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Sources and Mycoflora associated with some sundried vegetables in storage
International Biodeterioration & Biodegradation 31 (1993) 255-263
Sources and Mycoflora Associated with some Sundried Vegetables in Storage A. Adebanjo & E. Shopeju* Department of BiologicalSciences,Ogun State UniversityP.M.B. 2002, Ago-Iwoye, Nigeria
(Received26 August 1992; revisedversion received4 December 1992; accepted 28 December 1992)
ABSTRACT The sources and mycoflora of the following sundried, preserved vegetables." Abelmoschus esculentus, Corchorus olitorius, Solanum macrocarpon and Capsicum annum were investigated. The major fungalflora of the stored vegetables over a period of 8 weeks were Aspergillus flavus, A. niger, A. fumigatus, Rhizopus oryzae, Penicillium oxalicum, Rhizomucor pusillus and Fusarium equiseti however, the mycoflora isolated from the fresh vegetables and after sundrying differed. Some micro-organisms were isolated immediately after sundrying and throughout storage. There was also evidence offield acquisition of some of these species which were carried into storage. Also, post-harvest contaminants, handling, air spora during sundrying and storage fungi were other sources of contamination of these vegetables.
INTRODUCTION Aflatoxin-producing fungi, especially Aspergillus flavus is responsible for about 50% of fungal colonies isolated from some horticultural products (Kuku, 1980). Similarly, A. flavus-contaminated melon seeds have also been reported to produce aflatoxin Bl and G1 (Ogunsanwo et al., 1989). As a preliminary step towards establishment of the presence of aflatoxin in *Present address: Cadbury Nigeria PLC, Ikeja, Lagos, Nigeria. 255
256
A. Adebanjo, E. Shopeju
some sundried preserved vegetables, especially during storage, it is necessary to determine the source and type of mycoflora of the vegetables that are so preserved. This is necessary because, in Nigeria, vegetables are abundant in the rainy season but scarce in the dry season. This necessitates preservation, normally by sundrying, especially among the large rural populace, in order to provide vegetables during the dry season (Ihekornje & Ngoddy, 1985). However the literature available is mainly concerned with field disease problems (Joaquim, 1976; Odebunmi-Osikanlu, 1977; Ikediugwu, 1981; Arinze, 1987). Up until now, there appear to have been no reports on the micro-organisms associated with sundried stored vegetables, despite the fact that this is an age-long traditional preservation method. The present work has been directed towards the sources and mycoflora of some sundried preserved vegetables in order to rectify this apparent omission.
MATERIALS A N D METHODS Samples of four freshly harvested fruits and leaf vegetables: Capsicum annum (Sombo), Solanum macrocarpon (Igbagba), Corchorus olitorius (Jute) and Abelmoschus esculentus L. Moench (Lady's finger) were collected between December 1989 and March 1990 at three market locations in Ijebu-Igbo, Ibadan and the College of Agriculture Teaching and Research Farm, Ago-Iwoye, Nigeria. The moisture contents of the vegetables were determined by oven-drying 100-g edible portions of each until a constant weight was obtained.
Isolation of organisms associated with the fresh vegetables All the fresh vegetables were chopped and approximately 0.5 cm 2 samples were plated directly on potato dextrose agar (PDA). Plates were incubated at 28-4-2°C for 5 days and the organisms growing on the plate were examined and recorded. The percentage or frequency of isolation of organisms from each vegetable were also recorded. The organisms were subcultured on PDA until pure cultures were established. Identification was carried out by observation under the microscope and reference to Barnett (1967). The organisms were also sent to the International Mycological Institute (IMI) of the Commonwealth Agricultural Bureau (CAB) London for confirmation.
Mycoflora of sundried vegetables
257
Isolation of organisms associated with the sundried vegetables The four fresh vegetables mentioned above were chopped into pieces with a sharp sterile knife in sterile metal trays and sundried from 9.00a.m.6.00p.m., daily for 5 days. Sundrying of each vegetable was done in separate trays placed on raised platforms outside the laboratory to prevent interference by domestic animals and lizards. C. olitorius and S. macrocarpon were pulverised by hand wearing sterile rubber gloves. Okra and pepper were prepared using a Waring blender. Approximately 0.05 g of each were plated at four places in each plate, in triplicate, using the direct plate method. After 120 hours incubation at 28 -4- 2°C, the Petri dishes were examined for growth of micro-organisms. The frequency of isolation was again recorded and the organisms subcultured on PDA until pure cultures were obtained. Identification and confirmation of the species isolated were carried out as described above.
Isolation of organisms associated with sundried stored vegetables After sundrying for 5 days, the vegetables were removed to the laboratory and allowed to cool for 3 h. With a sterile spatula, each vegetable, in the pulverised form, was packed into a sterile test-tube bearing a cotton wool plug and covered with aluminium foil. The test-tubes were stored at 28 4- 2°C and isolations were made on PDA by the direct plate method on a weekly basis for 8 weeks. RESULTS The moisture content of the vegetables is shown in Table 1. TABLE 1 Moisture Content* of Dry Corchorus olitorius, Abelmoschus esculentus, Capsicum annum and Solanum macrocarpon Before Storage
Vegetable
Mean moisture content (%)
Abelmoschus esculentus Corchorus olitorius Solanum macrocarpon Capsicum annum *Values are means of three replicates.
10.0 :t: O. 12 10.4 -4- 0.05 11.0 -4- 0.36 8.5 -4- 0.35
A. Adebanjo, E. Shopeju
258
Organisms associated with the fresh vegetables Aspergillus niger, A. flavus, Rhizopus oryzae, R. stolonifer, Fusarium solani and F. equiseti were isolated from all the fresh vegetables except S. macrocarpon. Cercospora corchori and C. abelmoschi were obtained from jute leaves and okra fruits, respectively (Table 2). Sclerotium rolfsii was obtained from C. olitorius and S. macrocarpon. Choanophora cucurbitarum was isolated only from fresh okra and Colletotrichum capsici from fresh pepper. Aspergillus flavus was isolated from all the vegetables except Capsicum annum. Alternaria solani was obtained from 'igbagba' only (Table 2). Organisms associated with sundried vegetables After sundrying for 5 days, practically the same number and species of mycoflora isolated from the fresh vegetables were, isolated although at TABLE 2 Fungi Isolated From Fresh Corchorus olitorius, Abelmoschus esculentus, Capsicum annum and Solanum macrocarpon
Vegetable Corchorus olitorius
Capsicum annum
Abelmoschus esculentus
Solanum macrocarpon
Fungi isolated Cercospora corchori Rhizopus oryzae Fusarium solani Aspergillus niger Sclerotium rolfsii A spergillus fla vus Aspergillus niger Rhizopus stolonifer Colletotrichum capsici Fusarium equiseti
% isolation* 13.3 6.6 26.6 6.6 6.6
6.6 6.6 6.6 13.3
13.3
Rhizopus oryzae Aspergillus niger Cercospora abelmoschi Choanephora cucurbitarum Fusarium equiseti Aspergillus flavus
13.3 13.3 6.6
Alternaria solani Rhizopus oryzae Sclerotium rolfsii Aspergillus flavus
26-6 13.3 6.6 6-6
6.6 6.6 6.6
*The figures represent the frequency of isolation of each fungus from plated pieces/samples of each vegetable. Values are means of three replicates.
259
Mycoflora of sundried vegetables TABLE 3 Fungi Isolated From Corchorus olitorius, Capsicum annum, Abelmoschus esculentus and Solanum macrocarpon After Sundrying for 5 Days
Vegetable Abelmoschus esculentus
Fungi isolated
% isolation*
Aspergillus niger Rhizopus oryzae Rhizomucor pusillus Fusarium equiseti Aspergillus flavus
26-6 26-6
Corchorus olitorius
Aspergillus niger Aspergillus flavus Rhizopus oryzae Penicillium claviforme
13.3 26.6 33.3 13-3
Solanum macrocarpon
Aspergillus niger Aspergillus flavus Rhizopus oryzae Penicillium digitatum
13.3
Capsicum annum
Aspergillus niger Aspergillus fumigatus Rhizopus oryzae
13.3 33.3
13.3
13.3 33.3
13.3 13.3 13.3 33-3
*The figures represent the frequency of isolation of each fungus from plated pieces/samples of each vegetable. Values are means of three replicate samples.
higher percentages. However, S. rolfsii was not isolated from all the sundried vegetables while Penicillium claviforme and P. digitatum were obtained from jute and 'igbagba', respectively (Table 3). Organisms associated with sundried stored vegetables The number and kinds of mycoflora isolated from Abelmoschus esculentus was almost constant throughout the storage period except for the 3rd-5th weeks during which fewer and different fungi were obtained (Table 4). C. olitorius and S. macrocarpon had fewer species and number ofmycoflora in the first 3 weeks of storage and these increased till the end of the experimental period. C. annum was host to only three organisms over the whole period of storage (Table 4). The number of fungi isolated from the preserved vegetables was greater than from fresh vegetables (Tables 2 and 4). The percentage microbial occurrence in A. esculentus was 60-83% within the first 2 weeks of storage rising to 99% by the third week, declining to 66% in the fourth week and peaking at 83% by the eighth week (Table 5).
260
A. Adebanjo, E. Shopeju TABLE 4 Fungi* Isolated from Four Sundried Stored Vegetables Over an 8-week Period Vegetable
Abelmoschusescu&ntus
Corchorusolitorius
Weeks 1
2
3
4
5
6
7
8
An Af
An Af
An Aft
An Aft
An Aft
Ro
Ro
Ro
Ro
Ro
Po Rp
Po Rp
Mp --
An Af Af Ro Rp
An Af
Ro
An Aft Af Ro Rp
Po Rp
Po Rp
Po Rp
An Aft
An Af Ro Fe Po
An Aft Ro Po --
An Aft Af Ro Fe
An Aft Af Ro Fe
An Aft Af Ro Fe
An Aft Af Ro Fe
An Aft Af Ro Fe
Ro
Fe Po
Po
Po
Po
Po
Po
--
--
--
Rp
Rp
Rp
Rp
Rp
An Aft Af Fe
An Af Aft Ro Fe
An Aft Af Ro Fe
An Aft Ro Fe
An Aft Ro Fe
Ro
An Aft Af Ro Fe
Po
Po
An Aft Af Ro Fe
An Aft Af Ro Fe
Po
--
Po
Po
--
Po
Po
Rp
.
An Af Ro
An Af Ro
An Af Ro
An Af
-
So&num macroca~on
-
Capsicum annum
-
-
An Af Ro
-
-
-
-
An Af Ro
-
-
.
-.
An Af Ro
.
An Af Ro
.
Ro
*An, Aspergillus niger; Af, Aspergillus fumigatus," Aft, Aspergillus flavus; Ro, Rhizopus oryzae; Po, Penicillium oxalicum," Rp, Rhizomucor pusillus; Fe, Fusarium equiseti. C. olitorius h a d 5 5 % m i c r o b i a l l o a d in the first 2 weeks, w h i c h increased to 6 0 % b y week 4 a n d to 7 5 % b y week 8 ( T a b l e 5). S. macrocarpon h a d 5 3 % m i c r o b i a l invasion within 2 weeks o f storage, 5 0 % b y week 4 a n d 6 0 % b y the 8th week. C. annum h a d 8 3 % m i c r o b e s within the first 4 weeks o f storage a n d 7 5 - 8 3 % in weeks 6-8 ( T a b l e 5). G e n e r a l l y , m i c r o b i a l o c c u r r e n c e was highest in A. esculentus a n d C. annum o v e r the p r e s e r v a t i o n period. F o r all the s t o r e d vegetables, the highest p e r c e n t a g e m i c r o b i a l o c c u r r e n c e was r e c o r d e d within the first 3 weeks ( T a b l e 4).
DISCUSSION Fusarium equiseti was isolated f r o m fresh o k r a fruits in this s t u d y a l t h o u g h F. solani was earlier r e p o r t e d to cause fruit rot, in the field, b y J o a q u i m
60-0+0-0 50.0+0-0 50.0 + 0.0 60-04-0.2
1 83.3+0.1 55.5+0.0 53.3 ± 0.1 83.8±0-1
2 99.3+0.1 47-54-0.0 57.1 ± 0.0 99-9+0.0
3 66.6±0.0 60-0±0-1 50.0 + 0.0 83.3 ± 0.1
4
5 66-6±0.1 42.8±0.1 47.5 4- 0.1 75.0±0.0
Week
75.0±0.0 60.0±0.2 55.5 4- 0.1 83-34-0-1
6
83.3±0.1 66.6+0,0 57.1 ± 0.0 83-3-4-0-1
7
83.2 ± 0-1 75-0±0.0 60-0 ± 0.0 83.3±0-1
8
*Some of the fungi most commonly isolated from the sundried stored vegetables were Aspergillusflavus, Aspergillusfumigatus, Aspergillus niger, Penicillium oxalicum, Rhizopus oryzae, Rhizomucor pusillus, Fusarium equiseti. All values are +standard error.
A. esculentus C. olitorius S. macrocarpon C. annum
Vegetable
TABLE 5 Mean Percentage Microbial* Occurrence in Four Sundried Vegetables Over 8 Weeks of Storage at 28 ± 2°C
bo O~ m
~"
~. ~"
262
A. Adebanjo, E. Shopeju
(1976) and Odebunmi-Osikanlu (1977). This phenomenon could be due to location, and agroecological differences in the sources of the vegetable, apart from the dynamic nature of the pathogens. Cercospora corchori and C. abelmoschi isolated from fresh jute and okra, respectively, in this study were also reported to cause leaf spot and leaf mould on these vegetables in the field (Joaquim, 1976; Odebunmi-Osikanlu, 1977). Besides, C. corchori was found to be seed-borne in Corchorus olitorius (Joaquim, 1976). Similarly, Sclerotium rolfsii isolated from C. olitorius in this study had also been reported as a field disease of the crop (Odebunmi-Osikanlu, 1977). The isolation of Choanephora cucurbitarum from fresh okra fruits is in agreement with Odebunmi-Osikanlu (1977) and Adebanjo (1985). Aspergillusflavus isolated from the majority of the fresh vegetables clearly indicates the significance, in the air, of this species. It is reasonable to conclude that most of the fungi isolated from these fresh vegetables were pathogens acquired in the field. Isolation of almost the same number and species of mycoflora from both fresh and sundried vegetables means that some of these fungi were actually carried into storage and were capable of surviving during the 5-days sundrying. Apart from this, Rhizopus sp., Aspergillus niger, A. fumigatus have been reported as post-harvest pathogens of pepper in Nigeria (Adisa, 1985; Arinze, 1987). Mucor pusillus, Penicillium digitatum and A. flavus isolated from the vegetables after sundrying were probably part of the air spora. An increase in the number of fungi isolated from the preserved vegetables over time indicated their importance as storage fungi. These included Fusarium equiseti found in warm climates on pepper (Uma, 1981), A. fumigatus, and A. niger on Asparagus (Seenappa et al., 1980). The microbial occurrence on the preserved vegetables was highest within the first 3 weeks of storage because there was an abundance of fresh vegetable material on which the organisms could grow. The decline recorded in the number of microbes present at the later part of the storage ,period was probably an indication of exhaustion of the nutrients they contained. Peaks of microbial occurrence, towards the end of the storage period, for some of the vegetables could perhaps be an indication of increased microbial load resulting from cell multiplication/growth. The relatively thick slices of okra fruit compared with the thin flat-leaved vegetables used for this study accounted for their higher microbial loading. The present study clearly indicates the presence of A. flavus and other Aspergillus spp. on the vegetables at harvest, during sundrying and in storage. Since some of these organisms have been shown to produce aflatoxins in other horticultural products, this aspect requires further investigation with a view to highlighting the possible health hazards it might pose.
Mycoflora of sundried vegetables
263
REFERENCES Adebanjo, A. (1985). Premature fruit abortion, a new disease of okra in Nigeria caused by Choanephora cucurbitarum. J. Plant Prof. Tropics, 2, 131-3. Adisa, V. A. (1985). Microorganisms associated with the spoilage of Capsicum annum and C. frutescens in Nigeria. Fitopatologia Brasileira, 10, 427-32. Arinze, A. E. (1987). Microorganisms associated with harvested pepper fruits in Southern Nigeria. Fitopatologia Brasiler, 12, 91-3. Barnett, H. L. (1967). Illustrated Genera of Imperfect Fungi. 2nd edn, Burgess Publishing Company, Mineapolis, USA. Ihekoronje, A. I. & Ngoddy, D. O. (1985). Integrated Food Science and Technology for the Tropics, Macmillan Publishers Ltd. pp. 115-39. Ikediugwu, F. E. O. (1981). A shoot disease of Amaranthus spp. in Nigeria associated with Choanephora cucurbitarum. J. Hort. Sci., 56, 289-93. Joaquim, M. (1976). Seed-borne mycoflora of Nigerian local vegetables and their importance in disease development. PhD Thesis, University of Ibadan, Nigeria, pp. 21-42. Kuku, F. O. (1980). Technical Report, National Institute of Stored Products Research, Ibadan, Nigeria, 7, 63-68. Odebunmi-Osikanlu, Y. O. K. (1977). The more important diseases of selected local vegetables in Nigeria. Nigerian Journal of Plant Protection, 3, 79-83. Ogunsanwo, B. M., Faboya, O. O., Idowu, O. R., Ikotun, T. & Akano, D. A. (1989). The fate of aflatoxins during the production of 'Ogiri' a West African fermented melon seed condiment from artificially contaminated seeds. Die Nfihrung, 33, 983-8. Seenappa, M. L., Stobbs, W. & Krayah, G. (1980). Aspergillus colonization of kinds of pepper during storage. Phytopathology, 70: 218-22. Uma, Ngwanna, U., (1981). Four post-harvest diseases of the Nigerian red pepper (Capsicum annum L.). Plant Disease, 65, 915-16.
Sources and Mycoflora Associated with some Sundried Vegetables in Storage A. Adebanjo & E. Shopeju* Department of BiologicalSciences,Ogun State UniversityP.M.B. 2002, Ago-Iwoye, Nigeria
(Received26 August 1992; revisedversion received4 December 1992; accepted 28 December 1992)
ABSTRACT The sources and mycoflora of the following sundried, preserved vegetables." Abelmoschus esculentus, Corchorus olitorius, Solanum macrocarpon and Capsicum annum were investigated. The major fungalflora of the stored vegetables over a period of 8 weeks were Aspergillus flavus, A. niger, A. fumigatus, Rhizopus oryzae, Penicillium oxalicum, Rhizomucor pusillus and Fusarium equiseti however, the mycoflora isolated from the fresh vegetables and after sundrying differed. Some micro-organisms were isolated immediately after sundrying and throughout storage. There was also evidence offield acquisition of some of these species which were carried into storage. Also, post-harvest contaminants, handling, air spora during sundrying and storage fungi were other sources of contamination of these vegetables.
INTRODUCTION Aflatoxin-producing fungi, especially Aspergillus flavus is responsible for about 50% of fungal colonies isolated from some horticultural products (Kuku, 1980). Similarly, A. flavus-contaminated melon seeds have also been reported to produce aflatoxin Bl and G1 (Ogunsanwo et al., 1989). As a preliminary step towards establishment of the presence of aflatoxin in *Present address: Cadbury Nigeria PLC, Ikeja, Lagos, Nigeria. 255
256
A. Adebanjo, E. Shopeju
some sundried preserved vegetables, especially during storage, it is necessary to determine the source and type of mycoflora of the vegetables that are so preserved. This is necessary because, in Nigeria, vegetables are abundant in the rainy season but scarce in the dry season. This necessitates preservation, normally by sundrying, especially among the large rural populace, in order to provide vegetables during the dry season (Ihekornje & Ngoddy, 1985). However the literature available is mainly concerned with field disease problems (Joaquim, 1976; Odebunmi-Osikanlu, 1977; Ikediugwu, 1981; Arinze, 1987). Up until now, there appear to have been no reports on the micro-organisms associated with sundried stored vegetables, despite the fact that this is an age-long traditional preservation method. The present work has been directed towards the sources and mycoflora of some sundried preserved vegetables in order to rectify this apparent omission.
MATERIALS A N D METHODS Samples of four freshly harvested fruits and leaf vegetables: Capsicum annum (Sombo), Solanum macrocarpon (Igbagba), Corchorus olitorius (Jute) and Abelmoschus esculentus L. Moench (Lady's finger) were collected between December 1989 and March 1990 at three market locations in Ijebu-Igbo, Ibadan and the College of Agriculture Teaching and Research Farm, Ago-Iwoye, Nigeria. The moisture contents of the vegetables were determined by oven-drying 100-g edible portions of each until a constant weight was obtained.
Isolation of organisms associated with the fresh vegetables All the fresh vegetables were chopped and approximately 0.5 cm 2 samples were plated directly on potato dextrose agar (PDA). Plates were incubated at 28-4-2°C for 5 days and the organisms growing on the plate were examined and recorded. The percentage or frequency of isolation of organisms from each vegetable were also recorded. The organisms were subcultured on PDA until pure cultures were established. Identification was carried out by observation under the microscope and reference to Barnett (1967). The organisms were also sent to the International Mycological Institute (IMI) of the Commonwealth Agricultural Bureau (CAB) London for confirmation.
Mycoflora of sundried vegetables
257
Isolation of organisms associated with the sundried vegetables The four fresh vegetables mentioned above were chopped into pieces with a sharp sterile knife in sterile metal trays and sundried from 9.00a.m.6.00p.m., daily for 5 days. Sundrying of each vegetable was done in separate trays placed on raised platforms outside the laboratory to prevent interference by domestic animals and lizards. C. olitorius and S. macrocarpon were pulverised by hand wearing sterile rubber gloves. Okra and pepper were prepared using a Waring blender. Approximately 0.05 g of each were plated at four places in each plate, in triplicate, using the direct plate method. After 120 hours incubation at 28 -4- 2°C, the Petri dishes were examined for growth of micro-organisms. The frequency of isolation was again recorded and the organisms subcultured on PDA until pure cultures were obtained. Identification and confirmation of the species isolated were carried out as described above.
Isolation of organisms associated with sundried stored vegetables After sundrying for 5 days, the vegetables were removed to the laboratory and allowed to cool for 3 h. With a sterile spatula, each vegetable, in the pulverised form, was packed into a sterile test-tube bearing a cotton wool plug and covered with aluminium foil. The test-tubes were stored at 28 4- 2°C and isolations were made on PDA by the direct plate method on a weekly basis for 8 weeks. RESULTS The moisture content of the vegetables is shown in Table 1. TABLE 1 Moisture Content* of Dry Corchorus olitorius, Abelmoschus esculentus, Capsicum annum and Solanum macrocarpon Before Storage
Vegetable
Mean moisture content (%)
Abelmoschus esculentus Corchorus olitorius Solanum macrocarpon Capsicum annum *Values are means of three replicates.
10.0 :t: O. 12 10.4 -4- 0.05 11.0 -4- 0.36 8.5 -4- 0.35
A. Adebanjo, E. Shopeju
258
Organisms associated with the fresh vegetables Aspergillus niger, A. flavus, Rhizopus oryzae, R. stolonifer, Fusarium solani and F. equiseti were isolated from all the fresh vegetables except S. macrocarpon. Cercospora corchori and C. abelmoschi were obtained from jute leaves and okra fruits, respectively (Table 2). Sclerotium rolfsii was obtained from C. olitorius and S. macrocarpon. Choanophora cucurbitarum was isolated only from fresh okra and Colletotrichum capsici from fresh pepper. Aspergillus flavus was isolated from all the vegetables except Capsicum annum. Alternaria solani was obtained from 'igbagba' only (Table 2). Organisms associated with sundried vegetables After sundrying for 5 days, practically the same number and species of mycoflora isolated from the fresh vegetables were, isolated although at TABLE 2 Fungi Isolated From Fresh Corchorus olitorius, Abelmoschus esculentus, Capsicum annum and Solanum macrocarpon
Vegetable Corchorus olitorius
Capsicum annum
Abelmoschus esculentus
Solanum macrocarpon
Fungi isolated Cercospora corchori Rhizopus oryzae Fusarium solani Aspergillus niger Sclerotium rolfsii A spergillus fla vus Aspergillus niger Rhizopus stolonifer Colletotrichum capsici Fusarium equiseti
% isolation* 13.3 6.6 26.6 6.6 6.6
6.6 6.6 6.6 13.3
13.3
Rhizopus oryzae Aspergillus niger Cercospora abelmoschi Choanephora cucurbitarum Fusarium equiseti Aspergillus flavus
13.3 13.3 6.6
Alternaria solani Rhizopus oryzae Sclerotium rolfsii Aspergillus flavus
26-6 13.3 6.6 6-6
6.6 6.6 6.6
*The figures represent the frequency of isolation of each fungus from plated pieces/samples of each vegetable. Values are means of three replicates.
259
Mycoflora of sundried vegetables TABLE 3 Fungi Isolated From Corchorus olitorius, Capsicum annum, Abelmoschus esculentus and Solanum macrocarpon After Sundrying for 5 Days
Vegetable Abelmoschus esculentus
Fungi isolated
% isolation*
Aspergillus niger Rhizopus oryzae Rhizomucor pusillus Fusarium equiseti Aspergillus flavus
26-6 26-6
Corchorus olitorius
Aspergillus niger Aspergillus flavus Rhizopus oryzae Penicillium claviforme
13.3 26.6 33.3 13-3
Solanum macrocarpon
Aspergillus niger Aspergillus flavus Rhizopus oryzae Penicillium digitatum
13.3
Capsicum annum
Aspergillus niger Aspergillus fumigatus Rhizopus oryzae
13.3 33.3
13.3
13.3 33.3
13.3 13.3 13.3 33-3
*The figures represent the frequency of isolation of each fungus from plated pieces/samples of each vegetable. Values are means of three replicate samples.
higher percentages. However, S. rolfsii was not isolated from all the sundried vegetables while Penicillium claviforme and P. digitatum were obtained from jute and 'igbagba', respectively (Table 3). Organisms associated with sundried stored vegetables The number and kinds of mycoflora isolated from Abelmoschus esculentus was almost constant throughout the storage period except for the 3rd-5th weeks during which fewer and different fungi were obtained (Table 4). C. olitorius and S. macrocarpon had fewer species and number ofmycoflora in the first 3 weeks of storage and these increased till the end of the experimental period. C. annum was host to only three organisms over the whole period of storage (Table 4). The number of fungi isolated from the preserved vegetables was greater than from fresh vegetables (Tables 2 and 4). The percentage microbial occurrence in A. esculentus was 60-83% within the first 2 weeks of storage rising to 99% by the third week, declining to 66% in the fourth week and peaking at 83% by the eighth week (Table 5).
260
A. Adebanjo, E. Shopeju TABLE 4 Fungi* Isolated from Four Sundried Stored Vegetables Over an 8-week Period Vegetable
Abelmoschusescu&ntus
Corchorusolitorius
Weeks 1
2
3
4
5
6
7
8
An Af
An Af
An Aft
An Aft
An Aft
Ro
Ro
Ro
Ro
Ro
Po Rp
Po Rp
Mp --
An Af Af Ro Rp
An Af
Ro
An Aft Af Ro Rp
Po Rp
Po Rp
Po Rp
An Aft
An Af Ro Fe Po
An Aft Ro Po --
An Aft Af Ro Fe
An Aft Af Ro Fe
An Aft Af Ro Fe
An Aft Af Ro Fe
An Aft Af Ro Fe
Ro
Fe Po
Po
Po
Po
Po
Po
--
--
--
Rp
Rp
Rp
Rp
Rp
An Aft Af Fe
An Af Aft Ro Fe
An Aft Af Ro Fe
An Aft Ro Fe
An Aft Ro Fe
Ro
An Aft Af Ro Fe
Po
Po
An Aft Af Ro Fe
An Aft Af Ro Fe
Po
--
Po
Po
--
Po
Po
Rp
.
An Af Ro
An Af Ro
An Af Ro
An Af
-
So&num macroca~on
-
Capsicum annum
-
-
An Af Ro
-
-
-
-
An Af Ro
-
-
.
-.
An Af Ro
.
An Af Ro
.
Ro
*An, Aspergillus niger; Af, Aspergillus fumigatus," Aft, Aspergillus flavus; Ro, Rhizopus oryzae; Po, Penicillium oxalicum," Rp, Rhizomucor pusillus; Fe, Fusarium equiseti. C. olitorius h a d 5 5 % m i c r o b i a l l o a d in the first 2 weeks, w h i c h increased to 6 0 % b y week 4 a n d to 7 5 % b y week 8 ( T a b l e 5). S. macrocarpon h a d 5 3 % m i c r o b i a l invasion within 2 weeks o f storage, 5 0 % b y week 4 a n d 6 0 % b y the 8th week. C. annum h a d 8 3 % m i c r o b e s within the first 4 weeks o f storage a n d 7 5 - 8 3 % in weeks 6-8 ( T a b l e 5). G e n e r a l l y , m i c r o b i a l o c c u r r e n c e was highest in A. esculentus a n d C. annum o v e r the p r e s e r v a t i o n period. F o r all the s t o r e d vegetables, the highest p e r c e n t a g e m i c r o b i a l o c c u r r e n c e was r e c o r d e d within the first 3 weeks ( T a b l e 4).
DISCUSSION Fusarium equiseti was isolated f r o m fresh o k r a fruits in this s t u d y a l t h o u g h F. solani was earlier r e p o r t e d to cause fruit rot, in the field, b y J o a q u i m
60-0+0-0 50.0+0-0 50.0 + 0.0 60-04-0.2
1 83.3+0.1 55.5+0.0 53.3 ± 0.1 83.8±0-1
2 99.3+0.1 47-54-0.0 57.1 ± 0.0 99-9+0.0
3 66.6±0.0 60-0±0-1 50.0 + 0.0 83.3 ± 0.1
4
5 66-6±0.1 42.8±0.1 47.5 4- 0.1 75.0±0.0
Week
75.0±0.0 60.0±0.2 55.5 4- 0.1 83-34-0-1
6
83.3±0.1 66.6+0,0 57.1 ± 0.0 83-3-4-0-1
7
83.2 ± 0-1 75-0±0.0 60-0 ± 0.0 83.3±0-1
8
*Some of the fungi most commonly isolated from the sundried stored vegetables were Aspergillusflavus, Aspergillusfumigatus, Aspergillus niger, Penicillium oxalicum, Rhizopus oryzae, Rhizomucor pusillus, Fusarium equiseti. All values are +standard error.
A. esculentus C. olitorius S. macrocarpon C. annum
Vegetable
TABLE 5 Mean Percentage Microbial* Occurrence in Four Sundried Vegetables Over 8 Weeks of Storage at 28 ± 2°C
bo O~ m
~"
~. ~"
262
A. Adebanjo, E. Shopeju
(1976) and Odebunmi-Osikanlu (1977). This phenomenon could be due to location, and agroecological differences in the sources of the vegetable, apart from the dynamic nature of the pathogens. Cercospora corchori and C. abelmoschi isolated from fresh jute and okra, respectively, in this study were also reported to cause leaf spot and leaf mould on these vegetables in the field (Joaquim, 1976; Odebunmi-Osikanlu, 1977). Besides, C. corchori was found to be seed-borne in Corchorus olitorius (Joaquim, 1976). Similarly, Sclerotium rolfsii isolated from C. olitorius in this study had also been reported as a field disease of the crop (Odebunmi-Osikanlu, 1977). The isolation of Choanephora cucurbitarum from fresh okra fruits is in agreement with Odebunmi-Osikanlu (1977) and Adebanjo (1985). Aspergillusflavus isolated from the majority of the fresh vegetables clearly indicates the significance, in the air, of this species. It is reasonable to conclude that most of the fungi isolated from these fresh vegetables were pathogens acquired in the field. Isolation of almost the same number and species of mycoflora from both fresh and sundried vegetables means that some of these fungi were actually carried into storage and were capable of surviving during the 5-days sundrying. Apart from this, Rhizopus sp., Aspergillus niger, A. fumigatus have been reported as post-harvest pathogens of pepper in Nigeria (Adisa, 1985; Arinze, 1987). Mucor pusillus, Penicillium digitatum and A. flavus isolated from the vegetables after sundrying were probably part of the air spora. An increase in the number of fungi isolated from the preserved vegetables over time indicated their importance as storage fungi. These included Fusarium equiseti found in warm climates on pepper (Uma, 1981), A. fumigatus, and A. niger on Asparagus (Seenappa et al., 1980). The microbial occurrence on the preserved vegetables was highest within the first 3 weeks of storage because there was an abundance of fresh vegetable material on which the organisms could grow. The decline recorded in the number of microbes present at the later part of the storage ,period was probably an indication of exhaustion of the nutrients they contained. Peaks of microbial occurrence, towards the end of the storage period, for some of the vegetables could perhaps be an indication of increased microbial load resulting from cell multiplication/growth. The relatively thick slices of okra fruit compared with the thin flat-leaved vegetables used for this study accounted for their higher microbial loading. The present study clearly indicates the presence of A. flavus and other Aspergillus spp. on the vegetables at harvest, during sundrying and in storage. Since some of these organisms have been shown to produce aflatoxins in other horticultural products, this aspect requires further investigation with a view to highlighting the possible health hazards it might pose.
Mycoflora of sundried vegetables
263
REFERENCES Adebanjo, A. (1985). Premature fruit abortion, a new disease of okra in Nigeria caused by Choanephora cucurbitarum. J. Plant Prof. Tropics, 2, 131-3. Adisa, V. A. (1985). Microorganisms associated with the spoilage of Capsicum annum and C. frutescens in Nigeria. Fitopatologia Brasileira, 10, 427-32. Arinze, A. E. (1987). Microorganisms associated with harvested pepper fruits in Southern Nigeria. Fitopatologia Brasiler, 12, 91-3. Barnett, H. L. (1967). Illustrated Genera of Imperfect Fungi. 2nd edn, Burgess Publishing Company, Mineapolis, USA. Ihekoronje, A. I. & Ngoddy, D. O. (1985). Integrated Food Science and Technology for the Tropics, Macmillan Publishers Ltd. pp. 115-39. Ikediugwu, F. E. O. (1981). A shoot disease of Amaranthus spp. in Nigeria associated with Choanephora cucurbitarum. J. Hort. Sci., 56, 289-93. Joaquim, M. (1976). Seed-borne mycoflora of Nigerian local vegetables and their importance in disease development. PhD Thesis, University of Ibadan, Nigeria, pp. 21-42. Kuku, F. O. (1980). Technical Report, National Institute of Stored Products Research, Ibadan, Nigeria, 7, 63-68. Odebunmi-Osikanlu, Y. O. K. (1977). The more important diseases of selected local vegetables in Nigeria. Nigerian Journal of Plant Protection, 3, 79-83. Ogunsanwo, B. M., Faboya, O. O., Idowu, O. R., Ikotun, T. & Akano, D. A. (1989). The fate of aflatoxins during the production of 'Ogiri' a West African fermented melon seed condiment from artificially contaminated seeds. Die Nfihrung, 33, 983-8. Seenappa, M. L., Stobbs, W. & Krayah, G. (1980). Aspergillus colonization of kinds of pepper during storage. Phytopathology, 70: 218-22. Uma, Ngwanna, U., (1981). Four post-harvest diseases of the Nigerian red pepper (Capsicum annum L.). Plant Disease, 65, 915-16.