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ACAROLOGIA
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FINE STRUCTURE OF THE CUTICLE
AND STRUCTURAL CHANGES OCCURRING DURING MOULTING
IN THE MITE TETRANYCHUS URTICAE
1. FINE STRUCTURE OF THE CUTICLE
BY
Ursula MOTHES-WAGNER* and Karl-August SElTZ*
CUTICLE
ULTRASTRUCTURE
MITE
TETRANYCHUS
ABSTRACT : Structure of the cuticle of Tetranychus urticae (Acari, Tetranychidae)
is described by means of electron microscopy.
The cuticle is composed of an epicuticle and a lamellated procuticle. The epicuticle exhibits an inn er 'dense layer' which shows a weak fibrillar appearence, a
cuticulin layer, a wax layer and an outer cement layer. The procuticle can be differentiated in an outer exocuticle which extends to the ridges and lobes, characteristic of the cuticular surface, and an inner lamellated endocuticle. Lamellae are
formed of helicoid arranged electron lucent microfibrils embedded in an electron
dense matrix. Pores penetrate ali cuticular layers.
Structure of the cuticle is compared to published information on the insect cuticle.
CUTICULA
ULTRASTRUKTUR
MILBEN
TETRANYCHUS
ZUSAMMENFASSUNG : Die Feinstruktur der Cuticula von Tetranychus urticae (Acari,
Tetranychidae) wird anhand von elektronenmikroskopischen Aufnahmen dargestellt.
Die Cuticula besteht aus einer Epicuticula und einer lamellierten Procuticula. Die
Epicuticula besitzt eine innere 'dense layer', die eine feine Streifung aufweist, eine
Cuticulinschicht, eine Wachsschicht und eine iiul3ere Zementschicht. Die Procuticula lal3t sich in eine Exocuticula, die bis in die Leisten und Loben hineinzieht, und
eine innere, lamellierte Endocuticula gliedern. Die Lamellen werden aus elektronenhellen Microfibrillen gebildet, die in eine elektronendichte Matrix eingebettet sind.
Poren durchziehen alle cuticularen Schichten.
Die Feinstruktur der Cuticula von Tetranychus urticae wird mit Kenntnissen an der
Insektencuticula verglichen.
1976; LOCKE, 1974; NEVILLE, 1975 ;
1951 ; WIGGLESWORTH, 1948).
The main layers are an outer nonchitinous epicuticle and an inner chitinous procuticle composed
mainly of protein and chitin. According to the
physical properties, the cuticle may be soft, tanned or sclerotized. Many parallels may be drawn
BURN,
INTRODUCTION
Structure of the arthropod cuticle, a multilayered extracellular material, has been investigated and summarized in numerous reviews (ANDERSEN, 1977, 1979 ; HACKMAN, 1971 ; HEP-
RICHARDS,
• Department of Zoology, Philipps University, Lahnberge D-3550 Marburg, Federal ·Republic of Germany.
Acarologia, t. XXV, fasc. 3, 1984.
254between insect and mite cuticles but interpreting
arthropod cuticle in terms of what is known of
the insect cuticle can be misleading. Especially,
the recent reviews of HACKMAN (1982) and HACKMAN and FILSHIE (1982) on tick cuticle show that
knowledge on the cuticle of the Acari is limited.
Structure of mite cuticle has been investigated
by ALBERTI et al. (1981) according to the taxonomie relationship of mite species. In addition, the
cuticle of the spider mite Tetranychus urticae has
been of special interest (ANW ARULLAH, 1963 ;
CROWE, 1975 ; HENNEBERRY et al., 1963, 1965 ;
GIBBS and MORRISON, 1965 ; MAYR, 1971)
because it is suggested that one factor of resistance of mites towards acaricides is related to special cuticular structures. Although such precise
information exists, severa! points are discussed
with sorne controversy.
The first part of the present paper describes the
ultrastructure of the cuticle of Tetranychus urticae
and compares it with previously published information on mite and insect cuticle.
MA TE RIALS AND METHODS
The animais were reared by following the
method described earlier (MOTHES and SElTZ,
1980). Entire animais were fixed in 4 OJo collidine
buffered glutaraldehyde followed by osmication
for 15 min. After dehydration in graded acetone
solutions, polymerization was effected in Spurr's
medium (SPURR, 1969). Ultrathin sections were
eut with a diamond knife on an LKB ultratome
and stained with KMnO. and lead citrate (REY-
NOLDS, 1963), using a method modified from
SOLOFF (1973).
OBSERVATIONS
In describing the fine structure of the cuticle,
the nomenclature used for the layers is that which
other authors have used (ANWARULLAH, 1963 ;
GIBBS and MORRISON, 1959 ; MAYR, 1971 ;
ALBERTI et al., 1981 ; HACKMAN and FILSHIE,
1982) and is valid also for the insect cuticle
(ANDERSEN, 1977; 1979; FILSHIE, 1982). The
spider mite cuticle has a thickness of 0.5-2.5 ,um.
Procuticle thickness varies between 0.25 ,um and
2 ,um in the different body regions. The cuticle bears typical surface differentiations such as
ridges and lobes (MOTHES-WAGNER, 1982) which
are of taxonomie interest, and is composed of an
epicuticle and a procuticle (Fig. 1). The latter
can be differentiated into an outer exo- and inner
endocuticle (Fig. 3 a). The epicuticle exhibits different layers (Fig. 2 a, b, arrows) : the inner layer
is a "dense layer " (MA YR, 1971) which is overlain by an electron dense cuticulin and electron
lucent wax layer. The outer layer is an often
ruptured cement layer (Fig. 2 a, b). Fibrillar
material observed in severa! tick epicuticles could
be clearly demonstrated in the setal cuticle of T.
urticae (Fig. 2 c) and, with sorne difficulties, in
the body epicuticle (Fig. 2 c). Between epi- and
endocuticle, the exocuticle with its mostly granular material extends to the ridges (Fig. 2 a).
Especially in males, the exocuticle is penetrated by
electron dense plaques (Fig. 1), the origin and
FIG. 1 : Cross section of the dorsal opisthosomal cuticle of male. The outer layer is an epicuticle (arrow) which forms the taxonomically relevant ridges (RI) and lobes (LO). Epicuticular material extends to the procuticle (X). The inner layer is a lamellated
procuticle (PR). Cuticular glands (double arrow) and pores (asterisk). Hypodermis (HY) with lipoproteid granules ( +) and
bacteria (B).
FtG. 2 a-c : Epicuticle. 2 a-b. -
The epicuticle of the dorsal opisthosome is composed of an outer cement layer (arrow), a wax
layer (double arrow), a cuticulin layer (arrow head) and an inn er 'dense layer' (DL). The exocuticle (EX) with mostly granular
material extends to the ridges (Rl) and lobes (LO). - 2 c. - The epicuticle of seta shows a fine striation.
FtG. 3 a, b : Cross section of lateral opisthosomal cuticle.
Below the epicuticle (EP) the granular exocuticle (EX) with fibrillar
deposits ( +) is situated, followed by an inner endocuticle (EN) with helicoïdal arranged lïbrils (arrow).
FtG. 4 a, b : Cuticular gland apparently functioning in wax secretion (arrow) has contact with mcmbranous structures of the hypodermis (x).
FiG. 5 : Soft cuticle of joint.
adjacent cuticle ( + ).
Epicuticle (EP) extremely electron dense.
Procuticle (PR) thin and electron lucent compared to
-255-
256function of which is unknown. The endocuticle
shows a conspicuous lamellation which is due to
the helicoïdal arrangement of electron lucent
fibrils in an electron dense matrix (Fig. 1, 3 a,
b). In different body regions, from four to sixteen such lamellae may be seen. In other regions
they are lacking (lateral prosoma, ventral
cuticle). The cuticle is interspersed with pore
canals and cuticular glands (Fig. 1, 4 a) which
secrete the wax layer (Fig. 4 b). The cuticle of
the soft joints shows conspicuous alterations.
The surface profile is knobbed (MOTHES-W AGNER, 1982) and the procuticle is less compact
(Fig. 5). Fibrils are easy to detect and may show
an altered arrangement.
DISCUSSION
Presented results on the structure of the cuticle
of Tetranychus urticae agree with those published
by other authors (ANWARULLAH, 1963 ; GIBBS
and MORRISON, 1965 ; HENNEBERRY et al., 1963,
1965; MAYR, 1971) and can be compared to
information on the insect (ANDERSEN, 1977 ;
1979; FILSHIE, 1982) and tick cuticles (HACKMAN and FILSHIE, 1982).
Epicuticular layers are described in a variety of
ways by the above authors, although it appears
that the epicuticle is of uniform structure over the
whole animal and is the same in larval and nymphal instars. Existence of a wax layer is universally acknowledged (MA YR, 1971 ; GIBBS and
MORRISON, 1965 ; BOSTANIAN et al., 1973 ;
ANWARULLAH, 1963 ; ALBERTI et al., 1981),
although the nomenclature may be different (see
ALBERTI et al., 1981). Findings on the cement
layer, "dense layer ", and the polyphenol-containing cuticulin layer are either discussed with
sorne differences (ALBERTI et al., 1981 ; MA YR,
1971), or have not been mentioned (ANWARULLAH, 1963 ; BOSTANIAN et al., 1973). GIBBS and
MORRISON (1965) reported a nonchitinous cuticulin layer, which later was refuted by means of
biochemical tests (MAYR, 1971). ALBERTI et al.
(1981) investigated the cuticles of 71 mite species
and described an epicuticle as weil as a procuticle
which rarely can be differentiated into the two
layers of exo- and endocuticle. The procuticle
showed mostly a pronounced lamellation of microfibrils while the epicuticle was formed of
homogenous material. The epicuticle was covered by a secreted layer of cement and wax and was
suggested to be responsible for the special surface
differentiations (ALBERTI et al., 1981).
According to the presented results on T. urticae, the epicuticle is composed of four layers :
cement, wax, cuticulin and "dense layer ". The
latter seems to be composed of lipoproteins
(MAYR, 1971) or proteins (HACKMAN and FILSHIE, 1982), according to its staining behaviour.
Determination of these materials, however, can
only be done on the basis of histochemical reactions. The epicuticle of ticks contains fibrillar
structures, none of which have been reported
from other arthropod cuticles (HACKMAN, 1982).
In Tetranychus urticae too, electron lucent microfibrils can be detected in the '' dense layer '' and
the setal cuticle which is continuous with that of
the body. Du ring preliminary investigations on
the chitin content of T. urticae, evidence has arisen that the epicuticle contains chitin (MOTHESWAGNER, unpubl.), a condition which is also described for the arachnid Palamneus swammerdami
(Scorpionidae) (KRISHNAN et al., 1955). ln the
absence of definitive histochemical evidence, however, the presence of nonstaining microfibrils in
the epicuticle, or elsewhere, cannot be taken as
proof of the presence of chitin (HACKMAN, 1982).
The procuticle of T. urticae consists of an outer
unlamellated exocuticle which is of unknown
structure (CROWE, 1975), and an inner lamellated
endocuticle. HENNEBERRY et al. (1965) reported
the presence of seven endocuticular lamellae, but
MOTHES-WAGNER (1982) found that the number
of lamellae varies with different body regions, and
with sex (four to six for the female opisthosoma,
and sixteen for the male opisthosoma). The exocuticle takes part in the formation of the cuticular
ridges and lobes which are characteristic surface
differentiations of tetranychids. Function of
these ridges is suggested to be in a stabilization of
the cuticle. GIBBS and MORRISON (1965) reported
257a capability of stretching following food ingestion
which is due to the ridges. ln this context,
ALBERTI et al. (1981) showed that the cuticie of
ticks is polymorphous compared to that of spider
mites. The capacity for stretching is based upon
special structures of the basal layers. A thickened procuticle and an increased number of wax
channels which are characteristic for stretchable
soft-cuticles (ALBERTI et al., 1981) were not
observed in T. urticae. Additionally, electron
microscopical differences in appearence and
heighth of the ridges are not obvious between fed
and unfed animais.
According to MAYR (1971), procuticular lamellation is due to a helicoïdal arrangement of chitin
fibrils embedded in a proteinaceous matrix, as
reported for insects (ANDERSEN, 1977, 1979 ;
LOCKE et al., 1975, 1980 ; NEVILLE, 1963 a,
b, 1965). However, conclusions on components
forming endocuticular lamellation based only on
electron microscopical pictures should not be
made without simultaneous histochemical tests.
Investigations on the insect egg shell (MAZUR et
al., 1982) showed that a helicoïdal arrangement of
fibrils can be formed only by proteins. Consequently, one should consider the possibility that
the fine-structurally demonstrable fibrils in the
epi- and endocuticle of T. urticae may be due to
chitin or protein. Although the present results
do not answer the question of chitin content and
distribution in the cuticular layers of the spider
mite, histopathological effects of the chitin
synthesis inhibitor complex, Nikkomycin, are
conspicuous (MOTHES, 1981 ; MOTHES and SElTZ,
1982 ; MOTHES-WAGNER, 1982). At the moment, such effects could only be interpreted
with the suggeston that the cuticle of T. urticae
contains chitin (MOTHES· and SElTZ, 1982). Fine
structural investigations on the moulting processes
in T. urticae which are presented in a later
paper (MOTHES-WAGNER and SElTZ, 1984) do not
contribute to a solution of this question. To
what extent proteins and chitin (if present) are
linked, and which additional components (inactive
chitin synthetase, polyphenols, lipids, etc.) occur
in the cuticle of tetranychids, should be cleared in
subsequent investigations. Studies performed in
insects can be used as model systems, although
the size of tetranychids (0.1-0.4 mm) may create
tech ni cal difficulties.
ACKNOWLEDGEMENTS
The authors wish
GA Y TRAXLER for
study was supported
schungsgemeinschaft
to express their gratitude to Miss
correcting the manuscript. The
by a grant from the Deutsche For(DFG Se 162/13).
REFERENCES
ALBERTI (G.), STORCH (V.), RENNER (H.), 1981. Über den feinstrukturellen Aufbau der Milbencuticula
(Acari, Arachnida). - Zoo!. Jb. Anat. 105 : 183-236.
ANDERSEN (S. 0.), 1977. - Arthropod cuticles : their
composition, properties and functions. Symp. Zoo!.
Soc. London 39 : 7-32.
ANDERSEN (S. 0.), 1979. - The biochemistry of insect
cuticles. Ann. Rev. EntomoL 24 : 29-62.
ANWARULLAH (M.), 1963. - Beitrage zur Morphologie
und Anatomie einiger Tetranychiden (Acari, Tetranychidae). - Z. angew. Zoo!. 50 : 385-426.
BOSTANIAN (N. J.), MORRISON (F. 0.), 1973. - Étude
histologique du tégument et de l'effect des divers acaricides chez le tetranyque a deux points. - Can. J.
Zoo!. 51 : 1065-1067.
CROWE (J. H.), 1975. - Studies on acarine cuticles :
III. Cuticular ridges in the citrus red mite. - Trans.
Am. Microsc Soc. 94 : 98-108.
FILSHIE (B. K.), 1982. - Fine structure of the cuticle
of insects and other arthropods. In : Insect ultrastructure, VoL L - Plenum Press, NY - London, 281312.
GIBBS (K. E.), MORRISON (M. 0.), 1965. -The cuticle
of the two-spotted spider mite Tetranychus te/arius.
Can. J. Zoo!. 37 : 634-637.
HACKMAN (R. H.), 1971. - The integument of arthropoda. In : Chemical Zoology, VoL 6 (FLORKIN, M.
and SCHEER, B. T., eds.), Acad. Press, NY, 1-62.
HACKMAN (R. H.), 1982. - Structure and function in
tick cuticle. - Ann. Rev. EntomoL 27 : 75-95.
HACKMAN (R. H.) and FILSHIE (B. K.), 1982. - The
tick cuticle. In : Physiology of ticks (OBENCHAIN, F.
D. and GALUN, R., eds.). - Perg. Press, OxfordNY- Toronto-Sydney-Paris-Frankfurt.
HENNEBERRY (T. ].), ADAMS (J. R.), CANTWELL
(G. E.), 1963. -Comparative electron microscopy of
the integument of organo-phosphate resistant and
258non-resistant two-spotted spider mite, Tetranychus
urticae. - Proc. 1" Int. Conf. Acarol., Fort Collins
Colorado.
HEPBURN (H. R.) and CHANDLER (H. D.), 1976. Mechanical hysteresis of insect cuticles. - J. lnsect
Physiol. 22 : 221-226.
KRISHAN (G.), RAMACHANDRAN (G. N.), SANTANAN
(M. S.), 1955. - Occurrence of chitin in the epicuticle of an arachnid Palamneus swammerdami. Nature
176 : 557-558.
LOCKE (M.), 1974. - The structure and formation of
the integument in insects. ln : The physiology of
insects (ROCKSTEIN, M., ed.). - Acad. Press, NY.
LOCKE (M.) and HUIE (P.), 1975. - Staining elastic
fibers in insect connective tissue. Tiss. Cel!. 4 : 601612.
LOCKE (M.) and HuiE (P.), 1980. - Ultrastructural
methods in cuticle research. In : Cuticle techniques in
arthropods. (MILLER, T. A., ed.). - Springer, NYHeidelberg-Berlin.
MAYR (L.), 1971. - Untersuchungen zur Feinstruktur
der Cuticula der Bohnenspinnmilbe Tetranychus urticae. - Z. angew. Zoo!. 58 : 271-278.
MAZUR (G. D.), REGlER (J. C.), KAFATOS (F. C.),
1982. - Order and defects in the silkmoth chorion, a
biological analogue of a cholesteric liquid crystal. In :
Insect ultrastructure (KING, R. C. and AKAI, H.,
eds.). - Plenum Press, NY -London.
MOTHES (U .), 1981. - Feinstrukturelle Veranderungen
am Integument von Tetranychus urticae Koch (Acari,
Tetranychidae) nach Nikkomycin-Behancllung (AMS
0896 Bayer Leverkusen). - Mitt. Dt. Ges. Allg.
Angew. Entomol. 2 : 172-179.
MOTHES-WAGNER (U.), 1982. - Feinstrukturelle Untersuchungen an der phytopathogenen Spinnmilbe
Tetranychus urticae (Acari. Tetranychidae) und zur
Wirkungsweise der in der Erprobung befindlichen
Nikkomycine ais Bekampfungsmittel. Diss. Uni v.
Marburg.
MOTHES (U.) and SElTZ (K. A.), 1980. - Licht- und
elektronenmikroskopische Untersuchungen zur Funktionsmorphologie von Tetranychus urticae (Acari,
Tetranychidae). 1. Exkretions systeme. - Zoo!. Jb.
Anat. 104 : 500-529.
MOTHES (U.) and SElTZ (K. A.), 1982. -Action of the
microbial metabolite and chitin synthesis inhibitor
Nikkomycin on the mite Tetranychus urticae ; an
electron microscopical study. - Pestic. Sei. 13 : 426441.
MOTHES-WAGNER (U.) and SElTZ (K. A.), 1984. Fine structure of the cuticle and structural changes
occuring during moulting in the mite Tetranychus
urticae. - Zoomorph 104 : 105-110.
NEVILLE (A. C.), 1963 a. - Daily growth in locust
rubberlike cuticle influenced by an external rhythm.
- J. Insect Physiol. 9 : 177-186.
NEVILLE (A. C.), 1963 b. - Growth and deposition of
resilin and chitin in locust rubberlike cuticle. - J.
Insect Physiol. 9 : 265-278.
NEVILLE (A. C.), 1975. - Biology of arthropod cuticle. - Springer, Berlin.
REYNOLDS (E. S.), 1963. - The use of lead citrate at
high pH as an electron opaque stain in electron
microscopy. - J. Cell. Biol. 17 : 208-217.
RICHARDS (A. G.), 1951. - The integument of arthropocls. - Univ. Minnesota Press, Minneanapolis.
SOLOFF (B. L.), 1973. - Buffered potassium permanganate-uranyl-acetate-lead citrate staing sequence for
ultrathin sections. - Stain. Techn. 48 : 159-165.
SPURR (A. R.), 1969. - A low viscosity epoxy resin
embedcling for electron microscopy. - ]. Ultrastr.
Res. 26 : 31-43.
WIGGLESWORTH (V. B.), 1948. -The insect cuticle. Biol. Rev. 23 : 408-451.
Paru en octobre 1984.