore sections providing data on pre-, syn- and post-OA events.
Newly discovered Palaeocene/Eocene sections in Oman allow now for a detailed study of coastal
section those are stratigraphically complete and hence allow for a time-resolved sampling of
bivalve hardparts across the intervals of interest.
Here, we propose to investigate OA sensitive sessile (bivalves) and nannoplanktonic
(coccolithophores and calcareous dinoflagellates) calcifiers across these past punctuated
acidification events and to compare results with cultured bivalves exposed to various pH, CO32
and CO2(aq) levels. Following this approach it is intended to combine the strengths of controlled
laboratory experiments with the biologically and geologically significant temporal element
provided by the fossil shell material. Furthermore, the combination of data from sessile neritic
organisms with such from open oceanic planktonic ones allows for a more holistic view of
problems related to ocean acidification.
3.5.1 (Immenhauser).- The Bochum research team has longstanding experience in the
investigation of mollusc shell material as sensitive archives of global change (e.g., Immenhauser
et al., 2002, 2005; Hippler et al., 2008 etc.) and the application of non-traditional isotope systems
(δ44Ca, δ26Mg; e.g. Buhl et al., 2007). Previous collaborative research projects involve initiatives
such as EUROCLIMATE as well as DFG and NOW financed projects involving cultured and
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fossil shell material. The collaboration with Prof. Schmahl and Dr. Griesshaber at the LMU
München further strengthens the research team by involving experts in shell material sciences
(Griesshaber et al., 2007; Schmahl et al., 2008).
3.5.2 (Mutterlose).- The applicant has a long going expertise in the study of calcareous
nannofossils from both fossil and recent settings. Findings from a study of calcareous
nannofossils across the PETM interval under discussion have been published recently
(Mutterlose et al., 2007). These clearly show the following results: (1) Significant changes in the
composition of the nannoplankton assemblages. (2) Occurrence of malformed taxa. (3)
Extinction of various, deep dwelling taxa. These changes in the composition of the primary
producers are thought to reflect an acidification of the ocean water during a period of extreme
warmth and very high CO2 concentrations. The collaboration with Dr. P. Schulte from the
University of Erlangen will add more specific expertise on sedimentological aspects.
3.5.3 (Meier).- The proponents have longstanding experience in calcareous nannoplankton
research. Their knowledge includes investigation of plankton samples, core top calibrations and
reconstruction of palaeoecology and past carbonate fluxes. Recently they were trained to operate
the automated coccolithophore recognition system SYRACO. The results from the analysis of
coccolithophore morphometry and weight estimates within the ESF MERF project (Quaternary
Marine Ecosystem Response to Fertilisation) show the influence of alkalinity and carbonate ion
concentration on size and weight of coccoliths from the Mediterranean Sea.
Subproject 3.5.1 Comparison of Cultured and Fossil Bivalve Geochemistry and Shell
Ultrastucture
(Immenhauser in collaboration with Schmahl, Griesshaber)
Molluscs are sensitive organisms recording environmental change in their biomineralogy, shell
geochemistry and shell morphology (Immenhauser et al., 2005). In addition, by forming highly
time-resolved growth increments they represent one of the accurate archives of coastal neritic
settings (e.g., Witbaard et al., 1994; Vander Putten et al., 2000). Culturing experiments of
bivalves in tanks provide the opportunity to expose specimens to environmental conditions
representative of an acidic ocean as predicted for the year 2100. Biomineralization (growth) rates
and modes, metabolic activity, preproduction cycles and live span can be assessed under these
circumstances. An obvious pitfall, however, is the short (months to at best years) observational
window provided by such experimental setups. Because of this bias, important parameters such
as adaptational patterns and the effects of longer term (decennia and more) exposure to stressed
environments cannot be assessed adequately. In order to overcome these problems, we here
propose a combined field and culturing approach using the arguably best ocean acidification
analogue of more recent history of our planet: the thermal maximum at the Palaeocene-Eocene
boundary (PETM). For this purpose, it is intended to link with project of T. Brey in the context of
BioAcid providing cultured shell material exposed to variable and increasing levels of OA to us.
This project proposed aims at investigating the shell ultrastucture and shell geochemistry of
cultured bivalves kept under pre-OA conditions and to compare these findings with comparable
data from fossil shell material obtained from neritic sections from Oman including the
Paleocene/Eocene record of hypothermals and acidification events.
The metabolic activity of bivalves influences the fractionation of both Mg and Ca isotopes
incorporated in the bivalves shell (e.g., Immenhauser et al., 2005; Hippler et al., 2008). Under
favourable environmental conditions, the shell isotopic fractionation is largely in concert with
209
seawater temperature and salinity fluctuations. The isotope fractionation–environment relation of
shell calcite and aragonite is lost when the bivalve is exposed to a stressed environment. As a
consequence, δ26Mg and δ44Ca isotope ratios are an indirect proxy for the environmental state
(favourable versus stressed) in modern and fossil bivalves (Hippler et al., 2008). Furthermore,
highly detailed investigations of the shell ultrastucture of bivalves (nm to µm) reveal specifically
different patterns in the micro-scale arrangement of the crystallites that build bivalve shells under
normal (aerobic) metabolism versus stressed (temperature, pH, salinity etc.) environments
(Griesshaber et al., 2007). Detailed investigations of the shell ultrastucture using REM, FIB,
TEM and EBSD, material properties and distribution of organic components (München and
Bochum) are planned. With those two highly novel tools: geochemical analyses (δ26Mg, δ44Ca;
geochemical laboratories RU Bochum) and shell ultrastucture (material sciences LMU
München), the assessment of the performance and mineralization mode of fossil bivalves can be
assessed and calibrated knowing the experimental conditions resulting from the culturing
experiments by the twin project of T. Brey (Project 2.1.3). Additional collaboration with other
groups involving bivalve shells will further strengthen this research 4.1.2 (Wahl).
In overview, sub-project 3.5.1 intends to investigate the metabolic activity, shell geochemistry
and shell ultra-structure of:
(1) cultured mollusc shells (cultured in tanks under variable pH, CO32 and CO2(aq)) in the context
of the twin-project by T. Brey; and
(2) comparison of these geochemical and shell ultrastucture data from fossil bivalves representing
pre-, syn- and post-OA during past Palaeocene and Eocene OA events. Here, the focus is on
calcitic molluscs because of their stable shell mineralogy (as opposed to diagenetically instable
coral aragonite), their expanded life time (10’s to 100’s of years), and their sessile, benthic
habitat, making them valuable analogues for coastal calcifying ecosystems; and
(3) to link results with data from twin-subprojects 3.5.2 and 3.5.3.
Schedule
3.5.1
First Year
I
Fine-tuning of analytical facilities,
(München/Bochum)
Fieldwork in Oman (collection of OArelevant shell material)
CO2 perturbation culturing experiments
in Bremen (project T. Brey)
Analytical work in Bochum (nonconventional isotopes)
Analytical work in München (shellultrastucture)
Possible second field period (collection
of OA-relevant shell material)
data interpretation
210
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Milestones (3.5.1)
- All analytical facilities are fine-tuned to the project proposed
month 4-6
- Fieldwork phase I completed, sample material available
month 2-3
- Experimental (cultured) shells from Bremen available
month 10-12
- Possible fieldwork phase II completed
month 15
- Data set on shell ultrastucture and geochemistry available
month 20
- Comparison of results with those of partner projects
month 24
month 33
- Completion of project
month 36
Subproject 3.5.2 Biological response to short-termed ocean acidification events in the past:
biodiversity and evolution patterns of marine primary producers (calcareous nannofossils)
during the late Paleocene – early Eocene
(Mutterlose in collaboration with Schulte)
Work Programme
The applicants plan to study the calcareous nannofossils, stable isotope (δ13C, δ18O, δ11B)
patterns and the sediment petrography of the Late Palaeocene (NP 5) to Early Eocene (NP 12) of
an ODP Site in order to contribute to the understanding of the following questions:
(1) Does the isotope record provide evidence for δ13C excursions additional to the PETM?
(2) What is the sequence of nannofossil patterns (diversity, abundance, size evolution, extinction
and origination patterns) for the Palaeocene/Eocene hyperthermals?
(3) Do the excursion floras of the PETM reflect an acidification of the ocean waters?
(4) Are the Palaeocene/Eocene hyperthermals characterized by similar turnovers like the PETM?
(5) Were T°C, nutrients, CO2, pH or salinity controlling the composition of the assemblages?
(6) What are the concomitant sedimentological and mineralogical changes?
(7) What are the paleoceanographic and palaeoclimatic implications of these findings?
(8) Are these biotic changes a potential scenario for the future of our oceans?
For this purpose it is here proposed to investigate the isotopic composition (δ13C, δ18O, δ11B) of
calcareous nannofossils, and the mineralogy of the Late Palaeocene/Early Eocene interval from a
low latitudinal ODP Site. The pre- and post intervals of the hyperthermal events will be analysed
at a sample spacing of 5 – 10 cm as defined by the isotope stratigraphy. The events itself will be
studied with a spacing of at least 1 sample/5 cm, critical intervals at a resolution of 1 sample/1
cm.
211
Oxygen and carbon isotope data of benthic foraminifera will be analysed to construct a highresolution isotope stratigraphy and establish a long term palaeo-temperature record. The isotope
data will also be used for (1) gaining information on palaeo-environmental changes, (2) defining
the perturbations of the carbon cycle during the investigated interval, and (3) understanding the
chemical composition of potentially different water-masses through time. It is also attempted to
use the boron isotope record (δ11B) of foraminifera tests as a proxy for potential pH changes
About 400 samples will be analysed for calcareous nannofossils. In each settling slide 300-400
specimens will be counted in order to record the biodiversity and the absolute abundance of all
taxa encountered. The data obtained will include information about preservation and etching;
ranges of index taxa; diversity and abundance patterns; absolute abundances of nannofossils per
gram sediment; changes in the nannofossil assemblages; extinction/ origination of species; and
onset of malformed taxa. The dataset will be analyzed with respect to single species carbonate
content using the SYRACO system at Kiel University.
Grain size analysis of the <63 mm fraction will give evidence for either aeolian or hemipelagic
origin of the sediments. X-ray diffractometry of 200-250 samples will provide data of the bulk
rock and clay mineralogy, thus supplying information about the climatic regime (arid/humid).
The magnetic susceptibility (MS) will be determined (400 samples) to generate a high-resolution
MS-curve for the Late Palaeocene/Early Eocene. Variations of ferromagnetic and paramagnetic
minerals reflect climatically-induced changes in the depositional realm. This because mineralogy,
concentration and grain size of iron minerals are related to environmental conditions
The results of this study are potentially relevant for the projects involving modelling (1.3
Schneider, 5.2 Oschlies). The subprojects 1.1.3 (Müller), 1.1.4 (Reusch) and 3.1.1 (Schulz) cover
modern analogues of our past scenario of phytoplankton evolution and climate change, we thus
see the chance to bridge the gap between past-present-future by a contrast comparison of these
data.
Schedule
3.5.2
First Year
I
Sampling in Bremen
Mineralogical analyses
Magnetic data acquisition
Preparation and evaluation of smear
slides
Evaluation settling slides
Isotope studies
SEM studies
Evaluation of data/synthesis
212
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Milestones (3.5.2)
- Low resolution study samples analysed for calcareous nannofossils
month 12
- Magnetic data and mineralogical analyses completed
month 12
- High resolution study samples analyzed
month 26
- Isotope studies completed
month 24
month 24
- Compilation of study
month 36
Subproject 3.5.3 Nannoplankton response to modern and past ocean acidification events
(Meier in collaboration with Kinkel)
Work Programme
Subproject 3.5.3 makes use of fossil nannoplankton in the oceanic sediment record providing the
largest archive of any fossil organism group throughout the Mesozoic and Cenozoic (Young
1998; Kinkel et al., 2000; Bown et al., 2004; Baumann et al., 2005). The amount of calcite stored
in coccoliths or coccospheres can be estimated using light-optical measurements (Beaufort 2005).
For this the automated recognition system SYRACO (Beaufort & Dollfus, 2004) will be installed
to generate species specific data on morphometry and weight estimates of key taxa from different
OA events in Earth’s history. SYRACO provides data for the calcite stored in each coccolith or
coccosphere and can therefore be used to identify heterogeneity in calcite production between
individual liths and cells. The system reliably recognises the dominant modern species and can be
trained to recognise additional taxa. Due to the large amount of samples SYRACO can analyse
(about 50 slides per day), an unprecedented spatial and temporal resolution will be reached, and
statistically highly significant data will be produced. Parallel SEM investigations will allow
control on possible different genotypes, as well as dissolution or diagenetic overprint in fossil
assemblages. With this approach, the following hypotheses will be tested: Does single species
calcification depend on various environmental factors as opposed to ocean acidification alone?
and Does ocean acidification affect specific taxa only? By investigating single species response
to OA as well as generating estimates on total calcareous nannoplankton carbonate production it
is planed to identify possible common patterns of calcareous nannoplankton community reactions
to OA events. Recently Iglesias-Rodriguez et al. (2008) have challenged the assumption that OA
will lead to a reduction of coccolithophore calcification. However, their record only covers the
past 200 years and lacks control on species specific carbonate production. Thus it is not clear,
whether the observed increase in coccolithophore calcification in the past decades is a response to
anthropogenic CO2 emissions or reflects natural variability, and what the species specific
response is.
Therefore, the project aims to investigate natural variability versus the impact of OA and possible
climate feedback in three different scenarios: (1) Holocene (i.e. the last 10’000 years) during
times of minimum variability of the carbonate system. (2) Termination of the penultimate
glaciation (i.e. ~132’000 years B.P.), during which a doubling of the pCO2 concentration
occurred and is coupled with a strong and fast climatic change. (3) Paleocene/Eocene Thermal
Maximum (~55 million years ago), that is characterised by extreme OA events that led to
extinction of some species, whilst so-called disaster taxa including some coccolithophores and
213
many calcareous dinoflagellates show relatively little response to the OA event. For all three
scenarios, species specific carbonate weight estimates and morphometric data will be generated.
Measurements will also be carried out on cells and liths obtained from culturing studies on
coccolithophores and dinoflagellates proposed in BIOACID (subprojects 1.1.3 Müller, 1.1.4
Reusch, 3.1.1 Schulz and 4.2.2 Rost) to compare our observed nannoplankton response to natural
OA events to those created in the laboratory. Our results can also be of significance for those
projects investigating how pelagic calcification should be modelled (1.3 Schneider, 5.2 Oschlies).
All necessary samples for our project are already available (Holocene, Termination II) at our
institute or will be provided by partners within BIOACID (PETM, cultures).
Schedule
3.5.3
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Installation of SYRACO
Training of PhD student and SYRACO
Analysis of Holocene samples
Analysis of Termination II samples
Training of PhD student and SYRACO
for PETM species
Analysis of PETM samples
SEM control of sample quality
Continuous monitoring of culture
species
Image and data analysis, interpretation
Manuscript preparation, presentation at
conferences
Milestones (3.5.3)
- SYRACO operational
month 03
- PhD student trained in nannoplankton taxonomy
month 03
- Holocene dataset compilation finished
month 12
- Termination II dataset compilation finished
month 21
month 24
- PETM dataset compilation finished
month 30
- Project synthesis
month 36
214
First Year
Second Year
Third Year
Total
Personnel costs
3.5.1
3.5.2
3.5.3
€, PhD
€, HiWi
€, PhD
€, HiWi
€, PhD
€, HiWi
Subtotal
Consumables
3.5.1
3.5.2
3.5.3
Subtotal
€
(Analytical work
Bochum &
München)
€
(Analytical work
Bochum &
München)
€
(Analytical work
Bochum &
München)
€
€
(Isotopes etc,)
(Isotopes etc,)
€
€
€
(SEM etc.)
(SEM etc.)
(SEM etc.)
€
€
€
€
€
€
(Fieldwork,
meetings)
(Fieldwork,
meetings)
€ (Meetings,
workshops)
€
€
€
€
(Kiel, meetings)
(Kiel, meetings)
(Kiel, meetings)
€
€
(Training,
meetings)
(Training,
meetings)
€
€
€
€
€
n.a.
n.a.
n.a.
n.a.
€
n.a.
n.a.
n.a.
n.a.
€
(Isotopes etc,)
€
€
€
Travel
3.5.1
3.5.2
3.5.3
Subtotal
€ (Training,
meetings)
€
Investments
3.5.1
3.5.2
3.5.3
(camera micros)
€
(SYRACO)
€
€
215
First Year
Subtotal
Second Year
Third Year
Total
€
n.a.
n.a.
€
3.5.1
n.a.
n.a.
n.a.
n.a.
3.5.2
n.a.
n.a.
n.a.
n.a.
3.5.3
n.a.
n.a.
n.a.
n.a.
Subtotal
n.a.
n.a.
n.a.
n.a.
Other costs
TOTAL
3.5.1
Personnel costs: The research is ambitious and technically complex and is to be carried out by a
capable PhD student paid according to the financial regulations of the Ruhr-University Bochum.
Consumables: The combined geochemical and material properties research is cost intensive.
Coast for a δ26Mg isotope analysis are in the order of
€ depending on the number of
repetitions made. In general, each data point represents a total of five repeat analyses. The same
accounts for calcium isotope analysis. Shell ultrastucture work as carried out in collaboration
with München is both time and cost intensive. Hence most of the finances go into consumables
for analytical work (reference gas, high-precision thin sections etc.). In order to support the PhD
with time consuming work, a HiWi is hired for a moderate number of hours per week.
Travel: The PhD should participate at workshops and attend conferences. Fieldwork in Oman
and probably at other PETM onshore localities is performed.
Investment: Not applicable.
Other costs: Not applicable.
3.5.2
Personnel costs: The research will be performed by a PhD student paid according to the
financial regulations of the Ruhr-University Bochum.
Consumables: This money is requested for running the various analyses (stable isotopes, clay
min. etc.) suggested in the proposal. This will also cover the running costs for glass war,
adhesives etc.
Travel: We scheduled a) once per year a 3 weeks visit of the Bochum PhD student to Kiel, to
work on the automated microscope to gain data and discuss results; b) once per year a meeting of
the BioAcid group in Kiel, c) two short visits per year to Erlangen, d) at least one international
meeting per year to present findings.
Investment: For measuring the sizes of calcareous nannofossils a new microscope camera is
needed.
216
3.5.3
Personnel costs: The research will be carried out by a PhD student who will be paid according to
the German tariff (
- € per year). A student worker is requested for assistance
with operating the automated system, sample preparation and image analysis ( ,- € per year).
Consumables:
SEM analyses, about 150 hours at
SEM consumables
Glass slides, embedding material, filters
Travel: Training in Aix en Provence (1st year), regular meetings within partners (1st-3rd year),
international conferences (2nd & 3rd year).
Investment: The automated recognition system SYRACO is a prerequisite for generating single
species carbonate production estimates. The system can analyse up to 50 samples per day (for
comparison, traditional nannoplankton PhD studies would be based on a few hundred samples).
Considering the large number of samples to be analysed as proposed conventional counting and
imaging techniques are too slow. The system contains an automated microscope (
), a
high resolution b/w camera ( ), and a computer with monitor ( ).
vi. References
Agnini C, Fornaciari E, Raffi I, Rio D, Röhl U, Westerhold T (2007) High resolution nannofossil biochronology of middle Paleocene to early
Eocene at ODP Site 1262: Implications for calcareous nannoplankton evolution. Mar Micropaleontol 64:215-248
Baumann KH, Andruleit H, Boeckel B, Geisen M, Kinkel H (2005) The significance of extant coccolithophores as indicators of ocean water
masses, surface water temperature, and paleoproductivity: a review. Pal Z 79:93-112
Beaufort L (2005) Weight estimates of coccoliths using the optical properties (birefringence) of calcite. Micropaleontology 51:289-297
Beaufort L, Dollfus D (2004) Automatic recognition of coccoliths by dynamical neural networks. Mar Micropaleontol 51:57–73
Bown PR, Lees JA, Young JR (2004) Calcareous nannoplankton evolution and diversity through time. Coccolithophores - from molecular
processes to global impact. H Thierstein and J R Young. Berlin, Springer: 481-508
Buick DP, Ivany LC (2004) 100 years in the dark: Extreme longevity of Eocene bivalves from Antarctica. Geology 32:921-924
Calderia K, Wickett ME (2005) Ocean model predictions of chemistry changes from carbon dioxide emissions to the atmosphere and ocean. J
Geophys Res 110: doi:10.1029/2004JC002671
Hippler D, Witbaard R, Richter D, Buhl D, Immenhauser A (2008) Inter- and intra-species variability of Mg isotopes in marine biogenic
carbonates. Geochim Cosmochim Acta in press
Engel A, Zondervan I, Aerts K, Beaufort L, Benthien A, Chou L, Delille B, Gattuso JP, Harlay J, Heemann C, Hoffmann L, Jacquet S,
Nejstgaard J, Pizay MD, Rochell-Newall E, Schneider U, Terbrueggen A, Riebesell U (2005) Testing the direct effect of CO2
concentration on a bloom of the coccolithophorid Emiliania huxleyi in mesocosm experiments. Limnol Oceanogr 50:493-507
Fabry VJ, Seibel BA, Feely RA, Orr JC (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICES Journal of
Marine Science 65:414-432
Gibbs S, Bralower TJ, Bown PR, Zachos JC, Bybell LM (2006) Shelf and open-ocean calcareous phytoplankton assemblages across the
Paleocene-Eocene Thermal Maximum: Implications for global productivity gradients. Geology 34:233-236
Gibbs SJ, Bown PR, Sessa JA, Bralower TJ, Wilson PA (2006) Nannoplankton extinction and origination across the Paleocene-Eocene
Thermal Maximum. Science 314:1770-1773
Griesshaber E, Schmahl WW, Neuser RD, Pettke T, Blüm M, Mutterlose J, Brand U. (2007) Crystallographic texture and microstructure of
terebratulide brachiopod shell calcite: An optimized materials design with hierarchial architecture. Am Mineral 92:722-724
Iglesias-Rodriguez MD, Halloran PR, Rickaby REM, Hall IR, Colmenero-Hidalgo E, Gittins JR, Green DRH, Tyrell T, Gibbs SJ, Dassow vP,
Rehm E, Arbrust EV, Boessenkool KP (2008) Phytoplankton Calcificaiton in a High-CO2 world. Science 320:336-340
Immenhauser A, Kenter JAM, Ganssen G, Bahamonde JR, van Vliet A, Saher MH (2002) Origin and significance of isotope shifts in
Pennsylvanian carbonates (Asturias, NW Spain). J Sediment Res 72:82-94
Immenhauser A, Nägler TF, Steuber T, Hippler D (2005) A critical assessment of mollusk 18O/16O, Mg/Ca, and 44Ca/40Ca ratios as proxies
for Cretaceous seawater temperature seasonality. Palaeogeogr Palaeoclimatol Palaeoecol 215:221-237
Khim BK., Woo KS, Je JG (2000) Stable isotope profile of bivalves shells: seasonal temperature variations, latitudinal temperature grdaients
and biological carbon cycling along the east coast of Korea. Cont Shelf Res 20:843-861
Kinkel H, Baumann KH, Cepek M (2000) Coccolithophores in the equatorial Atlantic Ocean: response to seasonal and Late Quaternary
surface water variability. Mar Micropaleontol 39:87-112
Medlin LK, Sáez AG, Young JR (2008) A molecular clock for coccolithophores and implications for selectivity of phytoplankton extinctions
across the K/T boundary. Mar Micropaleontol 67:69-86
217
Mutterlose J, Linnert C, Norris R (2007) Calcareous nannofossils from the Paleocene-Eocene Thermal Maximum of the equatorial Atlantic
(ODP Site 1260B): Evidence for tropical warming. Mar Micropalaeontol 65:13-31
Nicolo MJ, Dickens GR, Hollis CJ, Zachos JC (2007) Multiple early Eocene hyperthermals: Their sedimentary expression on the New Zealand
continental margin and in the deep sea. Geology 35:699-702
Pancost RD, Steart DS, Handley L., Collinsion ME, Hooker JJ, Scott AC, Grassineau NV, Glasspool IJ (2007) Increased terrestrial methane
cycling at the Palaeocene-Eocene thermal maximum. Nature 449:332-335
Raffi I, Backman J, Pälike H (2005) Changes in calcareous nannofossil assemblages across the Paleocene/Eocene transition from the paleoequatorial Pacific Ocean. Palaeogeogr Palaeoclimatol Palaeoecol 226:93–126
Rexfort A, Mutterlose J (2006) Oxygen isotope of Sepia officinalis – a key to understanding the ecology of belemnites? Earth Planet Sci Lett
247:212-221
Riebesell U, Zondervan I, Rost B, Tortell PD, Zeebe RE, Morel FM (2000) Reduced calcification of marine plankton in response to increased
atmospheric CO2. Nature 407:364-367
Röhl U, Westerhold T, Bralower TJ, Zachos JC (2007) On the duration of the Paleocene-Eocene thermal maximum (PETM). Geochem
Geophys Geosyst 8:doi: 10.1029/2007GC001784
Schmahl WW, Griesshaber E, Neuser RD, Merkel C, Deuschle J, Götz A, Kelm K, Mader W, Sehrbrock A (2008) Crystal morphology and
hybrid fibre composit architecture of phosphatic and calcitic brachiopod shell materials- an overview. Mineral Mag in press
Vander Putten E, Dehairs F, Keppens E, Bayens W (2000) High resolution distribution of tracce elements in the calcite shell layer of modern
Mytilus edulis: Environmental and biological controls. Geochim Cosmochim Acta 64:997-1011
Witbaard R, Jenness MI, van der Borg K, Ganssen G (1994) Verification of annual growth increments in Arctica islandica L. from the North
Sea by means of oxygen and carbon isotopes. Neth J Sea Res 33:91–101
Young JR (1998). Neogene. Calcareous Nannofossil Biostratigraphy. PR Bown. Dordrecht, Kluwer Academic: 304-322
Zachos JC, Pagani M, Sloan L, Thomas E, Billups K (2001) Trends, Rhythms, and Aberrations in Global Climate 65 Ma to Present. Science
292:686-693
Zachos JC, Röhl U, Schellenberg SA, Sluijs A, Hodell DA, Kelly DC, Thomas E, Nicolo M, Raffi I, Lourens LJ, McCarren H, Kroon D
(2005) Rapid acidification of the ocean during the Paleocene-Eocene thermal maximum. Science 308:1611-1615
218
11.5: Theme 4: Species interactions and community structure in a
changing ocean
Regime shifts are rapid reorganizations of ecosystems from one relatively stable state to another.
Ecosystem regime shifts may not always be smooth, but can be abrupt resulting from complex,
non-linear processes (Scheffer et al. 2001) that are driven by specific alterations in the interaction
strength or mode between key species. For example, alternative stable states of coastal benthic
communities can be triggered by pathogens or altered predator prey relationships of key trophic
interactions (Sanford 1999). As such, relatively small changes in species composition may
transform ecosystems from consumer to producer-dominated communities and vice versa
(Scheibling 1986). To date, regime shifts have been often linked to climate forcing, e.g.
fluctuations in the North Atlantic oscillation, El Niño events, or global climate change. For
example, several warm water species have taken advantage of the warmer waters and expanded
their ranges polewards (Hays et al. 2005), whereas other, cold-water adapted, species have
retreated in the same direction (Perry et al. 2005). These range shifts had strong negative effects
on the recruitment of higher trophic levels (e.g. fish and seabirds) that feed on key boreal species,
such as cod feeding on the calanoid copepod Calanus finmarchicus (Beaugrand et al. 2003).
Regime shifts and other structural re-organizations of marine communities are caused by diverse
and imbalanced responses of different species or guilds to a changing environment. Even within
species, susceptibility to stress can differ between genotypes and between different ontogenetic
stages. This variability of stress effects at different organisational levels may lead to changes in
trophic interactions or competitive hierarchies (Tortell et al. 2002), and result in a conspicuous
shift in structural and functional properties of a community.
Not only temperature, but also ocean acidification (OA) is a potential stress factor with a diverse
set of individual reactions. As a result of the increasing pCO2, not only the stress related to
acidification, but also the change in the availability of different nutrients (Si, N, P) in relation to
carbon will change the competitive relationships between primary producers, especially in the
light of the reduction in anthropogenic input of phosphorus and nitrogen in many coastal seas.
Moreover, OA is accompanied by increases in water temperatures of ~2 to 5°C in many
temperate regions. Therefore, studying the impacts of OA on community level biological
interactions and cascades through the foodweb needs to consider potential synergisms between
increases in temperature and decreases in ocean pH. Temperature increase will, among others,
accelerate the metabolic rates of producers, consumers and remineralizers, whereas acidification
may, among other effects, negatively affect the nutritional quality of primary consumers and
detritus. Hence, the likelihood for consumer-prey mismatch situations may increase. In the
marine benthos, for example, macroalgae and seagrasses, which often lack efficient carbon
concentration mechanisms may benefit from increased pCO2 availability. In contrast, benthic
filter feeders such as bivalves, bryozoans, corals or tube building polychaetes may suffer declines
in their calcifying capacity (Shirayama & Thornton 2005), with concomitant decreases in
population growth and persistence. Along rocky shores in particular, where marine filter feeders
directly compete with marine plants for space, a climate change driven restructuring may shift
communities from filter feeder dominated consumer communities to macroalgae dominated
communities.
219
In short, ocean acidification may have important consequences for the biodiversity and
functioning of marine ecosystems: even moderate stress impacts on individual species may be
dramatically amplified (or buffered) by ensuing modulation of niches and interactions which
ultimately will provoke a restructuring of communities. As a consequence, important ecosystem
services (e.g. O2 production, CO2 sequestration, remineralization of nutrients, fisheries yields)
may be jeopardized. Extending our understanding of stress impacts from the individual or species
level to the level of interactions and community dynamics is crucial.
The main gaps in our knowledge
Our knowledge on the effects of OA has increased considerably in recent years, but there are still
enormous gaps that need to be filled with regard to complex community responses. This
knowledge is critical to assess the impacts of OA on all scales as well as to provide policy makers
with the tools and information necessary to implement the necessary actions. One of the main
gaps concerns the effect of OA on the interactions between species and the resulting community
structure. Whereas many studies exist on the effects of low pH and/or high CO2 conditions on
single organisms, not much is known about how these factors affect interactions between species
and, through that, communities. Indeed, the predicted shifts in pCO2 and pH in many species will
often only slightly impact performance and fitness of a given species. However, as shown for
other stressors (Christensen et al. 2006, Wahl 2008) ensuing modifications of species interactions
may substantially amplify or buffer the original stress (Wahl et al. 2004). How environmental
stress spreads through a community via shifts in composition and interaction is the main focus of
this theme.
We define two central research questions dealing with interactions between organisms. (1) First,
we ask how competitive interactions will change in a high CO2 world, and whether and how this
may lead to alterations in the structure and functioning of communities. Species competing for
similar resources may have completely different requirements to the physical environment. Most
obviously, calcifying organisms may be more affected by a decrease in pH than non-calcifying
organisms, and this should lead to a change in the competitive interactions between the two (Fig
4.1d). However, we can also envisage more subtle effects where slight differences in for example
carbon concentrating mechanisms may affect the competitive outcome between or even within
species. This question is very important with regard to highly diverse bacterial communities and
their key role as bioreactor for the remineralization of nutrients. Additionally, fitness shifts in
macroorganisms and/or compositional changes in the colonizer pool will lead to altered epibiotic
biofilms indirectly impacting the biotic and abiotic interactions of the host. Increasing CO2 levels
will also increase the C:nutrient ratios available for primary production, and as these nutrient
ratios can be considered an ecological niche (Loladze et al. 2004) we expect structural shifts in
algal communities. We will investigate these competition mediated effects at different functional
levels including macroalgae, benthic invertebrates, microalgae, and bacteria as we expect impacts
to differ between these functional groups. Since further differences are expected between
genotypes within species, microevolutionary aspects are also included.
220
10
b
0.8
0.6
9
8
0.4
nutrient replete
High C:N
High C:P
7
6
0.2
5
0
1
2
3
4
5
6
7
8
Respiration (µg O2 min-1 Animal-1)
Day of incubation
1.2
0
1e+5
2e+5
3e+5
4e+5
Rhodomonas salina (Ind ml-1)
c
d
70
60
Crustose coralline algae
Non-calcifying algae
1.0
0.0
6e+5
5e+5
50
0.8
40
0.6
30
0.4
20
0.2
Percentage cover
200 ppm
600 ppm
900 ppm
11
C:N Ratio (molar)
a
-1
1.0
12
Oxyrrhis marina growth rate (d )
10
0.0
0
nutrient replete High C:N
High C:P
Treatment of the food
400
765
CO2 treatment (ppm)
Fig. 4.1: Examples of previous relevant work. a) nutrient stoichiometry of Rhodomonas salina under different CO2
concentrations (Gülzow 2008); b) Oxyrrhis marina growing on food with different nutrient stoichiometry (Hantzsche
& Boersma submitted); c) Respiration of Acartia tonsa on different food sources (Boersma et al. unpubl); d)
competitive interactions between calcifying algae and non-calcifying algae under different CO2 conditions (Kuffner et
al. 2007)
(2) The second major aim of this theme is to elucidate the effects of OA on food web
interactions. Obviously, loss of species or the replacement of one species (strain) with another
one (diatoms versus dinoflagellates) as a result of OA will affect the flow of energy and matter
through the food web, but more subtle processes may be just as relevant. High CO2 availability to
primary producers may affect their quality as food for herbivores. Not only do we know that the
production of secondary metabolites is highly dependent on the nutrient conditions (Velzeboer et
al. 2001, Lippemeier et al. 2003), we also know that the concentration of essential components of
the food such as fatty acids is dependent on the growing conditions of the algae (Müller-Navarra
1995, Boersma 2000). Moreover, the stoichiometry of the macronutrients may change in primary
producers as a result of high CO2 availability (Fig 4.1a and: Burkhardt et al. 1999, Riebesell et al.
2000, Taraldsvik & Myklestad 2000, Urabe et al. 2003, Riebesell et al. 2007). As primary
producers are not homeostatic with respect to their nutrient composition, they usually reflect the
nutrient composition of their surrounding water, and hence are being expected to also show
higher carbon-to-nutrient ratios. High carbon-to-nutrient algae are known to be inferior food
quality for herbivorous organisms (Fig 4.1b and: Boersma 2000, Sterner & Elser 2002, Boersma
& Elser 2006). These consumers have to handle more energy (carbon) in relation to nutrients
needed for their assembly and acquisition machinery (phosphorus rich ribosomes and nitrogen
rich proteins and mitochondria, respectively) (Klausmeier et al. 2004), which creates costs and
leads to altered nutrient stoichiometry and reduced growth and reproduction of the consumers.
These food quality effects have only recently been shown to travel up food chains to predatory
221
beetles in terrestrial systems (Kagata & Ohgushi 2007), to parasites feeding/living on nutrient
limited hosts (Frost et al. 2008) and even to planktivorous fish (Malzahn et al. 2007a). The latter
finding is very important, as it implies that through OA, and the resulting increase in the
C:nutrient content of the algae (especially in the light of the re-oligotrophication of several
coastal seas) the production of zooplankton should decline as a result of lower food quality.
Moreover, the quality of this reduced stock of zooplankters for higher trophic levels should be
low, thus potentially impacting production of economically important fish species.
Changes in nutrient stoichiometry on all levels from water chemistry to secondary consumers are
likely to impact the community structure and functioning of microbial communities as well. Not
only as a result of direct stress effects, but also as a result of changes in excretory products of
consumers as a result of their homeostasis which will affect the available carbon pool for bacteria
(Fig 4.1c, different respiration rates under different food conditions). This is likely to favour new
species assemblages, and hence, bacterial community functioning.
Consequently, this theme comprises investigations of the horizontal (competitive interactions)
and vertical (food chains) translation of abiotic stress into structural changes of the benthic and
pelagic, microbial and macrobial communities considered (See Fig. 4.2 for an example in the
benthic environment).
Abiotic Stress
acidification, warming
Defence
Consumers
Food quality
Recruitment
Compensatory
growth
Competitors
Fucus
Growth
Epibiosis
Abundance &
Depth
distribution
Fig. 4.2: Horizontal and vertical interactions with the macroalga Fucus as an example of a target species: Abiotic stress
affects directly but often with different strength and/or sign the performance of interactors (target species Fucus, its
consumers, competitors and epibionts). The ensuing shifts in biotic interactions may have still stronger effects on the
survival and distribution of the target species than the direct stress itself.
Objectives
By means of manipulative experimentation from the individual up to the community level we
will:
1. Establish the role of differential sensitivities to OA at the community, species and
subspecies (ontogenetic stages, genotypes) level
2. Assess the synergistic effects of acidification and warming
222
3. Analyse changes in food quality and quantity of primary producers under various CO2
concentrations
4. Quantify the ensuing shifts in competitive abilities of benthic and pelagic model organisms
5. Quantify the ensuing shifts in the performance and ecological efficiency of energy transfer
between lower and higher trophic levels in pelagic and benthic systems
6. Assess and model cascading effects in coastal marine systems from secondary consumers
down to microbial communities via trophic, competitive and epibiotic interactions
7. Compare the effect of natural and experimental stress gradients on community structure
and dynamics
8. Compare the relative impact of acidification/warming stress on different types of
communities (benthic – pelagic, microbial – macrobial).
Approach
Different types of experimental designs will be applied to address changes in community
structure, competitive interactions as well as nutritionally mediated responses in consumers over
several trophic levels. The first type of setup will consist of using controlled laboratory systems
that subject organisms to different levels of acidification (and warming) and investigate how
sensitivities differ between genotypes, ontogenetic stages, and/or species and communities.
Secondly, primary producers cultured under different conditions will be fed to consumers in
sequential steps. This will allow quantification of the immediate impact of prey quality on the
performance of secondary producers using advanced analytical methods to determine food
quality parameters. The third type of experiments will consist of environmental samples from
natural pH or CO2 gradients and larger scale in-door mesocosm setups within walk-in
temperature controlled rooms where both temperature and several levels of acidification can be
manipulated in a factorial design. This setting can be used for both pelagic and benthic
community model systems, to study changes in community structure and interaction under
different CO2/temperature scenarios.
This theme involves two subprojects, both dealing with the gaps in our current knowledge
described above. One project (4.1) investigates processes in the benthic system, whereas the other
one (4.2) studies pelagic processes. Both projects comprise a number of complementary
subprojects, which differ with regard to their focus on community type, habitat and/or interaction
type. Within projects, the common habitats allow joint experiments, and combined sampling
campaigns. Between the projects the joint questions allow us the development of common
strategies and concepts. Moreover, investigating identical or similar questions over a range of
differently sized organisms with different generation times and reproduction modes also enables
us to test the robustness and generality of our findings as well as the dependency of our results on
the differences between systems. Strong collaborations will exist with different projects in the
other themes, more specifically with those projects investigating single species responses to OA
in themes 2 and 3, and the microbiological projects in theme 1.
223
Project 4.1: OA impacts on interactions in and structure of benthic communities
(M. Wahl)
i. Objectives
To assess the impact of ocean acidification (for some experiments in conjunction with predicted
warming) on
• performance of genotypes and populations
• competitive interactions within and between species
• associations such as epibiosis
• structure and function of communities
• microbial as compared to macrobial communities
Ocean acidification (in conjunction with warming) effects in benthic habitats can be found at
very different levels of organisation. Thus, the expected abiotic shifts may impact the activity of
important enzymes, calcification processes and the general performance or fitness of individuals
(Saborowski et al. 2004, Fabry et al. 2008). Such organismic responses to climate change are
conventionally categorized as either ecological or evolutionary. The former includes phenotypic
plasticity and dispersal, while the latter entails genetic change (Reusch & Wood 2007).
Evolutionary theory suggests that with increasing unpredictability of the environment, as
expected under global change, selection should favour genotypes that possess a broader tolerance
towards stress and other environmental challenges. Ontogenetic and genetic differences in the
susceptibility to acidification and warming may affect population dynamics and intra-specific
competition. Within populations, susceptibility to stress changes in the course of ontogeny (Fabry
et al. 2008) and often highest mortality occurs at early life stages (Gosselin & Qian 1997).
The direct effects of abiotic stress such as predicted during global change will usually not be fatal
at the individual or species level, but when they modify interactions or associations (such as
epibiosis) the impact of these stress factors may be substantially larger (Wahl 2008). When cooccurring and competing species are differentially impacted by this abiotic stress we may expect
shifts in the outcome of competitive interactions and, ultimately, in the structure and functioning
of the community (Jaschinski & Sommer 2008). Indeed, the modulation of interactions by shifts
in environmental conditions can be considered as ecological levers which may alter the purely
physiological impact of the stressor(s) in 2 directions: (i) at the level of the individual the direct
physiological effects may be masked, dampened or enhanced by changes in biological
interactions such as predation or competition and (ii) the impacts at the species level – even when
non-lethal – by interaction modulation will be projected onto the community level and may lead
to “surprising” (Christensen et al. 2006) re-organisations of the structural and functional
properties of a community. While population or community level effects have received little
attention, one particularly understudied component of benthic systems in this regard are
microbial communities (Inagaki et al. 2006).
We will compare the interaction-mediated responses of benthic populations and communities in a
variety of complementary systems: seagrass and macroalgae beds, macrobenthic and seafloor
microbial communities. Where possible, regional and systemic comparisons will be carried out
(e.g. tropical – temperate, deep sea – shore habitats).
224
Most of past research on hypercapnia effects has focussed either on tropical reefs or on
planktonic organisms (Fabry et al. 2008). However, some examples for interaction mediated
effects of environmental stress in benthic systems have been reported. Calcifying organisms, such
as coralline algae, may suffer a reduction in competitiveness in the course of environmental
acidification (Kuffner et al. 2007). Consequently, their non-calcifying local competitors may
indirectly benefit from OA even if they are only slightly less negatively impacted by this
environmental stress than the corallines. Competition between algae with and without carbon
concentration mechanisms is expected to shift when pCO2 rises (Zimmerman et al. 1997). A
higher C:N:P ratio as a possible consequence of CO2 enrichment may decrease food quality and
consumer fitness (e.g. Cruz-Rivera & Hay 2000). Shifts in predation pressure are expected. Also,
two (or more) stressors may interact and modulate their respective effects on a given response.
Thus, growth and shell properties play important roles for the competitiveness and the
susceptibility to predation of an important bioengineer such as mussels (e.g. Gutierrez et al.
2003). The interaction of acidification, warming and other stressors (e.g. eutrophication,
desalination) affect these traits in various and often unexpected ways with far-reaching
consequences for this species’ persistence (Kossak 2006, Wahl et al. work in progress).
Hence, it is essential to expand the OA impact studies beyond the species level to community
structure, comprising both “horizontal” interactions (competition among basal species, epibioses)
as well as the “vertical” aspects, i.e. food quality and trophic interactions. The interaction with
the second project in this theme will be very strong.
Principal foci will be shifts in nutritional quality/prey palatability and predation-mediated
environmental stress (4.1.1, 4.1.2), competitional shifts through species specific differences in
stress susceptibility (4.1.1, 4.1.3.), restructuring of populations by differences in stress sensitivity
between ontogenetic stages and genotypes (4.1.2, 4.1.3.), interaction-modulation through altered
epibioses (4.1.1, 4.1.2, 4.1.3.), restructuring of microbial and macrobial communities through
interaction-mediated stress effects (4.1.2, 4.1.4)
4.1.1
Ragnhild Asmus has long-standing experience in benthic ecology of soft bottom areas especially
temperate and tropical seagrass beds (Asmus & Asmus 2000). She also is experienced in field
experiments and community ecology with a focus on the ecology of benthic primary producers in
different soft bottom communities. She has large expertise in network analysis of food webs and
stable isotope techniques (Baird et al. 2004, 2007). Christian Wiencke has long standing
expertise in seaweed ecology and physiology and in research on anthropogenic effects (e.g.
increased UV radiation) on seaweed performance (Wiencke et al. 2006, Roleda et al. 2007,
Steinhoff et al. 2008). Lars Gutow studies plant-grazer interactions at both the ecological and the
physiological level with emphasis on the implications of nutritional quality and quantity on
meso-herbivore fitness (Gutow et al. 2006, Gutow et al. 2007).
4.1.2
Martin Wahl has extensive experience in benthic community ecology with a strong emphasis on
complex biotic interactions (defences, epibiosis, consumption, competition), their modulation by
abiotic factors, and the impact of environmental stress. His experimental approach comprises in
situ SCUBA experiments, climate simulation mesocosm experiments and analytical lab
techniques (Wahl et al. 2004, Sugden et al. 2008, Wahl 2008). Thorsten Reusch has broad
225
experience in ecological genetics of aquatic and marine organisms, including the development
and analysis of genetic marker data, the set-up of selection experiments and their analysis.
4.1.3
Kai Bischof is studying the eco- and stressphysiology of benthic macroalgae from polar,
temperate and tropical ecosystems (Bischof et al. 2006a). Interactive stress effects on macroalgal
photosynthesis, growth, reproduction and competition, as well as physiological acclimation
mechanisms (e.g. antioxidative strategies (Bischof et al. 2006b)) are the prime focus of his
research. By his workgroup tropical reef algae are successfully cultivated at the ZMT aquaculture
facility.
4.1.4
Alban Ramette has a broad expertise in high-throughput molecular characterization of microbial
communities in environmental samples and in the further statistical evaluation of multivariate
data (Ramette & Tiedje 2007). Antje Boetius has extensive experience with biogeochemical and
microbiological investigations of marine sediments, both in the field including in situ
measurements and in experimental approaches. In addition, previous molecular work has been
successfully accomplished at the proposed natural CO2 reservoir site (Inagaki et al. 2006).
Subproject 4.1.1 Effects of ocean acidification on trophic interactions in coastal seaweed
and seagrass ecosystems
R. Asmus, C. Wiencke, L. Gutow, H. Asmus, D. Hanelt, R. Saborowski, I. Bartsch
This subproject provides many opportunities for close cooperation with other BIOACIDprojects. The reference numbers of cooperating subprojects are given in brackets.
The combined effects of ocean acidification and increased seawater temperatures on marine
macrophytes from the North Atlantic will be investigated in multi-factorial experiments under
controlled laboratory conditions [3.2.4, 4.1.3]. In the facilities of the AWI at Sylt and
Bremerhaven selected seaweed and seagrass species will be raised under constant pCO2-levels
representing pre-industrial (280 ppm), current (380 ppm), and predicted future (700 ppm) pCO2conditions and constant temperatures of 10 and 20°C. An additional experimental temperature
of 25°C will be applied to the seagrass species. The selected species will include the North Sea
seagrass species Zostera marina and Z. noltii and representatives of the three major taxonomic
seaweed groups (i.e. Chlorophyta, Rhodophyta, and Phaeophyceae).
For each species photosynthetic activity, growth, and elemental composition (C, N, P) will be
measured for each pH/temperature combination [4.1.2, 4.1.3, 4.2.2]. Photosynthetic activity will
be measured by pulse amplitude modulated (PAM) fluorometry and O2-electrodes. Since both
methods do not measure carbon fixation directly, net photosynthetic rates will be inferred from
exemplary accompanying 14C-isotope measurements. Brown algae produce polyphenolic
compounds such as phlorotannins via the secondary metabolic pathway. Besides their structural
and UV-protective function phlorotannins act as feeding deterrents for many herbivores. Due to
this implication on algal palatability the phlorotannin content will be quantified in brown algae
by the Folin-Ciocalteu method.
226
In no-choice feeding experiments seaweeds grown under different pH/temperature treatments
will be offered to selected meso-herbivores (peracarids and gastropods) collected in natural
seaweed beds of Sylt and Helgoland. These experiments will focus on the treatment effects on
macrophyte nutritional quality. In order to avoid pH/temperature effects on the grazer
physiology the experiments will be conducted in untreated (but filtered) North Sea water at a
constant temperature of 15°C. Each individual grazer will be fed with one seaweed species from
one treatment only. In short-term experiments we will study the effects of altered food
chemistry on consumption rates, assimilation efficiency, and respiration of the grazers [4.2.1].
The effects of altered food quality on grazer fitness will be studied in long-term experiments.
Fitness will be determined in terms of growth, reproduction, and survival of the animals. In
population models, these life history parameters will be used to calculate fitness parameters
such as population growth rates.
We will study how the combined effects of temperature and acidification affect the activity of
highly important enzymes like cellulase, laminarinase, endo- and exopeptidase, phosphatase,
and lipase [1.2.1]. Digestion and assimilation of the diets by the grazers will be determined by
the activities and the characteristics of selected digestive enzymes in the digestive tract of the
animals, and by the chemical composition of the faecal pellets in comparison to the food (C, N,
P). Comparison between pH effects on the activities of endogenous digestive enzymes and of
those produced by heterotrophic bacteria will be performed in collaboration with project [1.2.1].
The consequences for the decomposition of faecal pellets and nutrient recycling will be studied
by the activity of digestive enzymes within faecal pellets and by the release of organic and
inorganic degradation products (carbohydrates, proteins, phosphate) [1.2.1].
In natural seagrass beds, the removal of seagrass epiphytes by grazers has strong effects on
seagrass performance. Seagrasses that were grown together with their epiphytes under different
pH/temperature regimes will be offered to selected meso-herbivores in feeding experiments in
order to investigate how ocean acidification in conjunction with increasing temperatures will
affect the balance between epiphyte growth and removal by grazers and the possible
implications on seagrass performance (measured as biomass increment, e.g. leaf area index)
[1.1.5, 4.1.2, 4.1.4]. These experiments will be conducted in untreated (but filtered) North Sea
water at a constant temperature of 15°C.
Ecosystem consequences of ocean acidification will be addressed in CO2-incubated benthic
chambers (5 to 500 dm3 volume) in Wadden Sea seagrass beds at Sylt [4.1.2, 5.1]. In situ and
benthic mesocosm enclosures (ca. 3 m3) of partial systems of a seagrass bed will be exposed to
different CO2 concentrations (from weeks to ½ year). System effects will be measured as
changes in biomass, abundance, oxygen and nutrients (N, P, Si) to compute growth, respiration,
productivity, and egestion of the community and particular community compartments [1.1.5,
4.1.4]. The results will be subject to a comprehensive network analysis (ENA) of altered trophic
and metabolic pathways in seaweed and seagrass systems in order to identify the sensitivity of
trophic pathways within these systems to acidification of coastal waters.
227
Schedule
First Year
4.1.1
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Incubation of macrophytes at different
pH/temperature combinations
Laboratory experiments on macrophytes
Laboratory experiments on herbivores
Laboratory experiments on fecal pellet decomposition
Field experiments on community effects in benthic
chambers
Analysis of experimental data
Network analysis of data from benthic chambers
Presentation of data and results, manuscript
preparation
Milestones (4.1.1)
- Incubated seaweed and seagrass species will be available for laboratory
experiments
- First experimental data on pH/temperature effects on macrophytes
- First results from benthic chamber experiments
- Experimental data on consumption and assimilation efficiency of grazers
- Experimental data on pH/temperature effects on fecal pellet decomposition
- Data set on pH/temperature effects on macrophytes completed
- Experimental data on the effects of altered macrophyte stoichiometry
on grazer fitness
- Final results from benthic chamber experiments
- Evaluation of data sets
month 6
month 15
month 21
month 24
month 24
month 30
month 33
month 33
month 35
Subproject 4.1.2 Acidification stress: Early life stage ecology in times of global change.
M. Wahl & T. Reusch
We will ask to which extent under different pCO2/temperature settings survival and performance
of early life stages of some key species are affected (year 1), how sensitivity varies among
genotypes (year 2), and how intra- and interspecific competitiveness is determined by differences
in sensitivity among genotypes and species, resp. (year 3). We expect early life stages to react
more sensitively to stress as compared to adults [in cooperation with projects 2.1.1, 2.1.3, 2.3.1].
Many of these aspects will be run in close cooperation with other projects (reference number in
brackets). In benthic hard bottom communities of the Western Baltic, barnacles (Balanus
improvisus, Bi) and mussels (Mytilus edulis, Me) are the ecologically most important suspension
feeders. The bladder wrack Fucus vesiculosus (Fv) is an important primary producer and
ecosystem engineer. The small calcareous polychaete Spirorbis spirorbis (Ss) is a suspension
feeder living preferentially as epibiont on macroalgae. All 4 species control a substantial amount
of the flow of matter between the open water and the benthos. They compete directly for
resources (food and/or attachment substratum) and indirectly by attracting or distracting shared
consumers.
All experiments will be run in micro- and mesocosms allowing the control of all relevant
variables. Survival and performance (year 1) will be studied on freshly settled juveniles obtained
228
from in vitro reproduction (Bi, Fv, Ss) or field sampling (Me in June – August). Acidification
will be achieved by bubbling with CO2 enriched air at defined and automated concentrations.
Acidification treatment levels as agreed project-wide (i.e. 380 – 560 – 980ppm) will be combined
with 2 temperature levels (ambient – predicted ∆5°C). Species performance (photosynthesis as
assessed by in situ PAM, respiration rates, growth, calcification) [2.1.3, 3.1.3, 3.1.4, 3.5.1, 4.1.3]
will be assessed in the absence of biotic stressors (fouling, grazing, competition). Although we
estimate the immediate stress impact to be small relative to its long-term and indirect effects we
will strive to develop a reliable quantification of the stress level experienced (potential stress
indicators: heat shock proteins, ion-regulation proteins, hypoxia-induced factor) [3.2.3].
Increased variance with stress strength will serve as an indication for inter-genotype variation in
sensitivity and allow selecting suitable species for the 2nd question. In the second year we will run
simplified pilot studies on the role of genetic diversity for stress resistance [1.1.4, 2.2.1] which
will be substantially deepened in a possible second 3-year period of BIOACID (i. e. using
genetic marker loci, either gene-linked or anonymous microsatellites, both of which are
currently being developed in Me and Fv). In a first step we will assess differences in stress
susceptibility between genotypes (equivalent to individuals in these sexually reproducing species)
using the performance parameters defined above. For this experiment, only species will be used
for which reproduction, embryogenesis and recruitment can be handled in the lab, i.e. Fv, Bi a/o
Ss. In a second and optional step [1.1.4], we assess potential evolutionary mediation of survival
and plasticity evolution using controlled crossings that serve as base to assemble genetically
diverse versus depauperate larval pools. This can be realized by either testing sibships alone or in
various combinations (Gamfeldt et al. 2005). Ideally, sibships from mother-father pairs which
differ in stress-sensitivity will be used. By obtaining half-sibs of identical female individuals
fertilized by different fathers, a heritability analysis of tolerance to acidification will provide
data on the predicted evolutionary response of the targeted key species to
acidification/warming. High heritability estimates in conjunction with strong selection
differential would indicate that persistence of benthic key species is possible due to
microevolution (Falconer & Mackay 1996). At the community level, we will finally (year 3)
investigate whether competitive hierarchies [3.2.2, 4.1.1] among selected sessile species (Bi,
Me, Fs) change under various acidification/warming scenarios. Competitiveness of a species
may be modulated by different levels of epibiosis resulting from weakened antifouling
defences or altered fouling pressure [4.1.1, 4.1.4, 4.2.1, 4.2.2]. To this purpose we will
assemble 2- or 3-species communities of Bi, Me a/o Fv at comparable initial density and
genetic diversity, and follow their structural changes over time under different pCO2/T
settings. [In a second 3-yr phase of BIOACID we will aim to disentangle the effects if species
identity and genetic diversity on competitiveness.].
Schedule
4.1.2
First Year
I
II
Second Year
III
IV
I
II
III
Third Year
IV
I
II
III
IV
Survival and performance
Genetic variability
Competition
Analysis & Publication
229
Milestones (4.1.2)
- Experimental data on CO2/pH sensitivity of juveniles
- Experimental data genetic variability of CO2/pH sensitivity
- Data set on changing competitiveness
- Evaluation of combined data sets, sensitivities & uncertainties
month 3
month 10
month 21
month 30
month 36
Subproject 4.1.3: Competitive success of calcifying and non-calcifying macroalgae under
shifting pH regimes in tropical vs. temperate regions
K. Bischof, A. Kunzmann, M. Nugues, T. Rixen, M. Teichberg
We will examine the physiological response of calcifying and non-calcifying macroalgae to
multiple abiotic stress and predict their respective competitive success on tropical reefs and
temperate rocky shore habitats in a scenario of ocean acidification.
The main questions we will address are:
1) To what extent is photosynthesis and growth of calcifying macroalgae affected by shifts in pH,
temperature and (PAR & UV) radiation in comparison to non-calcifying macroalgae, and
how are these responses modified under uni- vs. multifactorial stress? To what extent is
calcification affected in tropical vs. temperate algal species in the respective treatments?
2) What are the underlying physiological acclimation mechanisms involved in stress responses?
3) How will the competitive strength of calcifying and non-calcifying macroalgae be altered by
future ocean acidification?
4) How will local stress factors such as eutrophication interfere with the competitive success of
calcifying, and non-calcifying, particularly ephemeral opportunistic macroalgae under ocean
acidification?
We will examine the physiological response of commonly found calcifying and non-calcifying
macroalgae from the tropics and Helgoland (e.g. Padina, Halimeda, Corallina, Lobophora,
Hypoglossum, Entermorpha, Acrosiphonia), applying uni- and multifactorial stress experiments
in the laboratory and mesocosms at ZMT, University of Bremen, AWI Sylt and AWI Helgoland.
(coordinated with 4.1.1-4.1.4). In the second year of the project, field and mesocosm studies will
be conducted at a tropical field site (e.g. Similan Islands, Thailand). In the experiments we will
expose the algae to different CO2 regimes including the pre-industrial level (280 ppm), present
day conditions (380 ppm) and 560, 700 and 1000 ppm. In short-term experiments temperature,
UV-radiation and nutrient levels will be changed according to the degree of expected variation of
the respective factor within the respective habitat. In mesocosm ponds run at different pH and
temperature conditions and field set-ups we will offer sterile settlement tiles as well as coralline
algae as substrate and monitor algal recruitment, primary succession and epibiosis under varying
abiotic conditions and examine the interspecific competition of calcifying and non-calcifying
macroalgae. In all experiments we will measure photosynthesis (by oxygen evolution and
chlorophyll fluorescence), growth and uptake of dissolved inorganic C and N on an
230
individual/species specific and community base. In situ measurements of calcification will be
conducted in cooperation with Project 3.4.2. using microsensors. We will measure oxidised and
total glutathione content (GSH/GSSG) as an indicator of oxidative stress, and thus a measure of
general stress exposure, easily to compare between species. Studies on coral/alga interactions will
be conducted in cooperation with the projects 3.2.2 and 3.2.3, and by Dr. Maggy Nugues through
institutional funding by ZMT. We will closely cooperate with projects on trophic interactions
within seagrass and seaweed dominated communities under shifting pH scenarios [4.1.1], and
with 4.1.4. on bacterial communities on these organisms. An exchange of samples to study
changes in ultrastructure of the CaCO3 matrix of the different calcifying macroalgal species under
examination is envisaged with project 3.2.4.
Schedule
4.1.3
First Year
I
II
Second Year
III
IV
I
II
III
Third Year
IV
I
II
III
IV
Development of exp. infrastructure
Cultivation of macroalgae under
varying pH/temperature
Stress experiments in the laboratory
Cruise preparation
Field
experiments
(1st
year
nd
Helgoland/Sylt, 2 year Tropics)
Sample processing
data interpretation
Manuscript preparation, conferences
Milestones (4.1.3)
- CO2 incubation chamber at ZMT will be available to the project
- First laboratory data on pH/temperature effects on temperate and
tropical macroalgae
- Results from field & mesocosm experiments in temperate regions
(physiology vs. competition)
- Results from field & mesocosm experiments in tropical regions
(physiology vs. competition)
- Last laboratory studies & experimental work completed
- Data implementation:
1.) patterns in physiology and competition: temperate vs. tropical habitats
2.) common patterns within the other projects of cluster 4
- Evaluation of data sets, publication, modeling approaches
month 3
month 6
month 12
month 24
month 30
month 32
month 34
month 35
231
Subproject 4.1.4: Effects of ocean acidification on microbial community structure,
composition and activity in natural and experimental systems
A. Ramette & A. Boetius
Longterm CO2 effects on microbial diversity, biomass and function in the seabed
Transects of seabed through which CO2-rich porewaters seep from a natural CO2 reservoir
(sediment-hosted CO2 lake of the southern Okinawa Trough hydrothermal system (Inagaki et al.
2006)) will be used as natural laboratory for the study of long-term consequences of varied CO2
and pH levels on microbial community structure. Long term adaptation and consequent
structuring of natural benthic communities cannot be studied experimentally. A series of
molecular techniques will be applied to determine shifts in community structure (T-RFLP,
ARISA), community composition (sequencing of 16S rRNA genes), abundance (fluorescent in
situ hybridization, quantitative PCR), biomass and activity (sulfate reduction, respiration,
heterotrophy, CO2 fixation, methane production and consumption). In addition, biomass and
diversity of meio-, macrofauna and megafauna will be conjointly assessed to address thresholds
of high CO2 and low pH effects on benthic ecosystems. Adequate samples were obtained recently
from an expedition to Okinawa Trough (SO196, March 2008). This work will also contribute to
Theme 3 projects on buffering effects of different sediments in calcification/decalcification
processes and to Theme 1 with the possible detection of epsilonproteobacteria and
chemolithotrophic bacteria in the naturally enriched habitats, which are the target of the
experiments in Theme 1.1.1.
CO2 and pH thresholds in experimental model systems
Short-term shifts in microbial community diversity, composition and abundance will be
experimentally investigated as a function of pH, temperature and CO2 variation using flowthrough reactors as sedimentary mesocosms. These experiments will also assess changes in
ecosystem services mediated by microbes such as remineralization, sulfate reduction, methane
production or consumption. Bioreactors will be filled with standard heterotrophic sediments from
coastal areas (e.g. long-term research station, Sylt, Germany; Wadden Sea sediments of different
grain sizes, also see 4.1.1), for which gradients of pH, temperature and CO2 in the porewater will
be established. Using this approach, different factor combinations will be tested on microbial
communities, with for instance: a) different levels of CO2 concentration applied to the same
sediment samples, or conversely, b) the application of one CO2 concentration to different
sediment types (silicate vs. carbonate), c) Varying levels of heterotrophic activity under constant
CO2 concentrations, and d) combinations of increasing temperature and decreasing pH. In the
different experimental setups, abundance, composition and activity of the communities in each
bioreactor will be determined at different time intervals to assess resistance (extent of the shift
measured shortly after the end of the experiments), and resilience (observation of shifts after
long-term incubations). Those experiments will be done in collaboration with 4.1.1, 4.2.1, 4.2.2.,
3.1.2 and 3.4.2 allowing the comparison of different systems.
232
Effects of acidification on microbial communities associated with multicellular organisms
A subset of experimental analyses will deal with microbial communities associated with coldwater coral surfaces. Using Lophelia pertusa maintained in aquaria we will test how the
alteration of environmental parameters (pH, CO2 concentration, temperature) in the water column
may directly affect microbial communities that colonize coral branches. Control treatments
consisting of dead coral skeletons will be used to assess whether changes are substrate (i.e.
coral)-specific or not. Those experiments will be done in collaboration with Armin Form (Theme
3). In close collaboration with projects 4.1.1, 4.1.2 and 4.1.3, shifts in microbial communities
colonizing the surface of seagrass and macroalgae will be determined in parallel to the respective
shifting pH and temperature experiments. This integrated strategy will enable a direct observation
of the coupling between microbial and macrobial shifts in complex ecosystems, and the intensity
of the respective shifts.
Schedule
4.1.4
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Longterm CO2 effects on microbial diversity, biomass and function in
the seabed
DNA preparation, fingerprinting,
cell counts
Function measurements
Data analysis, manuscript
preparation
CO2 and pH thresholds in experimental model systems
Column setup, CO2 effects,
community analyses
Sediment effect
preparation
Effects of acidification on microbial communities associated with
multicellular organisms
DNA preparation, community
analyses
preparation
233
Milestones (4.1.4)
month 10
month 12
month 16
month 19
month 28
month 33
- Data on diversity and functional shifts in natural CO2 reservoirs
- Set up of flow-through columns
- Data on CO2 effects on communities
- Data on response of different sediment types
- Data on response of coral-associated microbes
First Year
Personnel costs
4.1.1
4.1.2
4.1.3
4.1.4
Subtotal
Consumables
4.1.1
4.1.2
4.1.3
4.1.4
Subtotal
Travel
4.1.1
4.1.2
4.1.3
4.1.4
Subtotal
Investments
4.1.1
4.1.2
4.1.3
4.1.4
Subtotal
Other costs
4.1.1
4.1.2
4.1.3
4.1.4
Subtotal
Total
234
Second Year
Third Year
Total
4.1.1
Personnel costs:
Experimental work on various aspects of this sub-project (primary
producers, enzymatic nutrient recycling, community processes) will be
conducted contemporaneously by master and diploma students at the
different facilities of the AWI at Bremerhaven, Helgoland and Sylt and at the
University of Hamburg. Funding is requested for 1 PostDoc (
)
for 36 months who will coordinate and supervise the students’ work and
assist in thesis preparation. This scientist will be responsible for data
management and synthesis of the results from different project
compartments. Experimental work of the PostDoc will encompass herbivore
feeding experiments and field investigations.
4.1.2
Personnel costs:
1 PhD student for 36 months.
Consumables:
Equipment for experiments, reagents, fees for external measurements
Travel:
Visit to other projects within BioAcid, 1 symposium in last year
Investment:
upgrading of existing culturing infrastructure for appropriate heating,
cooling, pCO2 and irradiation treatment plus, pCO2 measurement techniques
(e.g. CO2 and pH microsensor systems)
4.1.3
Personnel costs:
1 PhD student for 36 months. In the first and second year another
EUR/year are requested for student workers, assisting in culture work, in
field and laboratory experiments and routine analytic procedures
Consumables:
Lab chemicals (HPLC grade) for analysis of photosynthetic pigments and
cellular oxidative state (GSH/GSSG), pre cast gels and primary & secondary
antibodies for SDS-PAGE and Western Blotting for protein analysis
Travel:
for annual project workshop, conferences and field research. In the second
year, field research has to be conducted at a tropical field site, which requires
funding for airplane tickets, freight of scientific equipment, lab fees, local
field & diving support, and accommodation. In the third year, we will seek to
present our results in national and international conferences.
Investment:
Oxygen electrodes, CO2 and pH microsensor systems to measure
photosynthesis & calcification on small spatial scales, 1x Imaging-PAM for
measuring whole plant photosynthesis with an image analysis system,
Cryostats and pumps for aquaculture and mesocosm systems.
4.1.4
Personnel costs:
1 PhD student for 36 months.
Consumables:
Molecular biology reagents (DNA extraction, purification, quantification,
PCR enzymes, etc.), including capillary electrophoresis to separate PCR
amplicons (ARISA, T-RFLP). Consumables for DNA sequencing (done at
the MPI) are also included. Microbial activity measurements (sulfate
reduction, respiration, heterotrophy, CO2 fixation, methane production and
235
consumption) are also included in the estimated costs, as well as
consumables for microscopic counts (Acridine Orange, Fluorescent In Situ
Hybridization).
Travel:
One international conference and one BIOACID meeting per year for the
student
Investment:
Microsensor construction (pH, CO2) will be done at the MPI during the first
year (collaboration with Dirk de Beer; Theme 3). Flow-through columns and
related equipment (pump, tubes, etc.) will be purchased in year 1.
Project 4.2: OA effects on food webs and competitive interactions in pelagic
ecosystems (M. Boersma)
i. Objectives
We will investigate the role of OA shaping competitive interactions and food web dynamics in
pelagic ecosystems, working on microalgae, their grazers, and associated bacterial communities.
More specifically our objectives are:
•
To assess the sensitivity of different microalgal species and strains to OA
•
To provide a process-based understanding for the observed responses to OA by applying
a combination of different techniques and molecular tools
•
To establish the influence of OA on mixed assemblages in competition experiments at
the intra-population level as well as at the inter species level
•
To assess the consequences of OA on population community structure of key species as
well as on the nutrient stoichiometry of primary producers and primary consumers
•
To establish the way grazers deal with excess carbon when feeding on food with
imbalanced C:N:P ratios. We will investigate the reaction of bacterial communities on
different carbon sources originating from these grazers.
Species-specific differences in CO2/pH-sensitivity may directly impact phytoplankton succession
and distribution (Hansen 2002, Rost et al. 2003), as well as competitive interactions between
organisms. Not only direct sensitivities to CO2/pH as a stress factor influences succession and
competitive interactions, but also changes in the availability of essential nutrients may affect the
outcome of competitive interactions for these nutrients (Tilman 1982). Loladze and co-workers
(2004) took this one step further and showed that even the ratio of different nutrients can be
regarded as the limiting nutrient, and thus the outcome of competition may even depend on the
supply ratio of these nutrients. This is particularly relevant as the ratio of the available carbon to
other nutrients is bound to change as a result of OA, especially in coastal areas where a
concurrent trend of decreasing nitrogen and phosphorus inputs can be found (Wiltshire et al.
2008). This results in increasing C:N and C:P ratios of dissolved substances available for
phytoplankton growth.
236
These changing growing conditions are particularly relevant for dinoflagellates. Dinoflagellates
are a diverse and abundant group of protists with complex interactions in the food web. They can
form ‘red-tides’, and some species/strains are known to produce various toxins with large
consequences for ecosystems, human health and economy. Given the increasing number and
intensity of dinoflagellate blooms over the last decades (Hallegraeff 2003), it is remarkable that
relatively little is known about the underlying causes. It has been suggested that CO2/pH affects
photosynthesis, growth, and toxin production of phytoplankton (Lundholm et al. 2004, Hansen et
al. 2007), moreover nutrient (nitrogen) limitation can also affect toxin production of some species
as well (Parkhill & Cembella 1999), and it could be hypothesized that an increase in the C:N ratio
of the available resources should lead to a reduction of overall toxicity. Moreover, these different
environmental conditions could lead to strain or species replacements, with potentially large
secondary effects on competitors and consumers.
Not only do we expect species or strain replacements as a result of changing conditions, even in
genetically identical lineages other aspects affecting the quality as food for higher trophic levels
might change. Increasing CO2 concentrations will affect the nutrient stoichiometry of primary
producers. Provided that the levels of nutrients such as Si, N and P do not change, an increase in
C-uptake by autotrophs should cause a shift in the elemental composition of the primary
producers, and hence influence their quality as food for their consumers. They will have more C
relative to the nutrients, at the same time the total biomass of these producers will probably be
higher as a result of the increased C-availability. In most known cases an increase in the
C:nutrient ratio of primary producers causes a reduction in their quality as food for the next
trophic level (Boersma 2000, Malzahn et al. 2007a). Hence, the situation might arise that
consumers are faced with more food, which at the same time is of inferior quality. The aim of this
project will be to investigate the effects of these changes in food quality for consumers, whereas
we expect direct pH effects to be of less importance (Kurihara et al. 2004, Mayor et al. 2007).
An excess amount of carbon relative to nutrients in algae should lead to an increased excretion of
carbon by secondary producers. Depending on how this excess C is excreted (DIC/CO2 or DOC)
this could have major consequences for the flow of energy and matter (Darchambeau et al. 2003).
The nature of these excretory products has been under considerable debate (Sterner et al. 1998,
Plath & Boersma 2001, Darchambeau et al. 2003, Jensen & Hessen 2007), but still has not been
answered. DOC is potentially taken up rapidly by bacteria, which might benefit from increased
C:nutrient levels in algae, but virtually nothing is known here (Sterner et al. 1998, Riebesell et al.
2007). Since bacteria are homeostatic and “more like animals than plants” with respect to their
stoichiometry (Makino et al. 2003), changes in the elemental composition of excretory products
should cause shifts in bacterial community composition. To date, the main focus of the studies on
the response of bacterial communities has been direct, i.e. the direct effects of acidification on
diversity, and there seem to be no large effects (Allgaier et al. 2008). We have no idea on the
indirect effects, and the consequence of these population shifts due to “imbalanced nutrients” for
functional diversity and biogeochemical cycles are still unknown
Hence, potentially, the biological production of the world-seas will increase in the first trophic
level as a result of higher CO2 concentrations, but the second trophic level might not be able to
benefit from this increased biomass.
4.2.1 Maarten Boersma has considerable experience with experimental plankton ecology at
many different scales, ranging from laboratory experiments on individual organisms to
large-scale laboratory and field enclosures. Moreover, he is an expert on direct
237
manipulations of components of the food in algal-grazer interactions (Boersma 2000,
Boersma & Elser 2006, Boersma et al. in press). Arne Malzahn is a fisheries biologist
with expertise in experimental and field approaches in larval fish and plankton ecology.
His main research interests are ecological stochiometry and trophic transfer. He was the
first to show that limitations on the primary producer level can be transfered through
primary consumers to secondary consumers (Aberle & Malzahn 2007, Malzahn et al.
2007a, Malzahn et al. 2007b). Gunnar Gerdts is an expert on bacterial community
analysis of different habitats and pro-eukaryotic consortia. He has managed to investigate
bi-trophic systems of microalgae amended with 13C-carbonate and associated bacterial
community feeding on 13C containing algal exudates using stable isotope probing, which is
the first time that this approach has been applied to marine pelagic systems.
4.2.2. Björn Rost has studied the carbonate chemistry effects on marine phytoplankton over the
last decade, focusing on physiological key processes such as photosynthesis, carbon
acquisition and calcification (Rost et al. 2003, Rost & Riebesell 2004). He is an expert on
membrane-inlet mass spectrometry (MIMS), which allows monitoring gas exchange
processes in real-time under different environmental conditions, a technique that is central
for the planned experiments (Rost et al. 2007). Uwe John’s research emphasis is on the
molecular regulation of toxin synthesis and growth regulation of marine protists,
combining ecological and physiological experiments with functional genomics to reveal
the mechanisms behind algal bloom formation (Cembella & John 2006, Tillmann et al.
2007, John et al. 2008). He has established several assays for the estimation of the
molecular response of microalgae to environmental stressors using standard protein assays
and microarrays/quantitative PCR (qPCR), respectively.
Subproject 4.2.1: OA effects on pelagic community structure and food chains
M. Boersma, A. Malzahn, & G. Gerdts
One of the main questions targeted by this project is as to the effects of high CO2 induced
differences in algal quality on herbivores and bacteria. Furthermore, food with a high C:nutrient
should lead to an increase in the excretion of carbon by consumers, but to date there is no
information on these processes. The high CO2 world will be realized by culturing algae under
elevated CO2-concentrations, relevant to the different IPCC scenarios, and agreed in framework
of BIOACID. These algae will be fed to grazers. It is important to note that we will culture the
algae separately, harvest them and then feed them to the grazers at ambient pCO2. This is
important, as only in this way, we can guarantee that we are investigating food effects only, and
not direct effects of CO2/pH on the grazers, which will be dealt with in 2.1.2 (in cooperation with
this sub project)
We will carry out laboratory experiments with different algae-grazer systems, as it might well be
that different grazers have different mechanisms to void excess C, using at least partly identical
algal species/strains as the ones used in subproject 4.2.2. Therefore, in these experiments we will
assess the reaction of the grazers on the food sources of different quality in terms of nutrient
stoichiometry by analysing growth and reproduction of the consumer, coordinating techniques
with 4.1.1 and 4.1.2. At the same time we will measure respiration rates and DOC production to
better understand the relevance for the ecosystem. The two most important mechanisms being
238
respiration (CO2-production), and release as dissolved organic carbon. Whether one pathway or
the other is chosen has large consequences for the system as DOC is available for heterotrophic
bacterial production, CO2 for autotrophic bacteria only.
We will then proceed to investigate the bacterial community as well as the bacterial growth rates
and activities related to these different conditions. We hypothesise that bacterial activity will be
much higher in those cases that excess carbon is voided as DOC, and that bacterial diversity will
be different, for example by favouring those groups able to fix nitrogen. To investigate the effects
of different C:N:P ratios on bacterial populations as well as the influence of excreted DOC from
grazers, a state of the art methodological stable isotope probing approach will be applied
(Manefield et al. 2002, Radajewski et al. 2003, Sapp et al. 2008). Selected microalgae will be
incubated with different 13CO2 levels under defined N:P ratios and fed to grazers. Microalgal
exudates and grazer excretes (filtrates) will be incubated with natural marine bacterial
communities. Bacterial RNA or DNA will be extracted and “heavy” (13C-containing) RNA or
DNA will be separated from “light” (12C-containing) ribo/nucleic acid molecules by isopycnic
ultracentrifugation and gradient fractionation. Collected fractions will be analyzed for 13C/12C
isotope ratios by mass spectrometry. RNA will be transcribed to DNA by RT-PCR (Reverse
Transcriptase PCR) and analyses of the bacterial community will be performed by PCR with
(group) specific primers, followed by DGGE (Denaturing Gradient Gel Electrophoresis), ARISA
(Automated ribosomal intergenic spacer analysis) or DHPLC (Denaturing high performance
liquid chromatography) and 16S-rDNA sequencing of major bands.
The outcome of the experiments are of quantitative and qualitative nature:
a) Identification of those bacterial populations which benefit from enhanced carbon levels
in the coastal marine system or which are negatively affected.
b) Estimation of bacterial community compositions in conjunction with different C:N:P
ratios of bacterial nutrients
All molecular analyses will be performed in close cooperation with subproject 4.1.4, as well as
with the subprojects in theme 1 [1.2.1-1.2.4]. In contrast to the subprojects in Theme 1, we will
primarily focus on the analysis of shifts in bacterial community composition due to changes in
stoichiometry of algal exudates and excretes of grazers. In Theme 1, the stability of algal
exudates to hydrolysis in dependence of the water pH will be examined, which is a functional
response. We will examine if certain C:N:P ratios of "bacterial food" caused by elevated pCO2 in
the ecosystem will result in specific community compositions, which can be defined as a
structural response. Molecular techniques will be standardized and fingerprint-data will be shared
using advanced database software.
In a next step, we will use laboratory microcosms to investigate system effects. An increase in
bacterial activity should lead to increased micrograzers which are a good food source for
zooplankters, thus fuelling the microbial loop. The great advantage of a stronger microbial loop
for e.g. copepods will be that much of the excess carbon has already been respired by the
intermediate trophic levels, which should cause the micro grazers to be a better quality food. For
this we will use set-ups that are in the size of 10-50 l, stocked with different algae-grazer
combinations and inocula of natural seawater to enter a microbial community component to the
system.
239
Schedule
4.2.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Set-up of culturing facility,
instrument calibration
Collection and rearing of organisms
Algal-Grazer-Bacteria experiments
Cell counts, respirometry &
chemical analyses
DNA/RNA extraction and
preparation (SIP)
Molecular fingerprints &
sequencing
Microcosm experiments
data interpretation
Manuscript preparation, presentation
of results at conferences
Milestones (4.2.1)
- Experimental data set on CO2/pH sensitivity of grazer-algal interactions
- Data set on bacterial diversity and functional shifts
- Data set on microcosm experiments
month 6
month 15
month 24
month 30
month 33
Subproject 4.2.2: Competitive interactions in planktonic microalgae under OA-stress
B. Rost & U. John
We aim to examine the responses of different dinoflagellate species and strains to multiple
abiotic and biotic stressors. Applying a combination of different techniques and molecular tools
will provide a process-based understanding for the observed responses. In addition to
experiments with mono-clonal incubations, competition experiments will be conducted under
respective conditions to investigate the influence of stressors on the intra-specific level (i.e.
population) and species interaction. The project will strongly improve our predictive capabilities
on how different dinoflagellate species as well as mixed assemblages will respond to ocean
acidification. Findings about competitiveness and species/strain interaction of dinoflagellates
under the ocean acidification scenarios will be used for general comparison within theme 4
[4.1.1, 4.1.2, 4.1.3, 4.1.4, 4.2.1].
Representative species of marine dinoflagellates and corresponding strains will be grown under
different pCO2 levels, representing last glacial maximum (180 ppm), present-day (380 ppm) and
those predicted for the future (750-1000 ppm). Since other environmental factors, such as light
240
and nutrient availability, are also changing and possibly modifying the CO2/pH-related responses
interactive effects are investigated in a matrix approach. In these acclimations various parameters
like growth rates, photosynthesis, elemental ratios (C:N:P), photosynthetic pigments or toxin
production/toxicity (Tillmann & John 2002) are determined. These experiments will be
performed with mono-clonal cultures of species/strains. Data and samples will be provided to
1.2.2 (for comparative analysis of C:N:P stoichiometry), to 1.2.1 (for analysis of extracellular
organic material), to 4.2.1 (for feeding experiments) and to 3.5.3 (for morphological examination
of calcifying dinoflagellates). Subsequently, competition experiments will be conducted under
different pCO2 scenarios using strains and later species assemblages that responded differently to
the respective treatments. Because strains and some species cannot be differentiated
morphologically in such mixed assemblages, molecular markers such as AFLP and
microsatellites (John et al. 2004, Alpermann et al. 2006) will be used to qualitatively and
quantitatively distinguish species/strains and hence assess the success of certain
pheno/genotypes.
To get a process-based understanding of the responses to changes in CO2/pH different bio-assays
(in vivo) have been developed at the AWI. The membrane-inlet mass spectrometry (MIMS)
allows monitoring gas exchange processes in real-time under different environmental conditions
(experience exchange and data discussion with 3.1.1). One application allows distinguishing
between CO2 and HCO3- as carbon sources and determines the fluxes as function of
concentration. In other applications, the use of stable isotopes enables to measure activities of
carbonic anhydrase (CA) or photosynthetic electron fluxes (ETR). The advantage of our MIMS
approach is that several processes can be observed and quantified simultaneously, even with
sensitive species such as dinoflagellates (e.g. Rost et al. 2006). These methods unravel the carbon
and energy fluxes, a prerequisite to understand the CO2/pH-dependence in photosynthesis and
other down-stream processes. To further advance our process-understanding the expression of
proteins and genes of key enzymes (e.g. RubisCO, PKS) will be quantified using standard protein
assays and microarrays/quantitative PCR (qPCR), respectively. Those methods including Oligonucleotide DNA microarrays will provide, in combination with detailed physiology, a
comprehensive mechanistic overview of the cascade from gene expression to physiological
responses.
Schedule
4.2.2
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Experimental set up
Phenotypic characterisation/ MIMS
Genotypic characterisation
Interspecific competition
Intraspecific competition
Data evaluation and publication
241
Milestones (4.2.2)
- Data on species/strain-specific sensitivity towards stressors
- Competition experiments concluded
- Evaluation of influence of species/strain success due to stress tolerance
month 6
month 24
month 30
month 36
First Year
Second Year
Third Year
Total
Personnel costs
4.2.1
4.2.2
Subtotal
Consumables
4.2.1
4.2.2
Subtotal
Travel
4.2.1
4.2.2
Subtotal
Investments
4.2.1
4.2.2
Subtotal
Other costs
4.2.1
4.2.2
Subtotal
Total
4.2.1
Personnel costs:
2 PhD students, one candidate will perform the grazing experiments and
investigate the effects of different conditions on growth, reproduction and
excretory processes of grazers, the other one will investigate the responses of
the bacterial community on the differing conditions as a result of these
different grazing conditions.
Consumables:
Molecular biology reagents (DNA extraction, purification, quantification,
PCR enzymes, etc.), including electrophoresis reagents and consumables to
separate PCR amplifons (DGGE, ARISA, DHPLC). Consumables for DNA
sequencing are included. Costs for isotope ratio measurements and
consumables for determination of C,N & P and DOC.
242
Travel:
Covers the attendance of the annual project workshops and one conference
per year for the Ph.D. students.
4.2.2
Personnel costs:
1 PhD student for 36 months. The candidate will perform experiments on
physiological performance and competitive success of different
dinoflagellate species and strains as outlined in the proposal.
Consumables:
Lab chemicals for culturing and in vivo assays (e.g. MIMS). Lab chemicals
(HPLC grade) for analysis of secondary metabolites, pre cast gels and
primary & secondary antibodies for SDS-PAGE and Western Blotting for
protein analysis. Kits for RNA and DNA extraction, microarrays and the
hybridisation chemistry. Consumables for PCR and qPCR.
Travel:
Covers the attendance of the annual project workshops and one conference
per year for the Ph.D. student.
vi. References
Aberle N, Malzahn AM (2007) Interspecific and nutrient-dependent variations in stable isotope fractionation: experimental studies simulating
pelagic multitrophic systems. Oecologia 154:291-303
Allgaier M, Riebesell U, Vogt M, Thyrhaug R, Grossart HP (2008) Coupling of heterotrophic bacteria to phytoplankton bloom development at
different pCO2 levels: a mesocosm study. Biogeosciences Discuss 5:317-359
Alpermann TJ, John UE, Medlin LK, Edwards KJ, Hayes PK, Evans KM (2006) Six new microsatellite markers for the toxic marine
dinoflagellate Alexandrium tamarense. Mol Ecol Notes 6:1057-1059
Asmus H, Asmus R (eds) (2000) ECSA-Workshop on Intertidal Seagrass Beds and Algal Mats: Organisms and Fluxes at the Ecosystem Level,
Vol 54 (2/3)
Baird D, Asmus H, Asmus R (2004) Energy flow of a boreal intertidal ecosystem, the Sylt-Romo Bight. Mar Ecol Prog Ser 279:45-61
Baird D, Asmus H, Asmus R (2007) Trophic dynamics of eight intertidal communities of the Sylt-Romo Bight ecosystem, northern Wadden
Sea. Mar Ecol Prog Ser 351:25-41
Beaugrand G, Brander KM, Lindley JA, Souissi S, Reid PC (2003) Plankton effect on cod recruitment in the North Sea. Nature 426:661-664
Bischof K, Gomez I, Molis M, Hanelt D, Karsten U, Lüder U, Roleda MY, Zacher K, Wiencke C (2006a) Ultraviolet radiation shapes seaweed
communities. Rev Environ Sci Biotechnol 5:141-166
Bischof K, Rautenberger R, Brey L, Perez-Llorens JL (2006b) Physiological acclimation to gradients of solar irradiance within mats of the
filamentous green macroalga Chaetomorpha linum from southern Spain. Mar Ecol Prog Ser 306:165-175
Boersma M (2000) The nutritional quality of P-limited algae for Daphnia. Limnol Oceanogr 45:1157-1161
Boersma M, Aberle N, Hantzsche FM, Schoo K, Wiltshire KH, Malzahn AM (in press) Nutritional limitation travels up the food chain.
International Review of Hydrobiology
Boersma M, Elser JJ (2006) Too much of a good thing: on stoichiometrically balanced diets and maximal growth. Ecology 87:1335-1340
Burkhardt S, Zondervan I, Riebesell U (1999) Effect of CO2 concentration on C : N : P ratio in marine phytoplankton: A species comparison.
Cembella AD, John U (2006) Molecular physiology of toxin production and growth regulation in harmful algae. In: Granéli E, Turner JT (eds)
Ecology of Harmful Algae. Springer Verlag, p 215-228
Christensen MR, Graham MD, Vinebrooke RD, Findlay DL, Paterson MJ, Turner MA (2006) Multiple anthropogenic stressors cause
ecological surprises in boreal lakes. Glob Chan Biol 12:2316-2322
Cruz-Rivera E, Hay ME (2000) Can quantity replace quality? Food choice, compensatory feeding, and fitness of marine mesograzers. Ecology
81:201-219
Darchambeau F, Faerovig PJ, Hessen DO (2003) How Daphnia copes with excess carbon in its food. Oecologia 136:336-346
Fabry V, Seibel BA, Feely RA, Orr JC (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICES J Mar Sci
65:414-432
Falconer DS, Mackay TFC (1996) Introduction to Quantitative Genetics, Vol. Longman, London
Frost PC, Ebert D, Smith VH (2008) Responses of a bacterial pathogen to phosphorus limitation of its aquatic invertebrate host. Ecology
89:313-318
Gamfeldt L, Wallen J, Jonsson PR, Berntsson KM, Havenhand JN (2005) Increasing intraspecific diversity enhances settling success in a
marine invertebrate. Ecology 86:3219-3224
Gosselin LA, Qian PY (1997) Juvenile mortality in benthic marine invertebrates. Mar Ecol Prog Ser 146:265-282
Gülzow N (2008) Transfer of limitation signals from phytoplankton to zooplankton. Diplom Thesis, University of Kiel
243
Gutierrez JL, Jones CG, Strayer DL, Iribarne OO (2003) Mollusks as ecosystem engineers: the role of shell production in aquatic habitats.
Oikos 101:79-90
Gutow L, Leidenberger S, Boos K, Franke HD (2007) Differential life history responses of two Idotea species (Crustacea : Isopoda) to food
limitation. Mar Ecol Prog Ser 344:159-172
Gutow L, Strahl J, Wiencke C, Franke HD, Saborowski R (2006) Behavioural and metabolic adaptations of marine isopods to the rafting life
style. Mar Biol 149:821-828
Hallegraeff G (2003) Harmful algal Blooms: A global overview. In: Hallegraeff G, Anderson DM, Cembella AD (eds) Manual on Harmful
marine microalgae. UNESCO, Paris, p 25-50
Hansen PJ (2002) Effect of high pH on the growth and survival of marine phytoplankton: implications for species succession. Aquat Microb
Ecol 28:279-288
Hansen PJ, Lundholm N, Rost B (2007) Growth limitation in marine red-tide dinoflagellates: effects of pH versus inorganic carbon
availability. Mar Ecol Prog Ser 334:63-71
Hantzsche FM, Boersma M (submitted) Diet induced responses of physiology and growth of the heterotrophic dinoflagellate Oxyrrhis marina.
Hays GC, Richardson AJ, Robinson C (2005) Climate change and marine plankton. Trends Ecol Evol 20:337-344
Inagaki F, Kuypers MMM, Tsunogai U, Ishibashi J, Nakamura K, Treude T, Ohkubo S, Nakaseama M, Gena K, Chiba H, Hirayama H,
Nunoura T, Takai K, Jorgensen BB, Horikoshi K, Boetius A (2006) Microbial community in a sediment-hosted CO2 lake of the southern
Okinawa Trough hydrothermal system. Proc nat Acad Sci USA 103:14164-14169
Jaschinski S, Sommer U (2008) Top-down and bottom-up control in an eelgrass-epiphyte system. Oikos 117:754-762
Jensen TC, Hessen DO (2007) Does excess dietary carbon affect respiration of Daphnia? Oecologia 152:191-200
John U, Beszteri B, Derelle E, Van De Peer Y, Read B, Moreau H, Cembella AD (2008) Novel insights into evolution of protistan polyketide
synthases through phylogenomic analysis. Protist 159:21-30
John U, Groben R, Beszteri N, Medlin L (2004) Utility of amplified fragment length polymorphisms (AFLP) to analyse genetic structures
within the Alexandrium tamarense species complex. Protist 155:169-179
Kagata H, Ohgushi T (2007) Carbon-nitrogen stoichiometry in the tritrophic food chain willow, leaf beetle, and predatory ladybird beetle. Ecol
Res 22:671-677
Klausmeier CA, Litchman E, Levin SA (2004) Phytoplankton growth and stoichiometry under multiple nutrient limitation. Limnol Oceanogr
49:1463-1470
Kossak U (2006) How climate change translates into ecological change: impacts of warming and desalination on prey properties and predatorprey interactions in the Baltic Sea. Ph.D. thesis, University of Kiel, Germany
Kuffner IB, Andersson AJ, Jokiel PL, Rodgers KS, Mackenzie FT (2007) Decreased abundance of crustose coralline algae due to ocean
acidification. Nature Geosci 1:114-117
Kurihara H, Shimode S, Shirayama Y (2004) Effects of raised CO2 concentration on the egg production rate and early development of two
marine copepods (Acartia steueri and Acartia erythraea). Mar Poll Bull 49:721-727
Lippemeier S, Frampton DMF, Blackburn SI, Geier SC, Negri AP (2003) Influence of phosphorus limitation on toxicity and photosynthesis of
Alexandrium minutum (Dinophyceae) monitored by in-line detection of variable chlorophyll fluorescence. J Phycol 39:320-331
Loladze I, Kuang Y, Elser JJ, Fagan WF (2004) Competition and stoichiometry: coexistence of two predators on one prey. Theor Pop Biol
65:1-15
Lundholm N, Hansen PJ, Kotaki Y (2004) Effect of pH on growth and domoic acid production by potentially toxic diatoms of the genera
Pseudonitzschia and Nitzschia. Mar Ecol Prog Ser 273:1-15
Makino W, Cotner JB, Sterner RW, Elser JJ (2003) Are bacteria more like plants or animals? Growth rate and resource dependence of
bacterial C : N : P stoichiometry. Funct Ecol 17:121-130
Malzahn AM, Aberle N, Clemmesen C, Boersma M (2007a) Nutrient limitation of primary producers affects planktivorous fish condition.
Malzahn AM, Clemmesen C, Wiltshire KH, Laakmann S, Boersma M (2007b) Comparative nutritional condition of larval dab Limanda
limanda and lesser sandeel Ammodytes marinus in a highly variable environment. Mar Ecol Prog Ser 334:205-212
Manefield M, Whiteley AS, Griffiths RI, Bailey MJ (2002) RNA stable isotope probing, a novel means of linking microbial community
function to Phylogeny. Appl envir Microbiol 68:5367-5373
Mayor DJ, Matthews C, Cook K, Zuur AF, Hay S (2007) CO2-induced acidification affects hatching success in Calanus finmarchicus. Mar
Ecol Prog Ser 350:91-97
Müller-Navarra DC (1995) Biochemical versus mineral limitation in Daphnia. Limnol Oceanogr 40:1209-1214
Parkhill J, Cembella A (1999) Effects of salinity, light and inorganic nitrogen on growth and toxigenicity of the marine dinoflagellate
Alexandrium tamarense from northeastern Canada. J Plankton Res 21:939-955
Perry AL, Low PJ, Ellis JR, Reynolds JD (2005) Climate change and distribution shifts in marine fishes. Science 308:1912-1915
Plath K, Boersma M (2001) Mineral limitation of zooplankton: stoichiometric constraints and optimal foraging. Ecology 82:1260-1269
Radajewski S, McDonald IR, Murrell JC (2003) Stable-isotope probing of nucleic acids: a window to the function of uncultured
microorganisms. Curr Op Biotech 14:296-302
Ramette A, Tiedje JM (2007) Biogeography: An emerging cornerstone for understanding prokaryotic diversity, ecology, and evolution. Microb
Ecol 53:197-207
Reusch TBH, Wood TE (2007) Molecular ecology of global change. Mol Ecol 16:3973-3992
Riebesell U, Revill AT, Holdsworth DG, Volkman JK (2000) The effects of varying CO2 concentration on lipid composition and carbon
isotope fractionation in Emiliania huxleyi. Geochim Cosmochim Act 64:4179-4192
Riebesell U, Schulz KG, Bellerby RGJ, Botros M, Fritsche P, Meyerhofer M, Neill C, Nondal G, Oschlies A, Wohlers J, Zollner E (2007)
Enhanced biological carbon consumption in a high CO2 ocean. Nature 450:545-U510
Roleda MY, Wiencke C, Hanelt D, Bischof K (2007) Sensitivity of the early life stages of macroalgae from the northern hemisphere to
ultraviolet radiation. Photochem Photobiol 83:851-862
Rost B, Kranz SA, Richter KU, Tortell PD (2007) Isotope disequilibrium and mass spectrometric studies of inorganic carbon acquisition by
phytoplankton. Limnol Oceanogr Methods 5:328-337
Rost B, Richter KU, Riebesell U, Hansen PJ (2006) Inorganic carbon acquisition in red tide dinoflagellates. Plant Cell Env 29:810-822
244
Rost B, Riebesell U (2004) Coccolithophores and the biological pump: responses to environmental changes. In: Thierstein HR, Young JR (eds)
Coccolithophores: from molecular processes to global impact. Springer, Berlin, p 99-125
Rost B, Riebesell U, Burkhardt S, Sultemeyer D (2003) Carbon acquisition of bloom-forming marine phytoplankton. Limnol Oceanogr 48:5567
Saborowski R, Sahling G, del Toro MAN, Walter I, Garcia-Carreno FL (2004) Stability and effects of organic solvents on endopeptidases from
the gastric fluid of the marine crab Cancer pagurus. J Mol Cat B 30:109-118
Sanford E (1999) Regulation of keystone predation by small changes in ocean temperature. Science 283:2095-2097
Sapp M, Gerdts G, Wellinger M, Wichels A (2008) Consuming algal products: Trophic interactions of bacteria and a diatom species
determined by RNA stable isotope probing. Helg Mar Res
Scheffer M, Carpenter S, Foley JA, Folke C, Walker B (2001) Catastrophic shifts in ecosystems. Nature 413:591-596
Scheibling R (1986) Increased macroalgal abundance following mass mortalities of sea urchins (Strongylocentrotus droebachiensis) along the
atlantic coast of Nova Scotia. Oecologia 68:186-198
Shirayama Y, Thornton H (2005) Effect of increased atmospheric CO2 on shallow water marine benthos. J Geophys Res 110
Steinhoff FS, Wiencke C, Muller R, Bischof K (2008) Effects of ultraviolet radiation and temperature on the ultrastructure of zoospores of the
brown macroalga Laminaria hyperborea. Plant Biol 10:388-397
Sterner RW, Clasen J, Lampert W, Weisse T (1998) Carbon: phosphorus stoichiometry and food chain production. Ecol Lett 1:146-150
Sterner RW, Elser JJ (2002) Ecological Stoichiometry: The Biology of Elements from Molecules to the Biosphere, Vol. Princeton University
Press, Princeton, NJ
Sugden H, Lenz M, Molis M, Wahl M, Thomason JC (2008) The interaction between nutrient availability and disturbance frequency on the
diversity of benthic marine communities on the North East coast of England. J Anim Ecol in press
Taraldsvik M, Myklestad SM (2000) The effect of pH on growth rate, biochemical composition and extracellular carbohydrate production of
the marine diatom Skeletonema costatum. Europ J Phycol 35:189-194
Tillmann U, John U (2002) Toxic effects of Alexandrium spp. on heterotrophic dinoflagellates: an allelochemical defence mechanism
independent of PSP-toxin content. Mar Ecol Prog Ser 230:47-58
Tillmann U, John U, Cembella A (2007) On the allelochemical potency of the marine dinoflagellate Alexandrium ostenfeldii against
heterotrophic and autotrophic protists. J Plankton Res 29:527-543
Tilman D (1982) Resource competition and community structure, Vol. Princeton University Press, Princeton, NJ
Tortell PD, DiTullio GR, Sigman DM, Morel FMM (2002) CO2 effects on taxonomic composition and nutrient utilization in an Equatorial
Pacific phytoplankton assemblage. Mar Ecol Prog Ser 236:37-43
Urabe J, Togari J, Elser JJ (2003) Stoichiometric impacts of increased carbon dioxide on a planktonic herbivore. Glob Chan Biol 9:818-825
Velzeboer RMA, Baker PD, Rositano J (2001) Saxitoxins associated with the growth of the cyanobacterium Anabaena circinalis (Nostocales,
Cyanophyta) under varying sources and concentrations of nitrogen. Phycologia 40:305
Wahl M (2008) Ecological modulation of environmental stress: interactions between UV radiation, epibiotic snail embryos, plants and
herbivores. J Anim Ecol 77:549-557
Wahl M, Molis M, Davis A, Dobretsov S, Durr ST, Johansson J, Kinley J, Kirugara D, Langer M, Lotze HK, Thiel M, Thomason JC, Worm B,
Ben-Yosef DZ (2004) UV effects that come and go: a global comparison of marine benthic community level impacts. Glob Chan Biol
10:1962-1972
Wiencke C, Roleda MY, Gruber A, Clayton MN, Bischof K (2006) Susceptibility of zoospores to UV radiation determines upper depth
distribution limit of Arctic kelps: evidence through field experiments. J Ecol 94:455-463
Wiltshire KH, Malzahn AM, Wirtz K, Greve W, Janisch S, Mangelsdorf P, Manly BFJ, Boersma M (2008) Resilience of North Sea
phytoplankton spring blooms dynamics: an analysis of long term data at Helgoland Roads. Limnol Oceanogr in press
Zimmerman RC, Kohrs DG, Steller DL, Alberte RS (1997) Impacts of CO2 enrichment on productivity and light requirements of eelgrass.
Plant Physiol 115:599-607
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11.6: Theme 5: Integrated assessment – sensitivities and uncertainties
German Advisory Council on Global Change (WBGU, Berlin 2006) states: “Because of the
importance of the consequences of ocean acidification, research in this area should be intensified
considerably. As long as there is no general scientific consensus about the tolerable limit for the
effects of acidification, a margin of safety according to the precautionary principle should be
observed. The suggestion of the WBGU to prevent a pH decrease of more than 0.2 is oriented
toward the goal of avoiding an aragonite undersaturation in the ocean surface layer. If it is found
that other intolerable damages already occur before reaching aragonite undersaturation, then the
guard rail will have to be adjusted accordingly.”
As stated in the report, the tolerable window for ocean acidification defined by WBGU presently
relies on an extremely small data base. In fact, rather than using the limited data on observed
biological consequences of ocean acidification, the WBGU reaches its recommendation on the
basis of projected changes in water chemistry (aragonite saturation state). While this is an
appropriate approach in view of the scarcity of biological information, there is a clear need to
establish a reliable data base on tolerance levels for ocean acidification in key groups of oceanacidification sensitive marine organisms in order to reach a more informed recommendation.
Theme 5 of BIOACID will take the challenge of integrating the information gained under
Themes 1 to 4 in order to identify the potential thresholds associated with ocean acidification.
Uncertainties, probabilities and risks to the marine environment have to be assessed as well as
their feedback to climate systems. During the first 3-year phase of BIOACID, our main aim is to
develop and establish the tools that will allow us fulfil the BIOACID synthesis needs. For the
three subprojects proposed here, the synthesis tools to be established within BIOACID range
from meta-analysis techniques, over regional and global numerical ecosystem models to
economic methods of integrated assessment. These tools will help to better understand ongoing
changes in chemical and biological state of the North Sea from alkalinity fluxes originating from
the Wadden Sea over a synthesis model that integrates OA sensitivities at organism level into a
North Sea ecosystem model (5.1) to an economical impact assessment. (5.3). Newly developed
assessment tools will also be used to improve parameterisations of calcium carbonate production
in global biogeochemical climate models (5.2). By investigating the combined effects of
variations in temperature and ocean acidity, such parameterisations will allow to put better
constraints on possible threshold levels on ocean acidification in a warming world.
Objectives
• Synthesize information obtained in Themes 1 to 4 to achieve an integrated understanding of
biological responses to ocean change, integrating effects of ocean acidification and warming
• Develop a framework for integrating ocean acidification sensitivities at the organism level into
ecosystem modelling
• Identify critical threshold levels (‘tipping points’) of ocean acidification for irreversible
ecosystem changes providing sound information for adaptation and mitigation measures
• Define dangerous ocean acidification in terms of the goods and ecosystem services lost
247
5.1 Impact of Alkalinity fluxes from the Wadden Sea on the carbon cycle and the primary
production in the North Sea.
The central objective is to relate the Wadden Sea’s AT flux to the pelagic and benthic
environment, with emphasis on biogeochemical cycling of carbon in conjunction with other
nutrients, such as nitrogen. The impact of the AT flux from the Wadden Sea on the general North
Sea carbon cycling. and on the long-term trend with respect to the overall pH signal within the
North Sea will be investigated. A future scenario model simulation will be performed and
analysed with respect to critical ecological and biogeochemical regime shifts, as we impose an
atmospheric partial pressure (pCO2) of 1000 µatm.
5.2. Evaluating and optimising parameterisations of pelagic calcium carbonate production in
global biogeochemical ocean models.
The aim is to critically review existing parameterisations of calcification currently used in largescale biogeochemical climate models and to compile published experimental findings on
temperature- and pCO2-sensitivities of calcium carbonate production, its export and dissolution.
The ability of current parameterisations of calcification in biogeochemical models to reproduce
observed alkalinity fields will be quantitatively assessed and a Bayesian meta-analysis will be
applied to BIOACID experimental findings in order to condense distributed information into
plausible model parameterisations.
5.3. Viability-method for the impact assessment of ocean acidification under uncertainty Development of the method and exemplary application to the impact of acidification on the North
Sea cod fishery.
This project will further develop the ecological-economic viability-method towards a general
approach for integrated assessment of human actions influencing ocean acidification and the
consequences for human well-being that takes uncertainties about future development into
account.
The usefulness of the viability-method will be demonstrated by applying it exemplarily to the
assessment of acidification effects on the North Sea cod fishery.
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Project 5.1: Impact of Alkalinity fluxes from the Wadden Sea on the carbon cycle
and the primary production in the North Sea
PI: Johannes Pätsch (1), Co-PIs Helmuth Thomas (2), Markus Schartau (3)
(1) Institute of Oceanography, Hamburg; (2) Dalhousie University, Halifax, NS, Canada; (3)
GKSS Forschungszentrum Geesthacht
i.
Objectives
Recent findings suggest that the Wadden Sea at the southeastern North Sea acts as a major source
of total Alkalinity (AT) (Thomas et al., subm.). This will affect the carbon cycling within the
North Sea and is expected to have an impact on the overall primary production in pelagic and
benthic systems. During summer the AT release lowers the marine partial pressure of carbon
dioxide (pCO2) and the CO2 release to the atmosphere, in particular in the southern bight of the
North Sea. This AT flux buffers the pH decline, induced by anthropogenic CO2, as already
observed in the area of the North Sea (Thomas et al., 2007). Yet, the extent to which the AT flux
from the tidal flat areas of the Wadden Sea can buffer pH and its response to the biological
ecosystem has to be assessed.
The central objective is to relate the Wadden Sea’s AT flux to the pelagic and benthic
environment, with emphasis on biogeochemical cycling of carbon in conjunction with other
nutrients, such as nitrogen. With our modelling activity within BIOACID we specifically propose
the following:
•
We will use an existing ecosystem of the North Sea (ECOHAM) to extrapolate
information’s gathered from local observations to the entire North Sea.
•
We will quantify the impact of the temporally resolved AT flux from the Wadden Sea on
the general North Sea carbon cycling.
•
We will investigate to which extent this AT flux affects the long-term trend with respect
to the overall pH signal within the North Sea.
•
The response of primary producers to pH changes will be addressed by extrapolating
results from mesocosm and chemostat experiments in an attempt to crudely discriminate
non-calcifying from calcifying primary producers.
•
A future scenario model simulation will be performed and analysed with respect to
critical ecological and biogeochemical regime shifts, as we impose an atmospheric partial
pressure (pCO2) of 1000 µatm.
The proposed modelling activities integrate very well with the BIOACID-investigations of Prof.
Dr. Böttcher and Prof. Dr. Liebezeit on biogenic carbonates in the Wadden Sea. In addition, we
will consider characteristic parameter values of phytoplankton productivity that are derived from
data measured under different growth conditions by Dr. Engel and her group. This will provide
an excellent basis for assessing our modelling approach of “excess production” (Pätsch & Kühn,
2008) and “carbon overconsumption” (Schartau et al., 2007), which refers to the description of
carbon fixation under nutrient limited conditions (Fogg, 1983).
249
Continental shelves play a key role in the global cycling of biogeochemically essential elements
(Christensen, 1994; Jickells, 1998). From observations in the East China Sea, Tsunogai et al.
(1999) concluded that the global shelves act as a sink for atmospheric carbon dioxide and as a
source of carbon for the ocean (Borges et al., 2005). The North Sea, as part of the NorthwestEuropean shelf, has been characterized as a sink for atmospheric CO2 (Thomas et al., 2004).
These findings were confirmed by simulations, using the ecosystem model ECOHAM (Pätsch &
Kühn, 2008) during a Diploma study (Prowe, 2006). The dependency of the carbon fluxes in the
North Sea on the Alkalinity especially on the Alkalinity produced in the Wadden Sea is under
debate (Thomas et al., in prep). For the southern North Sea a modelling effort was successful
focusing on the effect of phytoplankton blooms on the carbonate system (Gypens et al., 2004).
On large scales, the multi-functional plankton modelling approach needs to be gradually and
slowly substantiated rather than precipitated; from this perspective, we will coordinate our
modelling activity during the proposed project in collaboration with our neighbor Institute of
Hydrology and Fisheries (Diekmann, et al., submitted), Institute for Coastal Research at GKSS,
and the Alfred Wegener Institute (Dr. Engel and her group) within BIOACID.
Johannes Pätsch has worked since two decades in the field of numerical modelling. In the last
years he has applied (Pätsch & Radach, 1997) and developed (Pätsch, et al. 2002)
biogeochemical models for different marine environments mainly shelf seas. He has gathered
experience in plankton dynamics and their mathematical expression in complex ecosystem
models during the European MAST project ERSEM (1990-1997). He has also applied statistical
methods for large data sets (Radach & Pätsch, 1997). The main focus of his work is on the
interplay between biogeochemical fluxes of carbon and nitrogen in various marine environments
(Pätsch et al., 2002; Pätsch & Kühn, 2008).
Helmuth Thomas has worked since many years in the field of carbon dynamics in marine
biogeochemical environments. Using field data sets, which resolve the North Sea system in time
and space, he established the first annual carbon budget of this shelf sea (Thomas et al., 2004,
2005). Furthermore he characterized the North Sea as a shelf sea which is highly vulnerable to
acidification processes (Thomas et al., 2007).
Markus Schartau has gathered experience in optimisation of ecosystem models based on
sophisticated data-assimilation methods (Schartau et al., 2001; Schartau and Oschlies, 2003). He
has participated in international projects that address model assessment and validation, but also
the modelling of functional plankton groups (e.g. Friedrichs et al, 2006; Hood et al., 2006).
During the last years his work focused on the development of biological models with variable
stoichiometry, with emphasis on carbon overconsumption in conjunction with abiotic organic
particle formation (Schartau et al., 2007). He is involved as principle investigator in the two
collaborating projects EPOCA (European Project on Ocean Acidification) and SOPRAN (Surface
Ocean Processes in the Anthropocene).
250
•
5.1.a: We will simulate the biogeochemistry of the North Sea with the ecosystem model
ECOHAM (Pätsch & Kühn, 2008) for recent years with prescribed Alkalinity fields (reference
run). The model includes a bulk formulation for calcifying phytoplankton. In order to improve
the already existing bulk parametrization of calcification this mechanism and the pCO2
sensitivities will be compared cautiously with M. Hoppema’s approach in project 3.4.3
(Buffering ocean acidification: Dissolution of carbonate sediments in the Southern Ocean).
•
5.1.b: In cooperation with the Institute of Hydrobiology and Fishery (Uni-Hamburg) and the
BIOACID-community we will perform cross-validation experiments with our already existing
plankton module, while separating between the three phytoplankton groups: calcifiers, silicifiers
and flagellates. The model-system with this plankton module will also be tested against the
reference run.
•
5.1.c: Results from experiments of A. Engel; project 1.2.1 (Production and decomposition of
exudates) will allow us to refine model parameterisations with respect to the production of
dissolved organic matter and its dependence on pH, temperature, and nutrient availability. Data
from M. and G. Nausch; project 1.2.3 (DOM availability and phosphorus utilization) will also
be regarded. All implementations and parametrizations, which concern the cycling of DOM, will
be aligned with the developments in the project 1.3 of B. Schneider (Modelling biogeochemical
feedbacks of the organic carbon pump).
•
5.1.d: In cooperation with the group of M. Böttcher; project 3.4.1 (Impact of biogenic
carbonates on pH buffering in an acidifying coastal sea (North Sea)) we will estimate the
recent temporally resolved flux of AT from the Wadden Sea into the open North Sea during low
tides. During high tides when the water is flushing the Wadden Sea we will provide the simulated
pH, as well as DIC and Alkalinity concentration to this group. The described boundary data will
be used to run our model with prognostic Alkalinity dynamics. For further model confirmation,
the GKSS will provide data from “ships of opportunity” (fluorescence, pH, temperature, salinity).
This data set can be updated in the near future (during BIOACID funding period) with pCO2
Ferry Box measurements, in collaboration with Prof. Friedhelm Schroeder (GKSS). At the
beginning of BIOACID, the sensors for pCO2 will not be in operational use and will also need to
be calibrated. Until now the carbon budget of the Wadden Sea is not fully understood. Therefore
the investigations by R. Asmus, project 4.1.1 (Effects of ocean acidification on trophic
interactions in coastal seaweed and seagrass ecosystems) are of great importance for our
studies.
5.1.e: In a first sensitivity study we will vary the amount of AT flux from the Wadden Sea, in
order to study the pH variation of the North Sea.. The impact of these pH variations will be
investigated with respect to 1) carbon fluxes in the North Sea and 2) primary production in
conjunction with the production of dissolved organic matter.
5.1.f: In a second sensitivity study we will run the model with an atmospheric pCO2 of 1000
atm. The impact of the corresponding pH changes on different primary producers will be
examined. We hypothesise a significant response in primary production that translates into
modifications in the overall carbon flux in the North Sea and across the boundaries of the North
Sea. The latter results can be identified as the exchange between the North Atlantic (open ocean)
and the Northwest European Shelf. We will compare the dynamics of this explicitly resolved
fluxes with those parameterised in global modelling approaches (project 5.2: Evaluating and
optimising parameterisations of pelagic calcium carbonate production in global
biogeochemical ocean models).
•
•
•
5.1.g: The results will be documented by an article within an international journal.
251
Work Schedule
5.1
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
5.1.a
5.1.b
5.1.c
5.1.d
5.1.e
5.1.f
5.1.g
Milestones (5.1)
- Establishing and evaluation of the reference run
- Implementation of the new plankton module
- Exchange of data with other institutions
- Sensitivity study 1
- Sensitivity study 2
252
3 months
18 months
27 months
30 months
33 months
First Year
Second Year
Third Year
Total
Personnel costs
PhD (1/2 job)
Three
months
Post-Doc
(Markus Schartau, GKSS)
Subtotal
Consumables
1 Personal Computer for the
PhD student
Travel
Uni Hamburg:
EGU meetings
Uni Hamburg:
BIOACID meetings
Uni Hamburg:
Cooperation Halifax
GKSS: EGU meetings
GKSS:
BIOACID meetings
GKSS:
Cooperation Halifax
Subtotal
Investment
Other costs
Total Uni Hamburg
Total GKSS
TOTAL
Personnel costs: The University of Hamburg will employ a PhD student on a half time position.
This student will be supervised by Johannes Pätsch and Markus Schartau. Markus Schartau will
give substantial advice and support in data assimilation and parameter optimisation techniques in
conjunction with the proposed model development. Since his contribution is not part of the
Helmholtz Association (HGF) research programme (PACES), one post-doc monthly salary will
be allocated to the GKSS per year.
Consumables: 1 PC: The employee needs access to our computer center. Simulation results will
be evaluated with the PC.
Travel: Results from the proposed BioAcid model simulations will be presented at EGU
meetings. Different aspects of the here proposed topics are likely to be addressed at the annual
253
meetings, allowing for presentation that are given not only by the PhD student but also by the
advisors. The PhD student is expected to gain international experience in the field of
biogeochemical modelling. The EGU meetings are a prominent place to learn this item.
For cooperation with Dalhousie University we intend to have at least one meeting per year, either
in Halifax, or in Hamburg. Travel money for GKSS partner will be allocated in order to join at
least one abroad meeting in Halifax, Canada.
vi. References
Anderson TR (2005) Plankton functional type modelling: running before we can walk? Journal of Plankton Research 27(11): 1073-1081
Borges AV, Delille B, Frankignoulle M (2005) Budgeting sinks and sources of CO2 in the coastal ocean: Diversity of ecosystems counts.
Geophys Res Let 32 L14601 doi:10.1029/2005GL023053
Christensen JP (1994) Carbon export from continental shelves, denitrification and atmospheric carbon dioxide. Continental Shelf Research 14:
547-576
Fogg GE (1993) The ecological consequence of extracellular products of phytoplankton photosynthesis. Bot Mar XXVI: 3-14
Gypens N, Lancelot C, Borges AV (2004) Carbon dynamics and CO2 air-sea exchanges in the eutrophied coastal waters of the southern bight
of the North Sea: a modelling study. Biogeoscience 1: 147-157
Hood RR, Laws EA, Moore, JK, Armstrong RA, Bates N, Brown C, Carlson C, Chai F, Doney SC, Ducklow H, Falkowski P, Feely RA,
Friedrichs M, Landry M, Nelson D, Richardson T, Salihoglu B, Schartau M, Wiggert J (2006) Functional Group Modelling: Progress,
challenges and prospects. Deep Sea Research II 52: 459-512
Jickells TD (1998) Nutrient biogeochemistry of the coastal zone. Science 281: 217-222
Friedrichs MAM, Dusenberry J, Anderson L, Armstrong R, Chai F, Christian J, Doney SC, Dunne J, Fujii M, Hood R, McGillicuddy D,
Moore K, Schartau M, Spitz YH, Wiggert J (2006). Assessment of skill and portability in regional marine biogeochemical models: The
role of multiple phytoplankton groups. Journal of Geophysical Research available: http://www.ccpo.odu.edu/RTBproject/Publications
Pätsch J, Kühn W (2008) Nitrogen and carbon cycling in the North Sea and exchange with the North Atlantic - a model study, Part I. Nitrogen
budget and fluxes. Continental Shelf Research 28: 767-787
Pätsch J, Kühn W, Radach G, Santana Casiano JM, Gonzalez Davila M, Neuer S, Freudenthal T, Llinas O (2002) Interannual variability of
carbon fluxes at the North Atlantic station ESTOC. Deep-Sea Res II 49(1-3): 253-288
Pätsch J, Radach G (1997) Long-term simulation of the eutrophication of the North Sea: temporal development of nutrients, chlorophyll and
primary production in comparison to observations. Journal Sea Research 38: 275-310
Prowe F (2006) Simulating and budgeting the carbon fluxes in the North Sea (2001/2002). Diploma Thesis University of Oldenburg: 1-128
Radach G, Pätsch J (1997) Climatological annual cycles of nutrients and chlorophyll in the North Sea. Journal of Sea Research 38: 231-248
Schartau M, Engel A, Schröter J, Thoms S, Völker C, Wolf-Gladrow D (2007) Modelling carbon overconsumption and the formation of
extracellular particulate organic carbon. Biogeosciences 4: 433-453; open access: www.biogeosciences.net/4/433/2007/bg-4-4332007.html
Schartau M, Oschlies A (2003) Simultaneous data-based optimisation of a 1D-ecosystem model at three locations in the North Atlantic Ocean:
Part 1) Method and parameter estimates. Journal of Marine Research 61(6): 765-793
Schartau M, Oschlies A, Willebrand J (2001) Parameter estimates of a zero-dimensional ecosystem model applying the adjoint method. Deep
Sea Research II 48: 1769-1800
Thomas H, Prowe AEF, van Heuven S, Bozec Y, de Baar HJW, Schiettecatte L-S, Suykens K, Kone M, Borges AV, Lima ID, Doney SC
(2007) Rapid decline of the CO2 buffering capacity in the North Sea and implications for the North Atlantic Ocean. Global Biogeochem.
Cycles 21 doi:10.1029/2006GB002825
Thomas H, Bozec Y, de Baar HJW, Elkalay K, Frankignoulle M, Schiettecatte L-S, Kattner G, Borges AV (2005) The carbon budget of the
North Sea. Biogeoscience 2: 87-96
Thomas H, BozecY, Elkalay K, deBaar HJW (2004) Enhanced open ocean storage of CO2 from shelf sea pumping. Science, 304, 5673: 10051008
Thomas H, Schiettecatte L-S, Suykens K, Kone YJM, Bozec Y, de Baar HJW, Borges AV (in prep.) Anaerobic oxidation of organic matter - a
major sink for atmospheric CO2 in the coastal ocean
Tsunogai S, Watanabe S, Sato T (1999) Is there a 'continental shelf pump' for the absorption of atmospheric CO2? Tellus 51B: 701-712
254
Project 5.2: Evaluating and optimising parameterisations of pelagic calcium
carbonate production in global biogeochemical ocean models
PI: A. Oschlies; Co-PIs: I. Kriest, W. Koeve; IFM-GEOMAR Kiel
i. Objectives
We propose
-
to critically review existing parameterisations of calcification currently used in largescale biogeochemical climate models,
-
to compile published experimental findings on temperature- and pCO2-sensitivities of
calcium carbonate production, its export to the interior of the oceans and its dissolution,
and to put these findings in perspective with the model parameterisations,
-
to quantitatively assess the ability of current parameterisations of calcification in
biogeochemical models to reproduce observed alkalinity fields, and to suggest improved
parameterisations that can reliably predict the response of pelagic calcium carbonate
production to variations in both temperature and ocean carbonate chemistry,
-
to initiate a Bayesian meta-analysis of BIOACID experimental findings
Two aspects of global change, warming and acidification of the ocean, will most likely affect
future global calcification rates and thereby influence the ability of the ocean to sequester
anthropogenic CO2. Current biogeochemical climate models use various, often pragmatic,
formulations of the production of particulate inorganic carbon. Parameterisations of the
sensitivity of calcification to acidification include nonlinear dependencies on the CaCO3
saturation state (Gehlen et al., 2007; Ridgewell et al., 2007), or linear relationships with pCO2
(Heinze, 2004). While these parameterisations all predict a decrease of calcification in response
to acidification, the models also include responses to temperature changes. These are accounted
for either explicitly or implicitly via (a) temperature dependent photosynthesis, in conjunction
with a linear coupling of autotrophic calcification (e.g., Archer et al., 1998), (b) direct
temperature dependence of the calcification rate (e.g., Maier-Reimer, 1993; Archer et al., 2000,
Aumont and Bopp, 2006), or (c) nonlinear effects between the temperature dependent growth and
loss rates of phytoplankton functional types, including coccolithophores (Gregg and Casey, 2007;
Aumont and Bopp, 2006). The combined effects of warming and acidification are uncertain and,
in current climate models, have been shown to result either in a simulated decrease (Gehlen et al.,
2007) or increase (Schmittner et al., 2008) of calcification over the next centuries.
Prof. Andreas Oschlies is a biogeochemical modeller with extensive experience in simulating
the interaction of physical transport and biotic processing of biogeochemical tracers at basin and
global scales. Data assimilation methods have been used to optimise the difficult-to-constrain
parameters of marine ecosystem models (Schartau and Oschlies, 2003a,b; Oschlies and Schartau,
255
2005), and mechanistic models have been developed to improve descriptions of nutrient uptake
and particle export (Kriest and Oschlies, 2007, 2008).
Dr. Iris Kriest is a biogeochemical modeller with particular interest in large scale particle flux
and particle dynamics (Kriest and Evans, 1999; 2000; Kriest, 2002; Kriest and Oschlies, 2008).
Recently she implemented, in close collaboration with Dr. S. Khatiwala, the tracer-transportmatrix approach of Khatiwala et al. (2005) and Khatiwala (2007) at the IFM-GEOMAR
Biogeochemical Modelling group. Currently she applies this approach to optimise
parameterisations of organic matter export and remineralisation profiles by evaluating simulated
nutrient and oxygen fields against observed data, a work that will directly feed into this project.
Dr. Wolfgang Koeve is a biological oceanographer with particular experience in the analysis and
evaluation of biogeochemical datasets from local to global scales (Koeve and Ducklow, 2001;
Koeve et al. 2002; Koeve, 2006). Focus of his recent work was on the coupling of nitrogen and
carbon cycles (Koeve, 2004; 2005), as well as on synthesis of organic and inorganic particle flux
estimates (Koeve, 2002).
Task 5.2.a
Parameterisations of pelagic calcium carbonate production currently used in biogeochemical
climate models will be analysed with respect to their sensitivity to temperature and carbonate
chemistry. These parameterisations are not always well documented in the relevant literature, and
a careful investigation of technical reports and computer codes will in many cases be required. A
first attempt to collect this information has shown that this is not a trivial task and that the results
of such a review of existing parameterisation would be of interest to many modellers that have
already been using such models for some time.
Task 5.2.b
A meta-analysis of hydrographic data and published experimental findings will be used in a first
step to evaluate existing parameterisations. In collaboration with WP9 of the European EPOCA
project (led by PI A. Oschlies), which will provide a synthesis of a number of mesocosm
experiments, we will compile the relevant information with the aim to develop improved
parameterisations of pelagic CaCO3 production and its sensitivity to environmental conditions.
This will, in particular, require careful consideration of the different experimental protocols,
which may help to reconcile apparently contradictory experimental findings (e.g., Riebesell et al.,
2000; Langer et al., 2006; Iglesias-Rodriguez et al., 2008).
Task 5.2.c
To obtain an objective assessment of the different parameterisations of calcification and its
sensitivity to variations in temperature and carbonate chemistry, we will combine the various
parameterisations with a new computational framework which allows efficient simulations of
biogeochemical tracers over hundreds to thousands of years, needed to model the impact of
changed carbonate production, export and dissolution parameterisations on the alkalinity field.
Export formulations to be investigated range from globally uniform remineralisation length
scales (Maier-Reimer, 1993) to complex ones invoking processes such as a link between organic
carbon to CaCO3 flux via the ballast particle flux hypothesis (Armstrong et al., 2001; Heinze,
2004), aggregation, or calcite dissolution in copepod guts or in marine aggregates (Aumont and
Bopp, 2006; Gehlen et al., 2007).
256
The agreement between simulated and observed alkalinity fields will then be quantified by a
“misfit function” that measures how well the individual parameterisations can reproduce current
regional variations in CaCO3 production, export and dissolution as a function of regional
variations in temperature and carbonate chemistry. Impacts of model errors in the circulation and
organic matter fluxes will be accounted for by a complementary analysis of the model-data
misfits of hydrographic and biogeochemical tracer distributions. All simulations will be carried
out under present environmental conditions, but it is expected that the emerging optimised
parameterisations will improve future climate scenario simulations performed elsewhere.
Task 5.2.d
While tasks5.2.a and 5.2.b out of necessity base their synthesis and modelling work on preBIOACID experimental and modelling efforts, task 5.2.d intends to use novel (novel with respect
to oceanographic applications) Bayesian meta-analysis methods to analyse the experimental
findings obtained within themes 1 to 4 of the BIOACID project. Twelve European PhD students
are currently trained in these techniques as part of the METAOCEANS Marie-Curie action. They
all work on the application of various meta-analysis techniques to different aspects of marine
ecosystems and are expected to finish their PhDs in 2010. We hope that BIOACID can take
advantage of the METAOCAENS project and offer a PostDoc position to one of theses welltrained young scientists.
A start as early as 1.5 years into the project should allow for a timely analysis of the experimental
findings already during the first BIOACID phase. This will allow the rapid identification of
remaining uncertainties as well as of the potential for uncertainty reduction. As such, it is
expected that this initiative will allow for an immediate feedback into experimental strategies,
which should be of benefit to the entire BIOACID consortium. This will also establish able
personnel and potent analysis methods in preparation for a second BIOACID phase that only will
allow the completion of a sound synthesis of the BIOACID experimental findings.
Interaction with other projects and subprojects
Concerning tasks 5.2.a to 5.2.c, this project will develop strong links to projects from theme 3
(Calcification), in particular to projects 3.1 (Cellular mechanisms of calcification; in particular
3.1.1 Coccolithophores) and 3.2 (Calcification under pH stress; in particular 3.2.1: Sub-polar
pteropods). Research carried out in project 3.4 (Microenvironmentally controlled
(de)calcification mechanisms; in particular 3.4.3) and 5.1 (Alkalintiy fluxes in the North Sea) will
help to clarify the longer term needs for (a) a sediment model compartment and (b) shelf to open
ocean interactions in the global modelling perspective. Bayesian meta-analysis (Task 5.2.d) shall
be applied to experimental work planned in Theme 1 (1.2 Turnover of organic matter).
Descriptions of acidification impacts on the production and export of organic and inorganic
carbon needed in 5.2 will rely on experimental evidence gathered particularly in subprojects 1.2.1
(Production and decomposition of exudates), 1.2.2 (Dissolved organic nitrogen release and
uptake unders stress), and 1.2.5 (Effect of decreasing calcareous/lithogenic ballast on aggregates
in the benthic boundary layer). The global modelling work of Project 5.2, which uses a stationary
present-day climate to evaluate model results against observations, has close links to the
prognostic climate model runs performed in Project 1.3 (Organic carbon pump feedbacks) and
will further benefit from long-term (geological) perspectives as provided by 3.5.2 and 3.5.3 (Past
ocean acidification events).
257
Schedule
5.2
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Review of IPCC-type model
parameterizations of marine calcium
carbonate cycling
Review of published experimental
findings (POC-net production, PIC
production, DOM, TEP, C:N, N2fix)
Set-up of prototype 3D model, first
exploratory model experiments
Preparation of datasets for model-data
comparison (from GLODAP,
CARBOOCEANS, and PANGAEA
sources).
Development of data analysis tools
Model experiments applying a variety
of formulations for CaCO3 production,
export and dissolution
In-depth analysis of model results
through comparison with observations
(GLODAP etc.) and subsequent
parameter optimization
Compilation of datasets from
BIOACID experiments from the first
1.5 years of project into one database
Milestones (5.2)
- Draft documentation (manuscript) on parameterization of marine
calcium carbonate cycling in IPCC type models
- Review (manuscript draft) on experimental work
- Prototype 3D model with carbonate cycle processes implemented,
major data analysis tools and datasets prepared
- Implementation of at least three distinct sets of carbonate models
- Three manuscripts submitted
month 9
month 18
month 21
month 24
month 36
First Year
Personnel costs
5.2.a–c
5.2.d
Subtotal
Consumables
5.2.a–c
5.2.d
Subtotal
Travel
5.2.a-c
258
Second Year
Third Year
Total
First Year
Second Year
Third Year
Total
5.2.d
Subtotal
Investments
Subtotal
Other costs
Subtotal
Total
5.2.a-c
Personnel costs: 1 full time researcher, Dr. W. Koeve, shall be employed to carry out the
planned review work (see objective 1+2), do the model experiments as well as model-data
analysis (objective 3) and publishing in peer reviewed journals. Both the review work as well as
the model-data analysis require a high level of scientific expertise provided by the candidate, who
has f.e. served as guest editor of a Deep-Sea Research II volume on carbon cycle synthesis,
convenor at scientific conferences, and reviewer for various funding agencies and scientific
journals
Consumables: Euro/yr for publication fees in open access journals (e.g. Biogeosciences) or
journals with fee-based open access option (e.g. Limnology Oceanography).
Travel: Euro/yr are requested for annual 2-week visits of Dr. Samar Khatiwala (Columbia
University, New York) to IFM-GEOMAR.
Euro/yr are requested for travel to international
and national conferences and workshops AGU Autum-2009, AGU Ocean Sciences- 2010, EGU2011, in order to present scientific results of the project.
Investment: no
Other costs: no
5.2.d
Personnel costs: 1 NN PostDoc for 1.5 years (second half of project). The PostDoc will be
responsible to carry out the Bayesian meta-analysis of BIOACID experimental findings.
Currently the EU Marie Curie action METAOCEANS trains PhD students in meta-analysis
techniques. METAOCEANS students will finish their PhDs in 2010, we are hence expecting a
number of well trained PostDocs available in summer 2010, being potential candidates for this
BIOACID task.
Consumables:
Euro for third year. Publication fees.
Travel:
Euro for third year only. Presentation of scientific results at national and international
meetings; potentially at the AGU autumn meeting in 2001 (San Francisco).
Investment: no
Other costs: no
259
vi. References
Archer D, Kheshki H, Maier-Reimer E (1998) Dynamics of fossil fuel CO2 neutralization by marine CaCO3. Global Biogeochem Cycles 12(2):
259-276
Archer D, Winguth A, Lea D, Mahowald N (2000) What caused the glacial/interglacial atmospheric pCO2 cycles? Rev Geophysics 38(2): 159189
Armstrong RA, Lee C, Hedges JI, Honjo S, Wakeham SG (2001) A new, mechanistic model for organic carbon fluxes in the ocean based on
the quantitative association of POC with ballast minerals. Deep-Sea Res II 49: 219-236
Aumont O, Bopp L (2006) Globalizing results from ocean in situ fertilization studies. Global Biogeochem Cycles 20: GB2017,
doi:10.1029/2005GB002591
Gehlen M, Gangto R, Schneider B, Bopp L, Aumont O, Ethe C (2007) The fate of pelagic CaCO3 production in the high CO2 ocean: a model
study. Biogeosciences 4: 505-519
Gregg W, Casey N (2007) Modeling coccolithophores in the global oceans. Deep-Sea Res II 54: 447-477
Heinze C (2004) Simulating oceanic CaCO3 export production in the greenhouse. Geophys Research Lett 31: L16308,
doi:10.1029/2004GL020613
P, Rehm E, Armbrust EV, Boessenkool KP (2008) Phytoplankton calcification in a high-CO2 world. Science 320: 336-340
Khatiwala SA (2007) A computational framework for simulation of biogeochemical tracers in the ocean. Global Biogeochem Cycles 21:
GB3001, doi:10.1029/2007GB002923
Khatiwala S, Visbeck M, Cane MA (2005) Accelerated simulation of passive tracers in ocean circulation models. Ocean Modelling 9: 51-69
Koeve W (2002) Upper ocean carbon fluxes in the Atlantic Ocean - the importance of the POC:PIC ratio. Global Biogeochem Cycles 16:
GB1056, doi:10.1029/2001GB001836
Koeve W (2004) Spring bloom carbon to nitrogen ratio of net community production in the temperate N. Atlantic. Deep-Sea Res I 51: 15791600
Koeve W (2005) The significance of the TEP-pump to deep ocean carbon fluxes. Mar Ecol Prog Ser 291: 53-64
Koeve W (2006) Stoichiometry of the biological pump in the North Atlantic - constraints from climatological data. Global Biogeochem Cycles
20: GB3018, doi:10.1029/2004GB002407
Koeve W, Ducklow H (2001) JGOFS synthesis and modelling: the North Atlantic ocean. Deep-Sea Res Part II 48: 2141-2154
Koeve W, Pollehne F, Oschlies A, Zeitzschel B (2002). Storm induced convective export of organic matter during spring in the northeast
Atlantic. Deep-Sea Res I 49: 1431-1444
Kriest I (2002) Different parameterizations of marine snow in a 1D-model and their influence on representation of marine snow, nitrogen
budget and sedimentation Representing phytoplankton aggregates in biogeochemical models. Deep-Sea Res I 49: 2133-2162
Kriest I, Evans GT (1999) Representing phytoplankton aggregates in biogeochemical models. Deep-Sea Res I 46:1841-1859
Kriest I, Evans GT (2000) A vertically resolved model for phytoplankton aggregation. P Indian Acad Sci – Earth 109: 553-469
Kriest I, Oschlies A (2007) Modelling the effect of cell-size-dependent nutrient uptake and exudation on phytoplankton size spectra. Deep-Sea
Res I 54:1593-1618
Kriest I, Oschlies A (2008) On the treatment of particulate organic matter sinking in large-scale models of marine biogeochemical cycles.
Biogeosciences 5: 55-72
Langer G, Geisen M, Baumann K-H, Kläs J, Riebesell U, Thoms S, Young JR (2006) Species-specific responses of calcifying algae to
changing seawater carbonate chemistry. Geochemistry Geophysics Geosystems 7: Q09006, doi:10.1029/2005GC001227
Maier-Reimer E (1993) Geochemical cycles in an ocean general circulation model. Preindustrial tracer distributions. Global Biogeochem
Cycles 7(3): 645-677
Oschlies A, Schartau M (2005) Basin-scale performance of a locally optimised marine ecosystem model. Journal of Marine Research 63: 335358
Ridgwell A, Zondervan I, Hargreaves JC, Bijma J, Lenton TM (2007) Assessing the potential long-term increase of oceanic fossil fuel CO2
uptake due to CO2-calcification feedback. Biogeosciences 4: 481-492
Schartau M, Oschlies A (2003) Simultaneous data-based optimization of a 1D-ecosystem model at three locations in the North Atlantic Ocean:
Part 1. Method and parameter estimates. Journal of Marine Research 61: 765-793
Schartau M, Oschlies A (2003) Simultaneous data-based optimization of a 1D-ecosystem model at three locations in the North Atlantic Ocean.
Part 2: Standing stocks and nitrogen fluxes. Journal of Marine Research 61: 795-821
Schmittner A, Oschlies A, Damon H, Galbraith E (2008) Global Biogeochem Cycles 22: GB1013, doi:10.1029/2007GB002953
260
Project 5.3: Viability-method for the impact assessment of ocean acidification
under uncertainty
PI: M. Quaas
Development of the method and exemplary application to the impact of acidification on the
North Sea cod fishery
i. Objectives
The objectives of the project 5.3 are
1. To further develop the ecological-economic viability-method towards a general approach
for integrated assessment of human actions influencing ocean acidification and the
consequences for human well-being that takes uncertainties about future development
into account.
2. To demonstrate the usefulness of the viability-method by applying it exemplarily to
assess effects of acidification on the North Sea cod fishery.
The tolerable-windows approach (Bruckner et al. 1999, Tóth et al. 2002) employed by the
German Advisory Council on Global Change (WBGU) in the Special Report “The Future Oceans
– Warming up, Rising High, Turning Sour“ has been conceptualized as a general framework that
allows an impact assessment for complex systems such as the ocean-climate system. While
uncertainties about the future development of such systems are of high relevance, the tolerablewindows approach does not explicitly address these uncertainties (a recent exemption is Kleinen
et al. 2007). By contrast, the ecological-economic viability approach (Baumgärtner and Quaas
2007), based on the concept of stocks (Faber et al. 2005) and on ecological population viability
analysis (Beissinger and McCullough 2002) allows to assess the sustainability of human actions
under conditions of uncertainty in a general and unified way. Despite many studies on cod
fisheries (e.g. Döring and Egelkraut 2008, Röckmann et al. 2008), the effects of acidification
have not yet been incorporated into a comprehensive ecological-economic modelling analysis.
The viability-method that shall be developed in the proposed project will build upon the results of
two ongoing projects funded by the BMBF within the key area Economic sciences for
sustainability: The proponent Martin Quaas is involved as PI in the project Sustainable use of
ecosystem services under uncertainty and partner in a sub-project of The Concept of Stocks as
Decision Support for Sustainability Policy. Within the first project, a general conception of
ecological-economic viability as a measure of sustainability under uncertainty has been
developed by the proponent and a coauthor (Baumgärtner and Quaas 2007) and to some extent
already been applied using methods of ecological-economic modeling (Quaas et al. 2007, Quaas
and Baumgärtner 2008).The proponent Martin Quaas is leading the Junior Research Group on
Sustainable Fisheries within Kiel's Cluster of Excellence Future Ocean, where currently
ecological-economic models of fisheries are developed.
261
Building on the ecological-economic concept of viability (Baumgärtner and Quaas 2007), a
“viability-method” shall be developed that allows assessing the consequences human actions both
leading to ocean acidification and dealing with the consequences of acidification. The
applicability of the method shall be proven by applying it to the case of the North Sea cod
fishery.
For this sake, an ecological-economic model shall be developed that incorporates the effect of
acidification, building on both available ecological knowledge and the results obtained in
BIOACID projects 2.3.1 and 2.3.2 (Effects on top predators) on the susceptibility of cod larvae to
acidification. By means of the viability-method, the impacts of different human actions shall be
assessed (including measures to mitigate acidification or adapting the management of the North
Sea cod fishery), taking into account the uncertainties about the future development of
acidification and the exact impact on cod recruitment.
Schedule
5.3
First Year
I
II
III
Second Year
IV
I
II
III
Third Year
IV
I
II
III
IV
Development of the viability-method
and adaptation to the issue of ocean
acidification
Assessment of the effects of
acidification on the North Sea cod
fishery using the viability-method
Milestones (5.3)
month 30
month 30
month 36
- Viability-method adapted for the issue of ocean acidification
- Ecological-economic model of North Sea cod fishery including acidification
- Two manuscripts submitted
First Year
Personnel costs
Subtotal
Consumables
Subtotal
Travel
Subtotal
Investments
Subtotal
Other costs
Subtotal
Total
262
Second Year
Third Year
Total
Personnel costs: 1 NN (e.g. Sandra Derissen) Postdoc for 1 1/4 years. The PostDoc shall conduct
the main part of the research in collaboration with Martin Quaas. As the BMBF-funded project
Sustainable use of ecosystem services under uncertainty will end in the second half of 2010, it is
very likely that one of the four current PhD students of that project can be hired as Postdoc, who
already has substantial experience in the methods involved.
Consumables: Euro in the last year, mainly to cover publication fees.
Travel: Euro in the last year to present scientific results at national and international conferences,
e.g. the biannual conferences of the European Society for Ecological Economics or the World
Congress of Environmental and Resource Economists.
Investment: no
Other costs: no
vi. References
Baumgärtner S. Quaas MF (revise and resubmit) Ecological-economic viability as a criterion of strong sustainability under uncertainty.
Ecological Economics
Beissinger S, McCullough D (eds.) (2002. Population Viability Analysis. Chigaco: University of Chicago Press
Bruckner T, Petschel-Held G, Tóth FL, Füssel HM, Helm C, Leimbach M Schellnhuber HJ (1999) Climate-change decision support and the
tolerable windows approach, Environmental Modeling and Assessment 4: 217–234
Döring R, Egelkraut TM (2008) Investing in natural capital as management strategy in fisheries: The case of the Baltic Sea cod fishery.
Ecological Economics 64: 634–642
Faber M, Frank K, Klauer K, Manstetten R, Schiller J, Wissel C (2005) On the foundation of a general theory of stocks. Ecological Economics
55(2):155-175
Kleinen, T Petschel-Held G, Bruckner T(2007) The probabilistic tolerable windows approach, submitted to Climatic Change
Quaas M F. Baumgärtner S (2008) Natural vs. financial insurance in the management of public-good ecosystems. Ecological Economics
65:397-406
Quaas MF and Requate T (in preparation). Sushi or fish fingers? How preferences for sustainability affect the sustainability of fisheries
Quaas M ., Baumgärtner S, Becker C, Frank K, Müller B (2007). Uncertainty and sustainability in the management of rangelands. Ecological
Economics 62:251-266
Röckmann C, Schneider UA, St.John MA, Tol RSJ (2008). Rebuilding the Eastern Baltic cod stock under environmental change - a
preliminary approach using stock, environmental, and management constraints, forthcoming in Natural Resource Modeling
Skonhoft A, Quaas MF, Requate T, Ruckes K (in preparation). A bioeconomic modelling analysis of how to optimally fish an age-structured
population
Tóth FL, Bruckner T, Füssel HM, Leimbach M, Petschel-Held G, Schellnhuber HJ (2002). Exploring options for global climate policy: a new
analytical framework. Environment 44 (5):22–34
263
12. Appendices
- Project structure
- Contact data of principal investigators
264
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Project coordination
Ulf Riebesell
0.1
Data management
Ulf Riebesell
0.2
Hans Pörtner
0.3
Subproject
Subproject PI
Code Links to Subprojects
Project and data
management,
training and
infrastructure
Ulf
Riebesell
0
development
Infrastructure
development
pH stat mesocosms
Franz-J.
Sartoris
0.3.1
Development of chemical
optical sensor technology
Athanas
for the determination of
Apostolidis /
pCO2 in fluids of marine
Christian
organisms and the marine
Huber
3.4.2 / 1.1.5 / 2.1.2 /
0.3.2 2.1.3 / 2.3.1 / 2.3.2 /
3.1.3 / 3.1.4 / 3.2.2
environment
Training and transfer
Michael
of knowhow
Meyerhöfer
0.4
Primary
production,
Maren
microbial
Voß
1
processes and
265
All
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
biogeochemical
feedbacks
Impact of changing climate
Acclimation versus
adaptation in
autotrophs
Thorsten
Reusch
1.1
on a
Günter Jost /
chemolithoautotrophic
Klaus
epsilonproteobacterium
Jürgens
1.1.1
1.1.2 / 1.1.3 / 1.1.4 /
1.1.5 / 1.2.4 / 4.1.4
from a pelagic redoxcline
The interplay between
1.1.3 / 1.1.1 / 1.1.4 /
carbon-and iron-availability Michael
1.2.2 / 1.2.4 / 1.3 /
and its impact on
Hippler / Julie
photosynthesis of primary
LaRoche
1.1.2 3.3.1
producers in the ocean
Long-term response of
phytoplankton on climate
change: a
1.1.2 /1.1.4 / 1.3 /
Marius Müller
1.1.3 3.1.1 /3.1.4 / 3.5.2 /
3.5.3/
multidimensional approach
Rapid evolution of key
Thorsten
phytoplankton species to a Reusch / Ulf
high pCO2 ocean
Interactive effects of CO2
concentration and
temperature on
microphytobenthic
266
1.1.2 / 1.1.3 / 3.1.1 /
1.1.4
Riebesell
0.3.2/ 1.1.1 / 3.4.1 /
Ulf Karsten /
Thomas
Hübener
3.5.2/ 4.1.2 / 4.2.1
1.1.5
3.4.2 / 4.1.1
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
biodiversity and
ecosystem function
Turnover of organic
matter
Anja Engel
1.2
Production and
decomposition of exudates
4.1.1/ 4.2.1 / 4.2.2 / 5.1
Anja Engel
1.2.1
Maren Voß
1.2.2 4.2.1 / 4.2.2 / 5.2
/ 5.2
Dissolved organic nitrogen
release and uptake under
stress
Monika
DOM availability and
Nausch /
phosphorus utilization
Günther
1.2.3
4.1.1 / 4.1.4 / 4.2.1. /
4.2.2.
Nausch
Microbial response to
DOM release and
aggregation
Effect of decreasing
calcareous / lithogenic
ballast on aggregates in
the benthic boundary layer
Hans – Peter
Grossart
1.1.1 / 1.1.2 / 1.3 /
1.2.4 3.4.2 / 4.1.4 / 4.2.1 /
5.1 / 5.3
Laurenz
Thomsen /
1.2.5
Giselher Gust
1.2 / 1.3 / 3.1.1 / 3.4.1.
/ 3.4.3 / 5.2
1.1.2/ 1.1.3/ 1.2.1/
Modelling
biogeochemical
Birgit
feedbacks of the
Schneider
1.3
organic carbon pump
no subproject
Birgit
Schneider
1.2.2/ 1.2.3/ 1.2.4/
1.3
1.2.5/ 3.1.1/ 3.2.1/
3.4.3/ 3.5.2 / 3.5.3/ 5.1/
5.2/ 5.3
267
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
Performance
characters:
reproduction,
Hans
growth and
Pörtner
2
behaviours in
animal species
Ocean Acidification and
Effects on grazers and
filtrators
Thomas Brey
2.1
Reproduction: Is the
Angela
beginning of Life in
Köhler
2.1.1
2.2.1/2.3.1 /3.1.3/ 3.1.4
/ 3.2.2 / 4.1.2
Danger?
The response of
zooplankton organisms to
Barbara
elevated CO2
Niehoff
concentrations
acidifying & warming
Gisela Lannig 2.1.3 2.3.2 / 3.1.3 / 3.1.4
/3.2.4 / 3.3.1 / 3.3.2/
Hyas araneus: Sensitivity,
physiological effects
adaptive capacities and
Daniela
evolutionary
Storch
of benthic
268
3.4.1 / 3.5.1 /4.1.2
Long-term
Felix Mark
2.2
consequences in
4.2.1
2.2.1/ 2.2.2 / 2.3.1 /
shallow waters
on different life stages
2.1.2 2.2.1 / 3.1.3 / 3.1.4/
0.3.1 / 0.3.2 / 2.1.2 /
Calcifying
macroorganisms in
0.3.1/ 0.3.2 /2.1.3 /
0.3.1 / 2.1.2 / 2.1.3 /
2.2.1 2.2.2 / 2.3.2 / 3.1.3 /
4.1.2
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
populations from different
crustaceans
latitudes
Cancer pagurus: Chronic
and acute responses –
Christopher
Adaptation versus
Bridges
2.2.2 2.1.3 / 2.2.1 / 3.1.3
Tolerance
Effects of changes in
ocean pH on the
development, growth,
Effects on top
predators (fishes,
cephalopods)
Catriona
Clemmesen
metabolism and
2.3
otolith/statolith formation
and composition of fish
0.3.1 / 0.3.2 / 2.1.1 /
Uwe
Piatkowski
2.3.1
2.1.3 / 2.3.2 / 3.1.3 /
3.1.4/ 3.3.1 / 4.1.2/ /
5.3
and cephalopod early life
stages: a comparative
approach
Mechanisms setting and
compensating for animal
0.3.1 / 0.3.2. / 2.1.3. /
sensitivity to ocean
acidification: functional
Magnus
capacities, thermal
Lucassen
interactions and
mechanism-based
modelling
269
2.2.1. / 2.2.2. / 2.3.1. /
2.3.2 3.1.3. / 3.1.4
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
Calcification:
Sensitivities
Dirk de
across phyla
Beer
3
and ecosystems
Inorganic carbon
Cellular mechanisms
Frank
of calcification
Melzner
acquisition for calcification
3.1
and photosynthesis in
1.1.3 / 1.1.4/ 1.2.5 / 1.3
Kai Schulz
3.1.1 / 3.4.2 / 3.5.2 / 3.5.3 /
marine coccolithophores:
4.2.2 / 5.2
towards a unifying theory
Transepithelial calcification
processes in the
hermatypic cold-water
1.2.5 / 3.2.2 / 3.2.4 /
Armin Form
3.1.2 3.3.1 / 3.3.2 / 3.4.2 /
coral Lophelia pertusa
4.1.4
(Scleractinia)
Calcification & ion
homeostasis in the phylum Frank
mollusca in response to
Melzner
2.1.1 / 2.1.3 / 2.2.1 /
3.1.3 2.2.2 / 2.3.1 / 2.3.2 /
3.2.1 / 3.4.2 / 4.1.2
ocean acidification
Sea urchin membrane
transport mechanisms for
Markus
calcification and pH
Bleich
regulation
270
3.1.4
1.1.3. / 2.1 / 2.3 / 3.2 /
3.3 /3.4.2 / 4.1.2.
Theme
Theme
Leader
Code
Project (Cluster)
Calcification under
Project PI
Ralph
Code
3.2
pH-stress: Impacts on Tollrian
ecosystem and
Subproject
Impact of ocean
Subproject PI
Ulf Riebesell
3.2.1 1.2.5 / 1.3 / 3.1.3 /
acidification and warming
3.2.4 / 3.3.1-3.4.3/
on sub-polar shelled
3.5.1 / 5.2
pteropods
organismal levels
Impact of ocean
acidification on
reproduction, recruitment
2.1.1 / 3.1.2 / 3.2.3 /
Ralph Tollrian 3.2.2 3.2.4 / 3.3.2 / 3.4.2 /
and growth of scleractinian
3.5.2 / 4.1.2 / 4.1.3
corals
Coral calcification in
Claudio
marginal reefs
Richter
Impact of ocean
acidification on coralline
3.4.2/ 4.1.3
Jan Fietzke
3.2.4 /3.2.2 / 3.2.3 / 3.4.2 /
4.1.3 / /4.1.1
Impact of ocean
Ultra-structural
acidification on the
changes and trace
partitioning in
3.1.2/3.2.2/3.2.4/3.3.2/
2.1.3 / 3.1.2 / 3.2.1
red algae
element / isotope
3.2.3
calcification mechanisms
Jelle Bijma
3.3
in marine calcifying
organisms and on ultra
calcifying organisms
structural changes of
(foraminifera, corals)
biogenic calcite
271
2.1.3 / 2.3.1 / 3.1.2 /
Jelle Bijma
3.3.1 3.1.3 / 3.1.4 / 3.2.1 /
3.2.4
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
The effect of decreasing
Anton
pH, salinity and
Eisenhauer
3.3.2 0.4 / 2.1.3 / 2.3.1 /
3.1.2 / 3.1.3 / 3.1.4
temperature on the trace
3.2.1 / 3.2.4
element and isotope
partitioning between
marine calcifying
organisms and seawater
Impact of biogenic
Microenvironmentally
controlled (de-
Michael
)calcification
Böttcher
3.4
carbonates on pH
Michael
buffering in an acidifying
Böttcher
3.4.1
1.2.5 / 2.1.3 / 3.4.2 /
3.4.3 / 5.1
coastal sea (North Sea)
mechanisms
Benthic (de-)calcification
driven by microbial
0.3.2 / 1.1.5 / 3.1 /
Dirk de Beer
3.4.2 3.2.3 / 3.2.4 / 3.3.2
processes
/4.1.3
Buffering ocean
acidification: Dissolution of Mario
carbonate sediments in
Hoppema
3.4.3
1.2.5 / 1.3 / 3.4.1 /
3.4.2 / 5.1 / 5.2
the Southern Ocean
Impact of present and
past ocean
acidification on
metabolism,
biomineralization and
biodiversity of pelagic
and neritic calcifiers
272
Comparison of Cultured
Adrian
Immenhauser
3.5
and Fossil Bivalve
Adrian
Geochemistry and Shell
Immenhauser
Ultrastructure
3.5.1 2.1.3 / 3.5.3 / 4.1.2
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
Biological response to
short-termed ocean
acidification events in the
past: biodiversity and
evolution patterns of
marine primary producers
Jörg
Mutterlose
3.5.2
1.1.3 / 1.1.4/ 1.3 / 3.1.1
/ 3.2.2 / 5.2
(calcareous nannofossils)
during the late Paleocene
– early Eocene
Nannoplankton response
to modern and past ocean
acidification events
Sebastian
Meier
3.5.3
1.1.3 / 1.1.4 / 1.3 /
3.1.1 / 4.2.2 / 5.2
Species
interactions and
community
structure: will
Maarten
ocean
Boersma
4
acidification
cause regime
shifts?
Effects of ocean
OA impacts on
interactions in and
Martin Wahl
4.1
acidification on trophic
interactions in coastal
structure of benthic
273
Ragnhild
Asmus
1.1.5 / 1.2.1 / 3.2.4 /
4.1.1 4.1.2 / 4.1.3 / 4.1.4 /
4.2.1 / 4.2.2 / 5.1
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
seaweed and seagrass
communitiesbenthos
ecosystems
1.1.4 / 2.1.1 / 2.1.3 /
2.2.1 / 2.3.1 / 3.1.3 /
Acidification stress: Early
life stage ecology in times
Martin Wahl
4.1.2 3.1.4 / 3.2.2 / 3.2.3 /
3.5.1 / 4.1.1 / 4.1.3
of global change
4.1.4 / 4.2.1 / 4.2.2
Competitive success of
calcifying and noncalcifying macroalgae
under shifting pH regimes
3.2.2. / 3.2.3. / 3.2.4. /
Kai Bischof
4.1.3 3.4.2. / 4.1.1 / 4.1.2 /
4.1.4 / 4.2.2
in tropical vs. temperate
regions
Effects of ocean
acidification on microbial
community structure,
Alban
composition and activity in Ramette
natural and experimental
1.1.1 / 3.1.2 / 3.4.2 /
4.1.4 4.1.1 / 4.1.2 / 4.1.3 /
4.2.1 / 4.2.2
systems
OA effects on food
webs and competitive
interactions in pelagic
ecosystems
274
Maarten
Boersma
OA effects on pelagic
4.2
community structure and
food chains
Maarten
Boersma
1.2.1 / 1.2.2 / 1.2.3 /
4.2.1 1.2.4. 2.1.2. / 4.1.1 /
4.1.2 / 4.1.4 / 4.2.2
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Subproject
Subproject PI
Competitive interactions in
planktonic microalgae
under OA-stress
Björn Rost
1.2.1 / 1.2.2 / 1.2.3 /
4.2.2 3.1.1 / 3.5.3 / 4.1.1 /
4.1.2 / 4.1.3 / 4.1.4 /
4.2.1
Integrated
assessment:
Andreas
Sensitivities and Oschlies
5
uncertainties
Impact of Alkalinity
fluxes from the
Wadden Sea on the
Johannes
carbon cycle and the
Pätsch
1.2.1 / 1.2.2 / 1.2.3 /
5.1
no subproject
1.3 / 3.4.1 / 3.4.3 /
4.1.1/ 5.2
primary production in
the North Sea
Evaluating and
optimising
1.2.1 / 1.2.2 / 1.2.5 /
parameterisations of
pelagic calcium
Andreas
carbonate production
Oschlies
5.2
no subproject
1.3 / 3.1.1 / 3.2.1 / 3.4
(3.4.3) / 3.5.2 / 3.5.3 /
5.1
in global
biogeochemical ocean
models
275
Theme
Theme
Leader
Code
Project (Cluster)
Project PI
Code
Martin Quaas
5.3.
Subproject
Subproject PI
Viability-method for
the impact
assessment of ocean
acidification under
uncertainty
276
no subproject
2.3.1 / 2.3.2
Appendix: Principal Investigators
Project
Name
Institute
Address
Email
Phone
0.1
Riebesell
Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel
Düsternbrooker Weg 20, 24105 Kiel
[email protected]
0431 600-4444
0.2
0.3
MARUM - Institute for Marine Environmental Sciences , University of Bremen
Alfred Wegener Institute for Polar and Marine Research
Presens Precision Sensing GmbH
Presens Precision Sensing GmbH
Leobener Strasse, POP 330 440, 28359 Bremen
Am Handelshafen 12, 27570 Bremerhaven
Josef-Engert-Str. 11, 93053 Regensburg
Josef-Engert-Str. 11, 93053 Regensburg
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
0421 218-65590
0471 4831-1307
0471 4831-1868
0471 4831-1639
0471 4831-1418
0471 4831-1312
0941 942 72 150 / 114
0941 942 72 150 / 114
0.4
Diepenbroek
Pörtner
Buck
Fisch
Krieten
Sartoris
Apostolidis
Huber
Meyerhöfer
1.1
Reusch
University of Münster
Hüfferstr. 1, 48149 Münster
[email protected]
0251 - 83 - 2 1095
Jost
Jürgens
Hippler
LaRoche
Müller
Reusch
Riebesell
Karsten
Hübener
Engel
Engel
Voß
Nausch, M.
Nausch, G.
Grossart
Thomsen
Gust
Schneider
Leibniz Institute for Baltic Sea Research (IOW), Warnemünde
University of Rostock
University of Rostock
Leibniz-Institute of Freshwater Ecology and Inland Fisheries (IGB), Berlin
Jacobs University Bremen gGmbH
Hamburg University of Technology (TUHH)
University of Kiel
Seestr. 15, 18119 Rostock
Hindenburgplatz 55, 48143 Münster
Hüfferstr. 1, 48149 Münster
Albert-Einstein-Str. 3, 18059 Rostock
Wismarsche Str. 8, 18051 Rostock
Alte Fischerhütte 2, OT Neuglobsow,16775 Stechlin
Campus Ring 1, 28759 Bremen
Schwarzenbergstraße 95 C, 21073 Hamburg
Ludewig-Meyn-Straße 10, 24118 Kiel
guenter.jost@io-warnemuende
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
0381 5197-270
0381 5197-250
0251 - 83 2 4790
0431 600-4212
0431 600-4526
0251 - 83 - 2 1095
0431 600-4444
0381 498 6090
0381 498 6210
0471 4831-1055
0471 4831-1055
0381 5197 209
0381 5197 227
0381 5197 332
033082 699 91
0421 200-3254
040 42878 6000
0431 880-3254
Brey
Am Alten Hafen 26, 27568 Bremerhaven
[email protected]
0471 4831-1300
Köhler
Bickmeyer
Niehoff
Lannig
Kurpromenade 201, 27498 Helgoland
Am Alten Hafen 26, 27568 Bremerhaven
[email protected]
[email protected]
[email protected]
[email protected]
0471 4831-1407
04725 819-3224
0471 4831-1371
0471 4831-2015
Mark
[email protected]
0471 4831-1015
Storch
Bridges
University of Düsseldorf
Universitätsstraße 1, 40225 Düsseldorf
[email protected]
[email protected]
0471 4831-1934
0211 81-14991
0.3.1
0.3.2
1.1.1
1.1.2
1.1.3
1.1.4
1.1.5
1.2
1.2.1
1.2.2
1.2.3
1.2.4
1.2.5
1.3
2.1
2.1.1
2.1.2
2.1.3
2.2
2.2.1
2.2.2
277
0431 600-4214
Appendix: Principle Investigators (continued)
Project
Name
Institute
Address
Email
Phone
2.3
Clemmesen
Piatkowski
Lucassen
Melzner
Schulz
Form
Melzner
Bleich
Tollrian
Riebesell
Tollrian
Richter
Fietzke
Bijma
Bijma
Eisenhauer
Böttcher
Böttcher
de Beer
Hoppema
Immenhauser
Immenhauser
Mutterlose
Meier
Kinkel
Wahl
R. Asmus
Wahl
Bischof
Ramette
Boetius
Boersma
Boersma
Rost
University of Kiel
Ruhr-University Bochum
Max Planck Institute for Marine Microbiology
University of Kiel
University of Kiel
University of Bremen
Hermann-Rodewald-Straße 5, 24118 Kiel
Universitätsstraße 150; 44801 Bochum
Universitätsstr.150, 44780 Bochum
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
[email protected]
0431 600 4558
0431 600-4571
0471 4831-1340
0431 600-4274
0431 600-4510
0431 600-1987
0431 600-4274
0431 880-2961
0234 32-14114
0431 600-4444
0234 32-24998
5.1
Pätsch
Institute of Oceanography, University of Hamburg
Bundesstr. 53, 20146 Hamburg
[email protected]
040 - 42838 6628
5.2
5.3
Oschlies
Quaas
University of Kiel
Wilhelm-Seelig-Platz 1, 24118 Kiel
[email protected]
[email protected]
0431 600-1936
0431 880 -3616
2.3.1
2.3.2
3.1
3.1.1
3.1.2
3.1.3
3.1.4
3.2
3.2.1
3.2.2
3.2.3
3.2.4
3.3
3.3.1
3.3.2
3.4
3.4.1
3.4.2
3.4.3
3.5
3.5.1
3.5.2
3.5.3
4.1
4.1.1
4.1.2
4.1.3
4.1.4
4.2
4.2.1
4.2.2
278
Columbusstrasse, 27568 Bremerhaven
Wischhofstraße 1-3, 24148 Kiel
Wischhofstraße 1-3, 24148 Kiel
Celsiusstr. 1, 28359 Bremen
Bussestrasse 24, 27570 Bremerhaven
Universitätsstraße 150, 44801 Bochum
Hafenstraße 43, 25992 List
Leobener Str., NW 2, 28359 Bremen
Kurpromenade, 27498 Helgoland
Kurpromenade, 27498 Helgoland
0471 4831-1304
0431 600-2106
0471 4831-1831
0471 4831-1831
0431 600-2282
0381 5197-402
0381 5197-402
0421 2028 - 802
0471 4831-1884
0234 32-28250
0234 32-28250
0234 32-23249
0431 880-2936
431 880-2878
0431 600-4577
04651 956-4308
0431 600-4577
0421 218-2859
0421 2028 - 863
0 421 2028 - 860
04725 819-3350
04725 819-3350
0471 4831-1809

bioacid

Transcription

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