(Sheldon)Isolated From Common Bean (Phaseolus Vulgaris .L)Seeds

SOME STUDIES ON THE FUNGUS Fusarium moniliforme
(SHELDON)ISOLATED FROM COMMON BEAN (Phaseolus vulgaris
.L)SEEDS.
BY
Mutasim Mohamed Osman
B.Sc (Agric) University of Manofiya (Egypt)
1981
A thesis presented in partial fulfillment of the
requirements for the Degree of M.Sc (Agric)
(Plant Pathology)
Supervisor
Prof. Ahmed Mohamed Baghdadi
From the Department of Crop Protection
Faculty of Agriculture, University of Khartoum.
2005
DEDICATION
To the memory of my father….
to my mother,
to my wife Inshirah,
and my sons, Awwab and Mohamed
Osman,
with great love and respect
i
ACKNOWLEDGEMENTS
First of all I am greatly indebted to the merciful “ALLAH”
who offered me everything to accomplish this study.
I would like to express my sincere gratitude and thanks to my
supervisor Prof. Ahmed Mohamed Baghdadi for his close and
careful supervision , guidance, patience , encouragement and help
throughout this study.
Deepest thanks are extended to:
- My brother …Dr. El-Dirdiri Mohammed Osman for his
assistances, encouragement, and financial support.
- Staff of Plant Pathology Labratory, Plant Protection
Directory, Khartoum North- Sudan , Mrs; Awatif Mohamed
Ibrahim , Dr. Soheir Ahmed Abd El-wahab ,Miss Mashair
Ahmed Abd El-Hafiz for their cooperation assistances and
help in identification of the isolated fungi.
- Plant Protection Directory. Ministry of Agriculture &
Forestry for financing this study.
- All my friends and colleagues for cooperation and
encouragement.
ii
Contents
page
Dedication …………………………………………………………
i
Acknowledgment ………………………………………………….
ii
Contents……………………………………………………………
iii
List of tables ……………………………………………………….
iv
List of figures……………………………………………….………
v
List of plates…………………………………………………….….. vi
Abstract………………………………………………….………….. vii
CHAPTER ONE: INTRODUCTION………….......…………..…… 1
CHAPTER TWO REVIEW OF LITERATURE…..…………..…… 3
2-1: Seed-borne diseases associated with common bean……. 3
2-2: Seed –borne fungi associated with common bean……...
3
2-3: Common bean diseases caused by Fusarium spp…….... 5
2-4: The fungus Fusarium moniliforme (Sheldon)……….…
6
2-5: Diseases caused by Fusarium moniliforme………….… 7
2-6: Control………………………………………….………… .7
CHAPTER THREE: MATERIALS AND METHODS:…………….. 9
3-1: Common been seed sample supply…………..………… 9
3-2: Detection of seed- borne fungi……………………..…….. 9
3-2-1: Dry seed inspection…………………………………….. 9
3-2-2: Blotter test………………………………………..…… 9
3-2-3: Agar plate test…………………………………….….. 10
3-3: Identification……………………………………….…… 10
3-4: Effect of different media on linear growth of Fusarium
moniliforme………………………………………………..11
3-5: Effect of fungicides on growth of Fusarium
moniliforme………………………………………...….…11
CHAPTER FOUR: RESULTS: …………………………………….....13
4-1: Detection of seed-borne fungi……………………...… …13
4-2: Media test…………………………………………… …..14
4-3: The effect of fungicides on the growth of Fusarium
moniliforme……………………………………..…………14
CHAPTER FIVE: DISCUSSION ………………………………… ..28
REFERENCES ……………………………………………………..…30
APPENDICES………………………………………………………....35
ARABIC ABSTRACT.
iii
LIST OF TABLES
Table
page
Table (1): Common bean seed samples tested………….………15
Table (2): Dry inspection test…………………………………....15
Table (3): Percentage incidence of fungi recovered by blotter
method………………………………………………………..….16
Table (4): Percentage incidence of fungi on agar plate method…17
Table (5): Effect of media type on the linear growth of Fusarium
moniliforme………………………………………..…21
Table (6): Effect of Tilt on the linear growth of Fusarium
moniliforme……………………………………….…..23
Table (7): Effect of Bayleton on the linear growth of Fusarium
moniliforme…………………………………………...25
iv
LIST OF FIGURES
Figure
Fig: (1): Effect of media type on the linear growth of
page
Fusarium moniliforme………………...……………………………..…….23
Fig: (2): Effect of fungicides on the linear growth of Fusarium
moniliforme………………………………………… ….27
v
LIST OF PLATES
Plate (1) Chain of microconidia of the fungus
Fusarium moniliforme ………………..……………..18
Plate (2) Macroconidia of the fungus Fusarium
moniliforme……………………………………..………………..18
Plate (3): Culture of Fusarium moniliforme (Upper side)…...….19
Plate (4): Culture of Fusarium moniliforme (under side)………..20
Plate (5): Effect of media type on growth of Fusarium
moniliforme……….………………………………………22
Plate (6): Effect of Tilt on growth of Fusariu monmiliforme.…..24
Plate(7):
Effect of Bayleton on growth of Fusarium
moniliforme………….…………………… …26
vi
ABSTRACT
Four samples of common bean (PhasIeolus vulgaris ) were used in this
study , two samples from Hudieba Research Station, one sample from
Berber area and one sample from Shendi area. Samples were tested by dry
inspection, blotter test and agar plate method. The dry inspection revealed
malformation, discoloration and some impurities. In blotter method
Fusarium moniliforme isolated might be for the first time in Sudan. Also the
following fungi were detected : Fusarium oxysporum, Macrophomina
phaseolina, Alternaria alternata, Drechslera specifer, Cladosporium sp,
Curvulavria lunata, Aspergillus niger and Rhizopus sp.
Disinfection of seeds with sodium hypochlorite before plating showed that
Fusarium moniliforme was a seed-borne fungus
Czapek's Dox agar was found to be the best medium for the growth of the
fungus Fusarium moniliforme. In vitro the fungicides Tilt and Bayleton
were found effective in controlling the growth of the fungus Fusarium
moniliforme.
vii
CHAPTER ONE
INTRODUCTION
One of the most important food legumes in the tropical countries is common
bean. It is a major supplement of diet protein in tropical countries
(Owera,1990). There are different names beside the name common bean
such as kidney, haricot, garden, dwarf, snap and dry bean (Mmbaga et al.
1990). Common bean is consumed in dry and green forms and the straw can
be used as fodder. Like other legume crops it has the ability of nitrogen
fixation in the soil.
Common bean (Phaseolus vulgaris L.) belongs to the family Leguminosae
and to the sub- family Papilionidae. The origin of common bean is Latin
America and Mexico (Kaplan et al, 1973). The cool tropical regions are the
best area's for cultivating beans, but they can be grown in temperate areas
for their immature pods (Allen 1983).
There are about 26837 thousand hectars cultivated with common bean all
over the world. The annual world production of bean dry seeds is about 18.3
million tons.
About one third of this production is in the center of origin beans (Latin
America and North America). Brazil and Mexico grow about 82% of the
total production (FAO, 2002).
In Sudan common bean is cultivated mainly in the northern part of the
country. The major
production area in Sudan is Shendi – Berber area
.(Mohamed & Salih,1990).
It is also cultivated in the southern region around Katari, Juba, Loka West,
Yambio, Tonj and Wau (Tarr, 1955). In Sudan common bean is considered
as the second important food legume used by the majority of families. It
provide them with a large portion of their protein needs.
In Sudan common bean has been reported to be infected by several
important fungi, such as
Alternaria spp, Ascochyta phaseolorum,
Auerobasidium pullulans , Curvularia spp,Drechslera spp, Fusarium spp,
Macrophomina phaseolina , Myrothecium sp, Periconia byssoides ,
Phyllosticta sp , Pythium sp, Ulocladium atrum and Uromyces
appendiculatus . Many serious diseases were found to be caused by most of
these
fungi
in
the
country
(Boughey,1946;
Tarr,
1955:
Abdelwahab,1987,Salih et al., 1990 ). Several of these fungi are seed-borne
and the Fusarium spp. represent the most important fungi among them.
Many species of Fusarium had been isolated from bean seeds in Sudan.
In this study the isolation of Fusarium moniliforme from common bean
seeds might be the first record in Sudan according to the available literature.
2
Therefore this study was carried out to provide some information about this
fungus.
3
CHAPTER TWO
LITERATURE REVIEW
2-1 Seed-borne diseases associated with common bean:
According to Richardson (1968) the following diseases were reported on
common bean as seed- borne diseases:
A. Fungal diseases:
Ascochyta leaf spots, grey mould, pod rot, ashy stem blight, charcoal rot ,
angular leaf spot, damping off, stem canker, white mould, downy
mildew, Cladosporium spot , anthracnose , wilt and root rot.
B. Bacterial diseases :
Bacterial wilt, brown stem, grease spot, bacterial brown spot, common
bacterial blight, fuscous blight.
C. Viral diseases:
Bean mosaic virus , cowpea mild mottle virus,
2-2 Seed- borne fungi associated with common bean:
Several fungi had been found to be carried on or in the seeds of common
bean. Many investigators discussed the diseases caused by seed- borne
fungi in common bean for example, Ellis et al (1976) showed that many
4
fungi can be borne internally or as surface contaminants on common
bean seeds, Ellis et al .(1976) and Winter et al (1974) suggested that the
following fungi : Fusarium solani , F. oxysporum , F equiseti , F.
semitectum, F. moniliforme, Macrophomin phaseolina Phyllosticta
phaseolina , Aspergillus flavus , Drechslera tetramera, Phoma sp.
Colletotrichum Lindemuthianum and Botrytis cinerea were seed-borne
fungi associated with common bean seeds in various countries . Booth
(1971) isolated F.equisti from bean plants . Fulco et al. (1977) found that
Fusarium spp, Phytophthora sp, Rhizoctonia sp. Colletotrichum sp. and
Diporthe sp were the predominant seed- borne fungi in the seeds of three
bean cultivars tested by the blotter method. Gomes and Dhingra (1983)
found that Rhizoctonia solani and Alternaris alternata were seed-borne
fungi on common bean. In Sudan, AbdElwahab (1987) isolated the
following fungi from the seeds of common bean : Alternaria alternata ,
A. tenusissima, A. dianthi ,Auerobasidium pullulans, Cladosporium sp.,
Curvularia brachyspora , Fusarium solani, F. semitectum, Fusariella
intermedia , Fuariella aegyptiaca, Myrothecium sp , Cephalosporium sp,
Rhizopus sp and Ulodadium atrum . Eltayeb (1993)
isolated the
following fungi: Fusarium scripi , F. proliferatum , F. oxysporum, F.
equiseti, F. brachygibbosurm,
Alternaria alternata , Macrophomina
5
phaseolina, Drechslera specifer, Cladosporium herberum,Stemphylium
sp. , Curvularia lunata, Chaetomium globosum, Aspergillus flavus,
A.niger, Rhizopus sp., and Penicillium sp.
In Kenya Buruchara (1989) recorded the following fungi from 26
samples of bean seeds using the standard blotter method : Fusarium
moniliforme ,F. semitectum , F.equiseti , F. solani, F. oxysporum,
Colletotrichum
lindemuthianam,
C.
dematium,
Phoma
sp
,
Cephalosporium sp, Verticillum sp , Cercospra sp, Rhizoctonia solani,
Macrophomina phaseolina , Botryodiplodia theobromae , Rhizopus sp,
Penicillium sp Aspergillus sp, and Cladosprium spp.
Also other investigators reported about seed-borne fungi in other hosts
not including common bean. For example, Khan et al. (1974) in Pakistan
isolated Aspergillus spp, F. moniliforme and F. semitectum from the
seeds of chick pea (Cicer aritinum). Mebalds (1987) reported that Phoma
medicaginis , Fusarium acuminatum and Stemphylium sp, were found to
be transmitted from Medicago truncatula seeds to the plants.
2-3 Common bean diseases caused by Fusarium spp.
In various developmental stages common beans are exposed to many
pathogenic fungi Fusarium spp are considered as the most important
pathogenic fungi of common bean plants. However, Schwartz et al
6
.(1989) demonstrated that the infection with Fusarium spp may occur on
seedlings and mature plants. Kendrich and Snyder (1942) reported that
bean yellow disease caused by Fusarium oxysporum F,sp phaseoli was
first reported in California in 1929. Schwartz et al (1989) showed that
F.oxysporum caused wilt disease in commercial bean fields . Gupta and
Saharan (1973) asserted that F.solani caused pre- emergence rot when
soil inoculation was used before sowing and caused seedling blight when
seedling inoculation was performed.
Also Ykema and Stutz (1991) stated that the isolates of Fusarium
equiseti, F. oxysporum and F. solani might cause root rot symptoms.
2-4 The fungus Fusarium moniliforme (Sheldon) :
This fungus is well known and has been worldwide in its distribution.
It is a seed- borne fungus which is encountered in many host plants such
as :
Allium cepa, Avena sativa , Capsicum spp, Corchorus capsularis,
Glysine max, Gossypium spp., Hibiscus esculentus, Hordeum vulgaris,
Lycopersicon esculentum, Oryza sativa. Phaseolus spp , Pisum sativum,
Ricinus communis, Solanum melongena, Sorghum bicolor , Trifolium
hybridum, Triticum aestivum and Zea mays (Ram et al,.1970).
7
2-5 Diseases caused by Fusarium moniliforme:
Many diseases have been known to be caused by Fusarium moniliforme
in graminae, leguminosae, malvaceae solanaceae, musaceae and
tiliaceae. These diseases are seedling blight scorch foot rot, stunting and
hypertrophy (Booth 1971), This fungus was found to be the most
prevalent pathogen in maize causing ear rot disease (Koehler,1942).
However, there are now many reports which indicate that Fusarium
moniliforme is a serious pathogen in cereal crops; Segall et al, (1955)
reported that Fusarium moniliforme causes bud rot in Zea mays in Puerto
Rico;Tarr (1962) reported that this fungus causes root and stalk rot in
Africa; Futrell and Webster,(1967) showed that it causes seedling blight,
stalk rot, head blight and scabbed seed in Nigeria. In other crops the
fungus was found to be associated with Vicia fabae (Boughey, 1946),
Where as in other crops the fungus was found to be associated with
symptoms in Vicia Fabae (Boughey, 1946),where it causes stem rot and
wilt in Saccharum officinarum (Singh et al, 1975).
2-6 Control:
8
Much work in literature was done for the control of Fusarium
moniliforme in various crops such as rice, maize cotton , wheat, sorghum
and other crops and vegetables.
In the control of seed – borne pathogens certain measures are used, these
include use of resistant cultivars, enforcement of quarantine regulations,
seed treatment and production of disease free seeds.
Use of seed health testing in the control of seed-borne pathogens has
been documented and has recently been showed that treating rice seed
with Thiram reduced F. moniliforme effect on germination. According to
Neergaard (1977) seed treatment is a benefit practice in seed protection
to avoid seed loss.
Dey et al.(1989) showed that maize seeds treated with Bavistin gave
good control against F. moniliforme, resulting in best improvement and
seedling survival.
9
CHAPTER THREE
MATERIALS & METHODS
3-1 Common bean seed sample supply :
Four seed samples of common bean were collected from different
production areas in River Nile State (North Sudan). Two samples from
Hudeiba Research Station, one sample from Shendi area and the fourth
sample from Berber area (Table 1).
3-2 Detection of seed- borne fungi:
3-2-1 Dry seed inspection:
Four hundred seeds from each sample were taken at random and were
examined under the stereoscopic binocular microscope for impurities
such as plant debris, sclerotia, malformation and seed discolouration.
3-2-2 Blotter test :
(a) Un- treated seeds:
Four hundred seeds from each sample were tested by the standard
blotter method (ISTA, 1966). Each seed sample was plated onto three layers
of moistened blotters in sterilized Petri- dishes (9cm- diameter) at a rate of
10 seeds per dish equidistantly placed. The plates were then incubated at 28
10
o
C + 3 oC under alternating cycles of 12 hours near ultraviolet light and 12
hours darkness for 7 days (Sohieb, 1983). On the 8th day the seeds were
examined under the stereoscopic binocular microscope and the incidence of
seed –borne fungi was recorded. The compound microscope was also used
to confirm identification of different fungi.
(b). Chlorine pre- treated seeds:
Four hundred seeds from each sample were first soaked for 5 minutes
in sodium hypochlorite (1% available chlorine ) and then washed repeatedly
in several changes of sterile distilled water, then the seeds were plated and
incubated for 7 days, examined under the stereoscopic binocular microscope
and the incidence of the seed- borne fungi were recorded.
3-2-3 Agar plate test:
Two hundred seeds of each sample were disinfected with sodium
hypochlorite as mentioned before and then plated on sterilized Petri-dishes
containing sterilized Czapek's Dox agar medium at a rate of 5 seeds per dish.
Seeds were then incubated for 7 day at 28 oC and on the 8th day were
examined using the stereoscopic binocular microscope and the isolated seedborne fungi were recorded.
11
3-3 Identification :
The identification of the isolated fungi was based on: Booth, (1971),
Ellis (1971) and the help of staff of plant Pathology Laboratory –Plant
Protection Directory- Khartoum North- Sudan.
3-4 Effect of different media on linear growth of Fusarium moniliforme.
Three types of media were tested in this study. These were potato
dextrose agar (PDA), Czapek's Dox agar and malt extract agar (MEA).
From each medium four plates (9cm.diameter) were prepared. Two
diameters were drawn on the back of each plate for centering the inoculum.
The plates were then inoculated with 5 mm. disc from 7 days old culture of
the fungus Fusarium moniliforme . the plates were then incubated at 28oC
and the rate of the fungus growth was estimated daily by measuring the
colony size along the two diameters on the back of each plate , then the
mean of colony diameter was calculated for each medium.
3-5 Effect of fungicides on linear growth of Fusarium
moniliforme: -
12
Two fungicides were tested in this experiment ,Bayleton and Tilt.
Four dilutions for each chemical were used. The dilutions were prepared by
taking one gram (or one ml.) from the chemical and then dissolved in one
liter of sterilized distilled water to give 1000 ppm. From this dilution 1,3,5
and 7 ml. were added to four flasks containing 99. 97, 95 and 93ml of
sterilized Czapek's medium, respectively. This gave a final concentrations
of 10,30,50 and 70 ppm. for each chemical, respectively. Then the content of
each flask was poured in sterilized 4 Petri-dishes and two diameters were
drawn on the back of each plate for centering the inoculum. Each plate was
inoculated with 5mm disc from a 7 days old culture of Fusarium
moniliforme. The plates were then incubated at 28oC and the rate of the
fungus growth was estimated daily as mentioned before. Other eight Petridishes (four dishes for each replicate) containing sterilized Czapek's medium
were inoculated with the fungus to serve as control.
13
CHAPTER FOUR
RESULTS
4-1 Detection of seed – borne fungi
(a) In dry seeds:
Four hundred seeds from each sample were tested for dry inspection
test. The test showed that 20% of these samples were lighter in weight
and smaller
in size with yellow and brown to black discoloration.
Malformation of seeds was also observed ( Table 2).
(b) Blotter test (untreated seeds):
The following fungi were isolated on the 8th day after plating : Alternaria
alternata , Aspergillus niger, Cladosporium sp, Curvularia lunata,
Derchslera specifer, Fusarium moniliforme, Fusarium oxysporum ,
Macrophomina phaseolina and Rhizopus sp.( Table 3).
(c) Blotter test (treated seeds):
Seeds were treated with sodium hypochlorite (1% chlorine) before
germination on the blotter. The following fungi were observed:
Alternaria alternata , Cladosprium sp ,Curvularia lunata , Drechslera
14
specifer , Fusarium moniliforme, Fusarium oxysporum, M. phaseolina
(Table 3).
(d) Agar test (treated seeds) :
The following seed-borne fungi were detected in the agar plates:
Alternaria alternata , Curvularia lunata Drechslera specifer, Fusarium
moniliforme, Fusarium oxysporum and M. phaseolina. (Table 4)
4-2 Media test:
The fungus Fusarium moniliforme was grown on three types of media
Czapek's Dox agar was found to be the best medium for the growth of the
fungus . In Table 5 the effect of media type on the growth of the fungus
can be seen.
4-3 The effect of fungicides on the growth of the fungus
Fusarium moniliforme:
Two fungicides were examined in this test, Tilt and Bayleton. Different
concentrations were tested, 10ppm, 30ppm, 50ppm and 70 ppm from
each fungicides.
15
Tilt was found inhibiting the growth of the fungus at the low conc.
(10ppm) and was found completely inhibiting the growth at the conc, 70
ppm (Table 6)
Bayleton was found reducing the growth even at the lowest concentration
(10ppm) but the fungal growth was not inhibited even at the highest
concentration (70ppm) (Table 7)
Table (1)
Common bean seed samples tested
Variety
Locality
1. Ibaria cv
Hudeiba
Station
Hudeiba
Station
Berber area
Shendi area
2. Mutwakil cv
3. local cv
4. local cv
Year of Harvest
Research 2003-2004
Research 2003-2004
2003-2004
2003-2004
Table (2)
Dry inspection test of four samples of common bean tested
Sample
1. Ibaria cv
2. Mutwakil cv
Healthy
seeds
86.5%
77.25%
Seeds with
sclerotia
4%
6.5%
16
Malformed
seeds
3.5%
7.25%
Discoloured
seeds
6%
9%
3. local cv
(Berber area)
4. local cv.
(Shendi area )
77.5%
12.5%
4.5%
5.5%
78.75%
9%
5.75%
6.5%
Table (3)
Mean percentage incidence of fungi recovered by blotter method
(treated & untreated seeds) for four CV. tested
Fungi
% incidence in
untreated seeds
% incidence in
treated seeds
1. Alternaria alternata
8
5
2.Aspergillus spp
10
0
3.Cladosporium sp
12
4
4.Curvularia lunata
6
3
5.Drechslera specifer
7
3
6. Fusarium moniliforme*
12
3
17
7. Fusarium oxysporum
24
9
8. Macrophomina phaseolina
11
5
9.Rhizopus sp
6
0
* from Ibaria cv. only
18
Table (4)
Mean percentage incidence of fungi on agar plate method (treated
seeds) by 1% sodium hypochlorite for four cv. tested
Fungus
% incidence
. Alternaria alternate
5
2.Curvularia lunata
2
3.Drechslera specifer
3
4. Fusarium moniliforme
2
5. Fusarium oxysporum
8
6. Macrophomina phaseolina
4
19
Plate .1 chain of microconidia of the fungus Fusarium moniliforme
20
Plate .2 macroconidia of the fungus Fusarium moniliforme
21
22
23
Table (5)
The effect of 3 types of media on the linear growth of F. moniliforme
(c.m)
2nd
3rd
4th
5th
6th
Czapek’s 0.6
1.5
2.2
6.2
7.2
8.2
PDA
0.4
1.1
1.8
4.8
5.6
6.4
MEA
0.3
1.0
1.5
4.2
5.2
5.8
media
1st
days
24
25
10
1st
8
2nd
3rd
6
4th
4
5th
2
6th
0
MEA
PDA
Czapek's
Dox Agar
fig. 3 Effect of media type on linear growth of the fungus Fusarium
moniliforme
Table (6)
The effect of Tilt on the linear growth of F. moniliforme (cm) in vitro
1st
2nd
3rd
4th
5th
6th
Control
0.6
1.4
2.2
6.2
7.2
8.4
10 ppm
0.2
0.5
0.7
1.0
1.2
1.5
30ppm
0.1
0.3
0.4
0.6
0.8
1.0
50ppm
0.1
0.2
0.3
0.4
0.6
0.8
Dose
Days
26
70ppm
0
0
0
0
27
0
0
Table (7)
The effect of Bayleton on the linear growth of
F. moniliforme(cm)
1st
2nd
3rd
4th
5th
6th
control
0.6
1.4
2.2
6.2
7.2
8.4
10 ppm
0.5
1.0
1.5
4.0
4.8
6.7
30 ppm
0.1
0.3
0.7
2.3
2.7
3.0
50 ppm
0.1
0.2
0.5
1.7
2.1
2.4
70 ppm
0.1
0.2
0.5
1.6
2.0
2.2
Dosage
Days
28
29
30
9
8
7
1st
6
2nd
5
3rd
4
4th
3
5th
2
6th
1
0
Control
10 ppm
30ppm
50ppm
70ppm
Fig.4 Effect of Tilt on linear growth of the fungus Fusarium moniliforme
growth (cm)
effect of Bayleton on linear growth of
F.momiliforme
9
8
7
6
5
4
3
2
1
0
1st
2nd
3rd
4th
5th
6th
control
10 ppm
30 ppm
50 ppm
70 ppm
Fig.4 Effect of Bayleton on linear growth of the fungus Fusarium
moniliforme
31
CHAPTER FIVE
DISCUSSION
In this study seed health testing was carried out for four samples of common
bean according to ISTA (1966) rules. The standard methods used were dry
inspection test, blotter method and agar plate method. In dry inspection a
percentage from 12.5-20% of the seeds were found associated
with
sclerotia, discolouration and malformation.
In the blotter and agar methods, several seed-borne fungi were isolated from
untreated or chlorine pre- treated seeds. This result coincides with Neergaard
(1977) who recorded that blotter method was superior to the agar plate
method. Fusarium moniliforme
was isolated from the variety Ibaria of
Hudeiba Research Station. The isolation of Fusarium moniliforme from
common bean seeds is considered as the first record in Sudan as far as the
literature is available (Osman, 2005). The investigation and emphasis on this
fungus Fusarium moniliforme was due to the fact that it is the first record.
Fusarium oxysporum was encountered in all samples in high percentages
(9-24%) compared to the other fungal species and this result coincides with
Neergaard (1977) who reported that the blotter method is ideal for detection
32
of different fusaria. Some other fungi such as Alternaria alternata,
Macrophomina phaseolina , Drechslera specifer , Cladosporium sp.,
Curvularia lunata, Aspergillus sp, and Rhizopus sp. were also detected in
rather low percentages. Several investigators had reported similar seed-borne
fungi associated with common bean(Winter et al 1974; Ellis et al 1976;
Abdel Wahab,1987; Eltayeb. 1993).
The physiological study conducted was for the suitable medium for the
growth of the fungus Fusarium moniliforme. Three types of media were
tested in this study, PDA, MEA and Czapek’s Dox agar. The later medium
was found to be the suitable medium for the growth of the fungus. This
finding agreed with Agarwal and Singh (1974), who found that Czapek’s
medium was highly selective for the isolation of the fungus Fusarium
moniliforme.
The characteristic features of Fusarium moniliforme were identical to those
described by Booth (1971) who reported that Fusarium moniliforme has a
culture pigmentation of peach , salmon, vinaceous or purple to violet colour.
Microconidia fusoid to clavate 5-12× 1.5-2.5 µ. Occasionally becoming 1
septate and produced in chains. Macroconidia when present in some strains
are in equilaterally fusoid, thin walled,3-7 septate, 25-60× 2.4-4 µ.
Chlamydospores absent.
33
The control of Fusarium moniliforme by the fungicides Tilt and Bayleton in
vitro was found effective by both fungicides. This result coincides with the
finding of Sohieb (1983) who found that Benlate had a toxic effect to the
linear growth of Fusarium moniliforme.
REFERENCES
Abdel Wahab, S.A.(1987). A survey of seed –borne fungi of some
legumonus crops of the Sudan M.Sc. thesis, Univ. Of
Khartoum.
Agarwal, V.K and Singh, O.V(1974), Relative percentage incidence of
seed-borne fungi associated with different varieties of
rice seeds .
Seed Research, 2: 23-25
Allen, D.J.(1983). The Pathology of Tropical Food Legumes: Diseases
Resistance in Crop Improvement. Wiley, Chichester, England,
413p .
Booth,C.(1971). The genus Fusarium. CMI,Kew surrey, England,237p.
Boughey,A.S.(1946). Apreliminary list of plant diseases in the AngloEgyptian Sudan.Mycological papers No.14,C.M.I Kew,
Surrey England.
Buruchara.R,A (1989). Preliminary information on seed- borne fungi of
bean (Phaseolus vulgaris) in Kenya.CIAT(Intertional
Center for Tropical Agriculture) African Workshop,
series No.7
Dey, S.K,Gill,G.S.Aulakh,K.S,Soki,S.S. KHera,A.S and Grewal,P.K
(1989) effect of seed dressing chemicals on germination
and seedling vigour of maize. Plant Disease Reptr. 4:3642.
Ellis,M.A., Galves,G.E.and Simclair, J.B.(1976). Effect of pod contact
34
with soil on fungal infection of dry bean seeds .Plant
Disease Reptr.60: 974-976.
Ellis,M.A. and Paschal, E.H.(1979). Effect of fungicides seed treatment
on internally seed- borne fungi affecting germination and
field emergence of pigeon pea. Seed Sci, and Technol.
7:75-81.
Ellis,M.B. (1971). Demataceous Hyphomycetes. C.M.I., Kew,Surrey,
England.
Eltayeb, E.M.,(1993) Detection of seed borne fungi associated with
common bean ( Phesolus vulgaris ) in the Northern
region of Sudan. M.Sc Thesis, University of Khartoum.
FAO (Food & Agriculture Organization of United Nations) (2002).FAO
Production Year book.
Fulco,W.,Das, S.,Gamergo,M.R, DE,O.,Eltmyer,M.B.and Brusamoum,
R.(1977). Health testing of seeds of three bean cultivars
from two regions in Rio Grande. Rev. PL.Path.58- 2025.
Futrell,M.C. and Webster,O.J.(1967). Fusarium scab of sorghum in
Nigeria ,PL.DIS.Rertr. 51:174 -178.
Gomes,J.L. and Dhingra,O.D.(1983) Alternaria alternata aserious
pathogen of white coloured snap bean (Phaseolus
vulgaris L.) seeds, Rev. PL.Path.63:2588
Gupta,V.K.and Saharan,C.S.(1973) seed rot , root rot complex of beans
(Phaseolus vulgaris L.) seeds, Rev. PL.Path.52:3883.
ISTA (International Seed Testing Association )(1966) International rules
of Seed Testing .Proc.Int.Seed Test Ass/ 31:1-15.
Kaplan, L.Lynch, T.F.and Smith,C.E.(1973).Early cultivated beans from
an Intermontane Peruvian Valley. Science 179:76-77.
Kendrich,J.B.and Snyder,W.C.(1942).Fusarium yellows of beans
35
.Phytopathology 32:1010-1014.
Khan.S.A.J.Mathur, S.B.and Neergaard,P.(1974) Survey on seed –borne
organisms of Pakistan. Seed Sci.and Technol. 2: 427-429.
Koehler, B.(1942) Natural mode of enterance of fungi into corn ears and
some symptoms that indicate infection.JAgric,Res 14:
421-442.
Mathur, S.B(1983).testing seed of tropical species for seed-borne
diseases. Seed Sci and Technol, 11:113-128
Mebalds,M.I.(1987) . Effect of seed mycoflora on the development of
disease in stands of Medicago truncatula. Seed Sci. and
Technol. 15: 811-820.
Mmbaga , M.E.T,Koiange, E.M.K ,Gondive,B.Mbuye,O.S., Mushi,C.S
and Kamala,R.(1990). Bean production and research highlights
in the northern Tanzania,CIAT African workshop series No.7.
Mohamed ,A.K. and Salih,H.S. (1990) Evaluation of dry bean Phaseolus
vulgaris L.cultivars in Sudan . CIAT African workshop series
No.7.
Neergaard,P.(1977).Seed pathology.VOl.1. the Greshman press Old
wording Surrey ,England.
Osman,M.M. (2005). Some studies on the fungus Fusarium
moniliforme (Sheldon.) isolated from common bean
(Phaseolus vulgaris.L) seeds. M.Sc Thesis, University of
Khartoum.
Owera,E.(1990) .Analysis serotypes and strains of bean common mosaic
virus (BCMV) in countries with CIAT's regional programe
on beans in Eastern Africa . CIAT African workshop series
No.7.
Ram,N.,Mathur,S.B.and Neergaard,P.(1970) Identification of Fusarium
spp. as they occur in blotter test .Proceeding of
36
International Seed Testing Association ,Seed Pathology,
35:121-144.
Richardson,N.M.(1968).An Annotated List of Seed –borne Diseases.
Proceeding of International Seed testing Association.33:64-65.
Salih,H.S.Bushara,A,G.and Khalid, A.M.E.(1990).common bean
(Phaseolus vulgaris L.) Production and research in the
Sudan CIAT African workshop series No.7.
Schwartz, H.F.Mc Millan, M.S. and Silbernagel,M.J.(1989). Occurance
of Fusarium wilt in clorado. PL.Dis.Reptr. 73:518.
Segall,R.H. and Veles, F.J.(1955).Gibberella Fujikuroi, the cause of pod
rot of corn in Puerto Rico. PL.Dis.Reptr. 39:238.
Singh,R.S.and Singh,N.(1975). An observation on the association of
Fusarium moniliforme with sugar cane wilt Indian
Phytopathology,28:271-272.
Sohieb.A.Y.(1983). Detection of Fusarium moniliforme (Sheldon) in okra
seeds (Hibiscus esculentus) its significance and
control. M.Sc. thesis, Unive, of Khartoum.
Tarr,S.A.J.(1955). The Fungi and Plant Diseases of the Sudan. C.M.I .
Kew, Surrey, England .
Tarr,S.A.J(1962). Diseases of sorghum, Sudan grass and Broom corn
.C.M.I. Kew. Surrey. England.
Winter,W.E.,Mathur,S.B. and Neergaard,P.(1974). Seed-borne organisms
of Argentina. Plant disease Reptr. 58:507-511
Ykema, R.E. and Stutz, J.C.(1991). Isolation, identification and
Pathogenicity of Fusarium spp from guayule in Arizona.
Plant Disease Reptr. 75:736-738.
37
APPENDICES
38
CULTURE MEDIA
1- Potato Dextrose Agar (PDA) was made from the following
constituents:
Potato Extract
250 gm.
Dextrose
20 gm.
Agar
15 gm.
Distilled water
1000ml.
2- Czapek’s Dox Agar:
Dipotasium Phosphate
Sodium nitrate
Magnesium sulphate
Potassium chloride
Ferrous sulphate
Sucrose
Distilled water
1.0 gm.
0.5 gm.
0.5 gm.
0.5 gm.
0.1 gm.
20.0 gm.
1000ml.
3- Malt Extract Agar (MEA):
Malt Extract
Agar
Distilled water
30 gm.
20 gm.
1000ml.
39
‫ﻣﻠﺨﺺ اﻻﻃﺮوﺣﺔ‬
‫ﺍﺠﺭﻱ ﻫﺫﺍ ﺍﻟﺒﺤﺙ ﻋﻠﻲ ﺍﺭﺒﻌﺔ ﻋﻴﻨﺎﺕ ﻤﻥ ﺒﺫﻭﺭ ﺍﻟﻔﺎﺼﻭﻟﻴﺎ ﺍﻟﺠﺎﻓﺔ ﻟﻤﻌﺭﻓـﺔ ﺍﻟﻔﻁﺭﻴـﺎﺕ‬
‫ﺍﻟﻤﺤﻤﻭﻟﺔ ﻋﻠﻲ ﺍﻭ ﺩﺍﺨل ﺍﻟﺒﺫﻭﺭ ﺍﻟﻌﻴﻨﺎﺕ ﺍﻟﻤﺴﺘﺨﺩﻤﺔ ﻓﻲ ﺍﻟﺒﺤﺙ ﺍﺜﻨﺘﺎﻥ ﻤﻨﻬﺎ ﻤﻥ ﻤﺤﻁـﺔ‬
‫ﺍﺒﺤﺎﺙ ﺍﻟﺤﺩﻴﺒﺔ ﻭﻋﻴﻨﺔ ﻤﻥ ﻤﻨﻁﻘﺔ ﺒﺭﺒﺭ ﻭﺍﻟﻌﻴﻨﺔ ﺍﻻﺨﻴﺭﺓ ﻤﻥ ﻤﻨﻁﻘﺔ ﺸﻨﺩﻱ ﺒﻭﻻﻴﺔ ﻨﻬـﺭ‬
‫ﺍﻟﻨﻴل‬
‫ﺍﺠﺭﻴﺕ ﺍﻻﺨﺘﺒﺎﺭﺍﺕ ﺍﻟﻤﻭﺼﻲ ﺒﻬﺎ ﻟﻔﺤﺹ ﺍﻟﺒﺫﻭﺭ )‪ (ISTA 1966‬ﻭﻫﻲ ﺍﺨﺘﺒـﺎﺭ ﻭﺭﻕ‬
‫ﺍﻟﺘﺭﺸﻴﺢ ﻭﺍﺨﺘﺒﺎﺭ ﻁﺒﻕ ﺍﻻﺠﺎﺭ ﺍﻀﺎﻓﺔ ﺍﻟﻲ ﺍﻟﻔﺤﺹ ﺍﻟﺠﺎﻑ ﻟﻠﺒﺫﻭﺭ ‪.‬‬
‫ﺍﻟﻔﺤﺹ ﺍﻟﺠﺎﻑ ﻟﻠﺒﺫﻭﺭ ﺍﻭﻀﺢ ﺍﻥ ﻫﺫﻩ ﺍﻟﻌﻴﻨﺎﺕ ﺘﺤﺘﻭﻱ ﻋﻠﻲ ﺒـﺫﻭﺭ ﻤﻠﻭﻨـﺔ ﻭﺍﺨـﺭﻱ‬
‫ﻤﺸﻭﻫﺔ ﻤﻊ ﻭﺠﻭﺩ ﺒﺫﻭﺭ ﻤﻜﺴﻭﺭﺓ ﻁﺭﻴﻘﺔ ﻭﺭﻕ ﺍﻟﺘﺭﺸﻴﺢ ﻭﺒﻌﺩ ﻤﻌﺎﻤﻠﺔ ﺍﻟﺒﺫﻭﺭ ﺒﻬﻴﻜﻠﻭﺭﻴﺩ‬
‫ﺍﻟﺼﻭﺩﻴﻭﻡ ‪ (Sodium hypochlorite) %1‬ﺍﻅﻬﺭﺕ ﻭﺠﻭﺩ ﺍﻟﻔﻁﺭﻴﺎﺕ ﺍﻟﺘﺎﻟﻴﺔ ‪.‬‬
‫‪Alternaria alternata , Curvlaria lunata,Drechslera specifer.‬‬
‫‪Fusarium moniliforme F.oxysporium and Macrophomina‬‬
‫‪phaseolina‬‬
‫ﻋﺯل ﺍﻟﻔﻁﺭ ‪ Fusarium moniliforme‬ﺭﺒﻤﺎ ﻴﻌﺘﺒﺭ ﺍﻟﻤﺭﺓ ﺍﻻﻭﻟﻲ ﺍﻟﺘﻲ ﻴﺘﻡ ﻓﻴﻬﺎ ﻋﺯﻟﻪ‬
‫ﻤﻥ ﺒﺫﻭﺭ ﺍﻟﻔﺎﺼﻭﻟﻴﺎ ﻓﻲ ﺍﻟﺴﻭﺩﺍﻥ ‪.‬‬
‫ﺍﻭﻀﺤﺕ ﺍﻟﺩﺭﺍﺴﺔ ﺍﻥ ﺒﻴﺌـﺔ ‪ Czapek's Dox Agar‬ﻫـﻲ ﺍﻟﻤﺜﻠـﻲ ﻟﻨﻤـﻭ ﺍﻟﻔﻁـﺭ‬
‫‪Fusarium moniliforme‬‬
‫ﺍﻭﻀﺤﺕ ﺍﻟﺩﺭﺍﺴﺔ ﺍﻥ ﻤﻌﺎﻤﻠﺔ ﺍﻟﻔﻁﺭ ‪ Fusarium moniliforme‬ﻓﻲ ﺍﻟﻤﻌﻤل ﺒﻤﺒﻴـﺩﻱ‬
‫ﺍﻟﺒﺎﻴﻠﺘﻭﻥ ﻭﺍﻟﺘﻠﺕ ﻗﺩ ﺍﺩﻱ ﺍﻟﺤﺩ ﻤﻥ ﻨﻤﻭ ﺍﻟﻔﻁﺭ ﺤﺘﻲ ﻓﻲ ﺍﻟﺘﺭﻜﻴﺯ ﺍﻻﺩﻨﻲ ﻤـﻥ ﺍﻟﻤﺒﻴـﺩﻴﻥ‬
‫)‪ ( 10ppm‬ﺒﻴﻨﻤﺎ ﺘﻭﻗﻑ ﻨﻤﻭ ﺍﻟﻔﻁﺭ ﺘﻤﺎﻤﺎ ﻋﻨﺩ ﺘﺭﻜﻴﺭ ‪ 70ppm‬ﻤﻥ ﻤﺒﻴﺩ ﺍﻟﺘﻠﺕ‪.‬‬
‫‪40‬‬
‫ﺑﻌﺾ اﻟﺪراﺳﺎت ﻋﻠﻲ اﻟﻔﻄﺮ‪Fusarium moniliforme.‬‬
‫اﻟﻤﻌﺰول ﻣﻦ ﺑﺬور اﻟﻔﺎﺻﻮﻟﻴﺎ اﻟﺠﺎﻓﺔ‪.‬‬
‫ﺗﻘـــــﺪﻳﻢ‪:‬‬
‫ﻣﻌﺘﺼﻢ ﻣﺤﻤﺪ ﻋﺜﻤﺎن‬
‫ﺑﻜﻼرﻳﻮس اﻟﻌﻠﻮم ﻓﻲ اﻟﺰراﻋﺔ‬
‫ﺟﺎﻣﻌﺔ اﻟﻤﻨﻮﻓﻴﺔ )ﻣﺼﺮ ( ‪1981‬م‬
‫أﻃﺮوﺣﺔ أﻋﺪت ﻟﻨﻴﻞ درﺟﺔ اﻟﻤﺎﺟﺴﺘﻴﺮ‬
‫ﻓﻲ اﻟﻌﻠﻮم اﻟﺰراﻋﻴﺔ ) أﻣﺮاض اﻟﻨﺒﺎت(‬
‫اﻟﻤﺸﺮف‪ :‬ب ـ أﺣﻤﺪ ﻣﺤﻤﺪ ﺑﻐﺪادي‬
‫ﻣﻦ ﻗﺴﻢ وﻗﺎﻳﺔ اﻟﻤﺤﺎﺻﻴﻞ ) أﻣﺮاض اﻟﻨﺒﺎت(‬
‫آﻠﻴﺔ اﻟﺰراﻋﺔ‬
‫ﺟﺎﻣﻌﺔ اﻟﺨﺮﻃﻮم‬
‫ﻳﻮﻟﻴﻮ ‪2005‬م‬
‫‪41‬‬
42