STUDIES OH POrr MMMWESn MiCROlIOLOGY TEdETJiSUS ...Prof. S. E. Saxena AGRICULTURE CENTRE ALIGARH...
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STUDIES OH POrr MMMWESn MiCROlIOLOGY or TEdETJiSUS JIIID FRUITS MIS«TATI0M Submitted in PartiAi Fulfilment of th» flMiairements for tho Award of the Degree, of 0lutttt of 9f|iIo«op|p IN BY f^NilSHl SHARMA Under the SupMVieion of Prof. S. E. Saxena AGRICULTURE CENTRE ALIGARH MUSLIM UNIVERSITV ALIGARH (INDIA) 1992
Transcript of STUDIES OH POrr MMMWESn MiCROlIOLOGY TEdETJiSUS ...Prof. S. E. Saxena AGRICULTURE CENTRE ALIGARH...
STUDIES OH POrr MMMWESn MiCROlIOLOGY or TEdETJiSUS JIIID FRUITS
MIS«TATI0M Submitted in PartiAi Fulfilment of th» flMiairements
for tho Award of the Degree, of
0lutttt of 9f|iIo«op|p IN
BY
f^NilSHl SHARMA
Under the SupMVieion of
Prof. S. E. Saxena
AGRICULTURE CENTRE ALIGARH MUSLIM UNIVERSITV
ALIGARH (INDIA)
1992
DS2422
CERTIFICATE
This is to certify that the dissertation entitled STUDIES
ON POST HARVEST MICROBIOLOGY OF VEGETABLES AND FRUITS
submittd in partial fulfilment of the requirements for
the award of the degree of M.Phil. (Agriculture) in
Microbiology of Agriculture Centre/ Aligarh Muslim
University, Aligarh is a bonafide record of the research
work carried out by Mrs. SHASHI SHARMA under my
supervision and guidance. No part of the dissertation
is submitted for any other degree or diploma. The
assistance and help rendered during the course of this
investigation and sources of literature' are duly
acknowledged.
LENA ) Dean
^acuity of Life Sciences Aligarh Muslim University
ALIGARH.
ACKNOWLEDGEMENTS
I wish to oxjjrosa my pruroiiniJ yrotlludo and lieartfolt souse of
regards to honourable Prof. S.K. Saxena, Dean, Faculty of Life
Sciences, Aligarh Muslim University, Aligarh, under whose guidance,
keen interest , constant supervision and encouragement, I have been
able to carry out th is study. For all this and much more which
is undefinable, I pay my gratitude to him.
I would l ike to express my appreciation to Prof. M.S.
Jairajpuri, F.N.A. Coordinator, Agriculture Centre and Prof. A.K.M.
Ghouse, Chairman, Department of Botany, A.M.U., Aligarh, for
providing all faci l i t ies .
1 feel delighted for continuous help and suggestions given
to me by seniors and friends. I am unable to thank all of them
individually and beg their Indulgence in this matter,
I wish to owe my extreme gratefulness towards my husband,
brothers , parents and parent-in laws for their good wishes, moral
support, patience and sacrifice without which this work could not
be completed.
I am also thankful to Mr. Kafcel A. Khan for typing the
dissertation in present form.
( SHASHI SHARMA ]
CONTENTS
SI . No. CHAPTERS PAGE NO.
1 . INTRODUCTION 1-7
2 . REVIEW OF JUTERATURE 8-48
3 . PLAN OF WORK 49
4 . MATERIALS AND METHODS 50-65
5 . BIBLIOGRAPHY 66-93
LIST OF TABLES
TABLE PAGE NO.
1. Bacteria that cause soft rot of vegetables 09
2. Bacteria that cause of diseases of fruits. 10
3 . Changes in Carbohydrate content of perishables
during pathogenesis. 21-25
4. Changes in protein, protein-bound and free amino
acids in perishables during pathogenesis. 28-34
5. Changes in organic acid in perishables
during pathogenesis. 36-38
6. Reduction in Ascorbic acid content of perishables
during pathogenesis. 40-41
7. In-vivo production of enzymes by post harvest
pathogens of perishables. 44-47
INTRODUCTION
Vegetables and fruits are natural sources of proteins,
carbohydrates , vitamins and minerals, therefore , occupy an
important position in human diets (Work S Carew, 1955). These
are h ighly per i shab le commodity and l iable to damage • both in
the field and after harvest upto the consumers. In the late*,
more damage i s done due to primit ive and defective methods of
handling and s torage . The damages to fruits during post harvest
are no less as compared to pre harvest losses due to diseases
(Sudhir Chandra, 1986). Post harves t losses could be due to
physical , physiological factors and to pathogenic microorganisas.
The phys ica l factor include the mechanical injuries , while
physiological factors the response to post harvest environment.
The microbiological deterioration and spoilage of fruits is the
third category, of course the interactions between the three are
more damaging (Sudhir Chandra, 1986).
Pimentel (1983) estimated that 10-21 percent losses occur
after h a r v e s t , Eckert (1978) enumerated the following consequences
of pos t -ha rves t deterioration of per ishables :
1. Reduced post harves t life of the product due to
accelerated ripening or senescence triggered by ethylene
released from a few diseased fruits in a package or a
storage room
2. Poss ible contamination of the edible product with a
mycotoxin elaborated by the disease inducing
microorganisms, e . g . patulin produced by Penicillium
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expansum in diseased apples (Sommer et^ al^. 1974) or
furanoterpenoid metabolites in sweet potatoes infected
with Ceratocystis fimbriata (Boyd, 1972).
3 . Softening of processed fruits by heat tolerant macerating
enzymes such as those secreted by Rhizopus stolcmifer
in incipient infections on peaches .
According to Smith et^ ^ . (1964) the re are more than
250 known pa ras i t i c diseases of frui ts that cause decay and
blemishes during t r ans i t , marketing and storage. In India precise
data on losses a r e not available but i t is estimated that ttie
average loss of fruits during post harves t periods is at 20-30%
(Mehta et Ql.-1975a>. About 107 million metric tcmnes of total food
produce a r e lost annually (FAO estimates) with high.', percentage
in t ropical countr ies . A loss of Rs. 200 crores annually has been
estimated (only in respect of fruit) due to post harves t diseases
including storage and transit in India.
Ratnam and Nema (1967) while assessing losses from
various s tores and markets observed considerable losses on apples
(12.4-26.7%), banana (15.3-26.7%), ci t rus (14.05-23.2%), mango
(23.3-29.8%), pear (10.32-23.4%), tomato (16.6-31.0%). The losses
were more during August (23.05%), September (26.72%). While
Thakur and Chenulu (1970) observed a single peak of high intensitj
of disease diiring the month i . e . June-July.
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According to Mandal and Das gupta (1981) perishable
fruits and . vegetables suffered an overall loss of greater than 25%.
They divided the fruits into 2 categories on the basis of
damage.
(a) Worst sufferer (above 10% loss) are msmgo, pom^ranate,
apple, pear , mandarian orange, sweet orange, kagzi lime, litchi,
grape, pumpkin, bi t tergourd, brinjal , tomato, c h i l l i , potato, ^u: l ic
and ginger.
(b) Medium sufferer (5-10%) include papaya, pineapple,
banana, guava mango, cucumber, cabbage, cauliflower, radish,
car ro t .
Mandal and Dasgupta (1981) also repor ted that most of
the pathogens affecting fruits are polyphagcus in nature but some
are most specific such as Absidia corymb if era on mandarin orange,
Botryosphaeria r i b i s on pear; Penicillium purpurogenum on mango.
Mandal (1981) found that pineapple, mandarin orange,
grape, tomato, pointed gourd need immediate transportat ion as they
are more l i ab le to damage. The refrigerated frui ts and vegetables
also need fast t ransport as they suffer most. The post harvest
losses to frui ts would depend upai the moisture content of fruits
(Frazier 6 Westhoff, 1989).
To minimize the losses the contact between micro
organisms and our foods (to prevent contamination) and also
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elimination of microorganisms from our foods or adjusting candlitiao
of storage to prevent the i r growth (Frazier and Westhoff, MB)
are essent ia l . If the microorganisms involved in damage are
pathogenic the i r association with our food supply is hazadoos
from a public health point of view. Many of our foods si^port
the growth of pathogenic microorganisms or serve as a vecto of
them. Emphasis should be given to prevent the i r entraittJB and
growth in our foods or eliminate them by processing (Frazier S
Westhoff, 1989).
The microorganisms not only spoil the fruits in
appearances but reduces the quality of fruits and the nutrient
value. Some of the microorganisms causing perishables disease
of fruits and vegetables cause the ailment of human being and
animals (Ilasgupta 6 Mandal, 1989).
Mandal and Dasgupta (1983) isolated at least 17 fang^
pathogens from per i shab les in West Bengal, which are known to
cause diseases in man and animals. Besides, there are tiacteria
obtained from the decomposing perishables which at the same time
may cause disease in animals (Mandal & Dasgupta, 1983 ) .
Alcaligens faecal ls , Enterobacter s p p . e t c . constitute natural
enteric flora in the warm blooded animals and may turn pathogoiic
(Frazier a Westhoff, 1989).
There are large number factors which affect post harvest
losses of per i shables fruit and vegetables (Bhargava 1962;
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Eckert, 1978; Liu 6 Ma, 1983). However, the magnitude A ]pHt
harvest loss i s influenced by initial quality of the
environmental, operat ional , physiological changes during
and genetical factor.
Amongst the physical factors , temperature and icbfte
humidity are known to play a crucial role in decaying prooESS ^
fresh products (Pantast ico, 1975; Harvey, 1978) by loCbKocaiig
growth of the pathogens as well as in pathogenesis. The iBicratM
attack to different crops becomes very slow at temperatures
5°C (Pathak, 1972; Harvey, 1978). The optimum temperatures
0°C are suited for many fruits and vegetables v i z . apple,
carrot, grape (Ecker t , 1978). Chilling injury is noticed in tropkal
fruits of mango, banana, lemon, tomato, grape fruit, pineapple whai
stored at 10-15°C and tielow 12°C in grape fruit which reduces iHBt
resistance against Al temar ia stem end rot (Schiffman-Nadel et ai.
1975; Eckert , 1978). Geotrichum and Erwinia s p . are iriubited
at 0°C but can r ap id ly cause disease when the produce is brou^t
back to high temperatures after storage at 0°C. Botrytis,
Penicillium, Selerotinia, Centrospora, Cladosporium etc . grow sloidy
at near freezing temperatures . Rhi .zopus stolonifer neither grovs
nor their spores germinate below 7.5°C.
Relative humidity near saturation resul ts in heavy losses
except at temperature near 0°C. RH equilibrium below 90% doeai't
permit microorganisms to grow on crop surface (Singh et eil., 1983).
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Deleterious effects of controlled atmosphere on enhancing
disease incidence have also been reported bacterial soft rot of
immature tomato is reported in atmosphere of carbon dioxide (5%)
+ oxygen (3%) [Pearson 5 Hall, 1975). Ethylene, a ripening
inducer, enhance rotting in Alternaria rot of tomatoes; steo end
rot and anthracnose of ci trus (Eckert , 1978).
Proper packaging and handling can reduce injuries and
moisture loss , prevent recontamination and maintain the desirable
storage atmosphere. Plast ic films of low permeability to water
vapour are widely used for packaging consumer units of fresh fruits
and vegetables in developed countries (Grierson, 1969; Eckert,
1975; Harvey, 1978) which reduces the spoilage of fruits. Singh
and Kainsa (1983) observed that bamboo basket for grapes reduces
their spoilage than wooden box.
The fruits and vegetables during post harvest period are
attacked by several pathogens at a time (DGM). The association
have been identified as belong to any one of the following
categories :
1. Non interfering coincidence
2. Predominance of one
3. Unilateral aggravation
4. Unilateral re tardat ion
5. Mutual aggravation
6. Synergism
7. Occupational pr iv i lege of the pioneer se t t ler
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8. Copathogenicity
9. Antagonism
Frui ts undergo tremendous biochemical changes during
ripening (Thind et al.l977) but also during infection. One of the most
obvious of the changes that occur during fruit ripening is the
conversion of insoluble carbohydrate to soluble sugars. In a
number of fungal diseases of plants the general resistance to
colonization has been related to t he sugar content of the tissue
(Horsfall 6 Dimond, 1957), In the same year Horsfall and Dimond
(1957) reported that diseases in which susceptibi l i ty was favoured
by high sugar levels were re fe r red to as high sugar disease and
the converse as low sugar d i s e a s e s . If increasing sugar levels
responsible for the suscept ibi l i ty to rotting in fruits , then such
post harves t disease would be high-sugar d iseases .
Considerable s tudies have been made on the association
of two or more microorganisms on fruits in causing damage (Mandal
8 Dasgupta, 1982). The phenomaion of antagonism has also been
used for adopting biological cont ro l . But information about the
chemical changes brought out i n s ide the fruits as a resul t of
infection with two or more pathogens special ly the antagonists
is very scanty. Most of the s tudies deal with changes brought
out due to infection with single pathogens. Therefore, in the
present studies an attempt will be made to determine the
biochemical changes in the frui ts as a resul t of infection of fruits
with 2 or more pathogens during post harves t operation.
REVIEW
OF
LITERATURE
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The surfaces of healthy fruits include the natural flora
plus contaminating microorganisms from soil and water. In
addition, some fruits contain plant pathogens or saprophytic
spoilage organisms which may grow subsequent to harvesting.
A few microorganisms' are present in the in ter ior of occasional
healthy fruits (Pantastico, 1975).
The microorganisms on the surfaces of freshly harvested
fruits and vegetables include not only those of the normal surface
flora but also those from soil and water . Pseudomonas,
Alcaligenes, Bacillus, Chromobacterium, Enterobacter,
Flavobacterium, Lactobacillus, Leuconostoc, Micrococcus, Sarcina,
Serratia, Staphylococcus, in addition to genera causing plant
pathogens make normal surface bacterial f lora. In case the
surfaces is moist or the outer surface has been damaged, growth
of some of these microorganisms may take place between harvesting
and processing of the vegetables. Some of the important
non-pathogenic diseases include brown heart of apples and pears,
black heeart of potatoes and red heart of cabbage (Tomkins, 1951|
Vaughn, 1963) .
Large number of bacteria have been reported to cause
rots of fruits and vegetables. Some of the de ta i l s are given in
table 1 and 2.
Table No. 1 Bacteria that cause soft rot of vepetables
Bacterium
Ervvinia carotovora sub s p . atroseptica
E. carotovora sub s p . carotovora
Pseudomonas marginalis
P. v i r id i f lava
P. cepacia
P. gladioli pv. allicola
B. polymyxa
Bacillus subti l is
Clostridium puniceum
Low temperature Clostridium
Produce affected
Most vegetables par t icular ly potatoes and some frui ts .
Most vegetables S some frui ts .
Many vegetables
Beans
Onion
Onion
Potatto, Pepper
Potato
Potato
Potato
Author
Peronbelon 5 Kelman (1980)
M
Doudoroff S Pelleroni (1974)
II
II
II
Brocklehurst S Lund (1982)
Gibson S Gordon_ (1974)
Lund et_ al_. (1981)
Brocklehurst S Lund (1982)
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Table No. 2 Bacteria that cause diseases of fruits,
Fruit Disease Pathogen Reference
Grape Bacterial blight Xanthomonas
Crown-gall
Other diseases
Pineapple Fruit collapse
Heart rot
Brown rot
Pink diseases
ampeline
Agrobacterium tumefaciens
Pseudomonas vi t icola
Xanthomonas vi t i s -vini fera
Eriwinia chrysanthemi
E. chrysanthemi
Pseudomonas ananas
6 or
E. ananas
Gluconobacter Acetobacter Enterobacter
Winkler et a l . (1974)
Teviotdale S Schroth (1981)
Hedge -6 Kulkarni (1985)
Nayudu (1972)
Lim fi Lowing (1983)
Lim (1974); Perombelon 8 Kelman (1980)
Serrano (1928)
P o m e g r a n a t e Bacterial blight X. campestris
Young e t^al . (1978J.
Rohrbach 8 Pfeiffer (1976) Ramesh 6 Ramkishan
E. carotovora sub s p . betavasculorum caused soft rot and
vascular necrosis of sugar beet (Thomson 5 Schroth 1981) in addition
to several diseases of fruits and vegetables. According to Chupp
and Sherf (1960) the E. carotovora group of bacteria are probably
the major single cause of microbial spoilage of vegetables.
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Post-harvest soft rots are also caused by E. carotovora (Ramsey
e^ al_., 1959; Smith e^ al_., 1966; Smith and Wilson, 1978). It is
responsible for the cause of blackleg of potatoes (Perombelon 8
Kelman, 1980). Cauliflower and cabbage have also been found
to be spoiled by the group of organisms (Keller and Knosel, 1980).
E. carotovora and E. chrysanthemi a- re reported to cause core
rot found in carrots (Towner and Beraha, 1976). Victoria 6 Granada (1983)
pointed out that E. chrysanthemi caused losses of tomatoes upto
60%. E. amylovora caused a severe outbreak of fire blight affecting
greater than 43000 apple and pear annually (Phocas, 1986).
Strains of Pseudomonas marglnalis are capable of causing
post harves t spoilage of many vegetables e .g. celery (Harrison
and Barlow, 1904), chicory (Friedman, 1951), Lettuce (Dowson,
1941; Ceponis, 1970; Beraha and Kwolek, 1975), cabbage (Bobbin
and Geeson, 1977) and lettuce (Hall et^ al^. 1971). P. viridiflava
was f i r s t described as the cause of reddish-brown necrotic lesions
on a market sample of green bean pods (Burkholder, 1930). It
also caused soft rot of cabbage and lettuce leaves (Wilkie et ed.
1973). P_. chichori caused pos t -ha rves t spoilage of cabbage (Smith
and Ramsey, 1956) and lettuce (.Ceponis, 1970; Grogan et
a l . , 1977). The other species l i k e £ . aeruginosa caused post
harves t spoilage of vegetables. An outbreak of spoilage of onion
bulbs in Australia in 1974 was a t t r ibu ted to £ . aeruginosa. In
a shipment of tomatoes from Florida .17% of the 'decay bacteria'
isolated were identified as £ . aeruginosa. 17% were £ . marginalis
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and 66% Erwinia sp . (Bartz, 1980).
There are a few repor t s that species of Bacillus are
able to cause spoilage of vegetables . The Bacillus failed to attack
carrot or turnip roots, cabbage heads and other plant parts.
B. subt i l i s group was isolated as the cause of soft rot in tomatoes
and caused extensive rot at 36-40^C (Madhok & Fazal, 1943).
Clostridium sp . were isolated from potatoes that showed
extensive soft rot (Campos et_ eil^., 1982; Lund, 1982b). These
are usually present alongwith E. carotovora, but in some craidition
act as primary cause of rots (PerombelonS Kelman, 1980). Several
morphological types of Clostridium have been isolated from rotting
potatoes (Lund et^ al^., 1981; PerombelonQ Kelman, 1979; Campos et
al_., 1982).
Nedumaran and Vidhyasekaran (1981) studied the production
of pect ic enzymes by Corynebacterium michiganens in tomato fruits.
Tomato lines with varying degrees of resis tance to bacterial wilt
l £ . solanacearum) have been repor ted by Tikoo et^ ed. (1984).
Tomato pith necrosis caused by £ . v i r id i f lava (Al vizatos, 1986) .
Calzolari (1986) discussed various bacterial diseases of
tomato i . e . Bacterial spot by Xanthomonas campestris. Bacterial
speck by Pseudomonas syringae, necrosis of the medulla by P.
corrugata and Bacterial canker due to Clavibacter michiganense
and Corynebacterium michiganense (Kurozawa, 1984). Psaidomonas
corrugata found to be causative agent of tomato pith necrosis
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(Olsson, 1986). According to Sasaki a Umekawa (1986) Bacterial canker
(Corynebacterium michiganense, Clavibacter michiganensis) occurred
6 when the population of pathogenic bacteria reached 10 cells/gm
tomato, but d idn ' t occur at lower leve l . Xanthomonas sp. 6
Pseudomonas sp . isolated in mixed culture from tomato (Gitaitis
et a l . , 1987).
FUNGAL SPOILAGE OF FRUITS AND VEGETABLES :
When apples and pears ripen they become susceptible
to a t tack by a variety of fungi to which they were resistant
during the i r period of development on the t ree (Edney 8 chambers
1981). Considerable damage due to rotting caused mainly by o
Gloe^sporium s p p . and Monilia fructigena on apples has been
repor ted (Preece, 1967). Swinburne (1970) repor ted average losses
of apples as 15% in refrigerated gas stores and 3% in air (bam)
s to r e s . The main species responsible for rotting included Nectaria
galligena and Penicillium expansum which accounted for more than
half of all the ro ts examined. A var ie ty of fungal forms including
Gloeosporium, M. fructigena, Penicillium, Botrytis cineria, hi.
galligena, Aspergillus, Rhizopus, Trichothecium and Phoma have
been known to cause rotting of apples as well as pear fruits.
Gloeosporium, Monilia and Penicillium were found to be main
causative agent of rotting (Babovic et_ al_., 1980; Vyas et a l . .
1976). Cryptosporiopsis and Phylactaena s p p . have been reported
to induce lenticel rot during post harvest period (Thind et a l . ,
1977). Both apple and pear have been shown to be susceptible
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to Sclerotium rolfsi i (Sumbali and Mehrotra , 1980) and
Monili-- a taxa (Kaul a Munjal, 1981).
The most important post ha rves t pathogen of soft fruits
i s Botrytis cinerea (Dennis a Mountford, 1975; Davis 8 Dennis,
1977; Mason a Dennis, 1978). Other fungi such as Mucor and
Rhizopus spp mainly infect s t rawber r ies and to lesser extant on
respber r ies (Dennis a Davis, 1977; Mason a Dennis, 1978) and
Cladosporium spp . infect the surface responsible for spoilage of
s t rawberr ies are Colletotrichum fragariae (Howard, 1972),
Dandrophoma obscurans, Pestalotia longischila, Altemaria
tenuissima (Howard and Albregts, 1974), Phytophthora nicotianae
(Matsuzaki et ^ . , 1981), P_. c i t rophthora (Kao and Leu, 1981),
Species of Rhizopus, Aspergil lus, Colletotrichum,
Botryodiplodia, Pestalotia, Phomopsis and Diplodia have been
reported to cause rotting of frui ts of mango during marketing,
t ransi t and storage (Verma a Kamal, 1951; Thakur, 1972;
Laxaminarayana 6 Reddy, 1975 ; Thakur a Chenulu, 1970) reported
that R. arrhizus alone was responsible for decay of 6.3% of mango
fruits in Delhi market. Species of Aspergil lus, Botryodiplodia,
Pestalotia, Phoma, Phomopsis and Rhizopus cause rotting of fruits,
Gloeosporium spp . not only caused anthracnose but rotting of fruits
as well (Dhingra and Mehrotra, 1980.;. Saxena a Saksena, 1983).
Besides these Curvularia, Monilia, Fusarium and As,pergillus,
Phytophthora, Macrophoma, Pesta lot ia , Dichotomyces and
Physalospora have also been reported to attack 15-20% ffuit during
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storage in Agra (Gupta a Madaan, 1970). According to Ooke (ISei)
R. slolonifer is also serious pathogen of post harvest iisears.
Colletotrichum musae, Fusarium roseum, Verticillium th«)br(M^
and Ceratocystis paradoxa. Gloeosporium ni us a rum caused fwst
harvest disease as well as anthracnose (Sudhir Chandra, 1386)
of banana Trichothecium roseum (Srivastava S Tandon, 1971). £.
.moniliforms (Khanna 5 Chandra, 1976) also attacked fruits in
storage. Botryodiplodia (Lantican 5 Quimo, 1978), F_. monlifonne
and F . roseum (Khanna & Chandra, 1976), A. t e rms (Sharma 6
Khan, 1981) have been known to cause post harvest losses of
frui ts . According to Sattar^iHaithami (1986) Macrophomina phaseoli
(Charcoal rot) i s responsible for banana fruit spotting.
Papaya fruits a re damaged most by Botryodiplodia,
Phytophthora, Trichothecium and Cladosporium. CoUetotridium
spp . and Gloeosporium s p p . have been reported to cause rotting
as well as anthracnose during storage (Grewal, 1954). Prasad
and Verma (1976) while making a survey of the diseases of fruits
during storage found that Alternaria, Chaetomium, Fusarium and
Rhizopus were responsible for the spoilage. Other pathogens
reported as post ha rves t cause of loss are Verticillium theobromae
(Srivastava 8 Tandon, 1971) J Macrophomina phaseolina (Kapur 6
Chohan, 1974), £ . solani (Quimio 5 Quimo, 1977), Colletotrichum
capsici (Lai et^ ^ . (1980). Trichothecium roseum (Saxena 5 Jain,
1981) 8 Chaetosphaeropsis truncata (Rai and Bihari Lai, 1984)
Cladosporium harbarum, Botrytis (Jainini, 1923); Botrytis cincrea,
Penicillium digitatum, £ . italicum, A. c i t r i , c i adosporiutn spp.
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(Adam, 1923), Diplodia natalensis (Abbot, 1931), caused
considerable decay of citrus fruits. Besides these other fruit
rot fungi of citrus which cause damage include B. theobromae,.
£ . moniliformae, F_. fumigatus, CoUetotrichum gloeosporbides,
Aspergillus niger (Srivastava 6 Tandon, 1969). Curvularia
tuberculata. C. lunata, A. tenuis, R . nigricans and R. stolonifer
(Garcha et ^ . ' 1980), Phytophthora c i t rophthora , £ . parasitica
(Feld 6 MangQ 1979). Geotrichum candidum (Kamal £t^ ^ . , 1978),
Penicillium digitatum, P . italicum, (Bai, 1979) .
Phytophthora spp are also found associated with post
harves t rotting of ci trus fruits (Uilasa, 1986).
Microbial spoilage is generally brought out by species
of Botryt is , Cladosporium, Gloeosporium, Al temar ia , Stemphylium
and Aspergillus (Mac clel lan 5 Hewitt, 1973; Prakash et al_.,
1976; Sudhir Chandra, 1986).
Geotrichum candidum, Aspergillus s p p . CoUectotrichum
spp . and Pestalotia s p p . have been reported to cause rotting of
fruits of l i th i (Jamaluddin et_^ a\_., 1975)
Phomopsis versoniana caused a severe fruit rot of
pomegranate (Mehta et a l . 1975) Gloeosporium s p p . Fusarium spp.
Penicillium spp . and Aspergillus spp . other well known organisms
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causing fruit rots [Srivastava a Tandon, 1971; Phi l ip , 1981) of
pomegranate.
Only a few fungal spp . such as Ceratocystis paradoxa
and R . stolonifer have been responsible for decay of pineapple.
Tandon (1967) reported Botryodiplodia theobroraae.
Pestalotia sapotae and Hendersonula toruloidea as the cause of
decay of sapota fruits .
Peaches are attacked by wound pathogens e .g. A. niger
and B. theobraomae (Sumbali 5 Mehrotra, 1980; Bhargava et a l . .
1979) which cause tremendous losses .
Botrytis cinerea i s the major cause of post harvest
fruit rotting of tomatoes (Chastagner and Ogawa, 1979; Dennis and
Davis, 1980). For outdoor grown fruits Alternaria spp . are mainly
causative pathogens (Bartz, 1972; Pearson 6 Hall , 1975; Dennis
Q Davis, I960). Other fruit rotting fungi are Stemphylium spp.
Fusarium s p p . Cladosporium s p p . and R. stolonif e r , Pythium spp.
(Pearson 8 Hall, 1974). Phoma destruct iva, Nigrospora oryzae,
Phytophthora, £ . nicotianae, Myrothecium carmichaeli (Kogan, 1979;
Tandon S Singh, 1981) and Geotrichum candidum, Rhizoctonia solani,
Botryodiplodia theobromae (Bartz, 1980). According to Singh et
a l . (1983) post harves t decay of r ipe tomato fruits is caused by
Cladosporium oxysporum.
Phocas (1986) reported the presence of Fulvia
fulva on tomato. Phoma andina v/as found to be new pathogen of
- 1 8 -
tomato (Loerakkar et_ al^., 1986). This pathogen caused necrosis
followed by mummification of f rui t .
The causative pathogen of brinjal are Phytophthora spp. ,
Rhizopus s p p . , Phoma exigua, Fusarium moniliforme. F_. solani.
Solanum melongena, Al temaria , Sclerotium rolfsi i and Phomopsis
spp.(Chowdhry a Hasija, 1981; Ali 8 Shukla, 1981; Da tar, 1981).
Sharma (1981) isolated £ . digitatum from Aegle
marmelos; Cladosporium herbarum from Avverhoa carambola;
Ulocladium chartarum from Carica papaya, Altemaria eribotryae
from Eribotrya japonica, A. tenuissima and Myrothecium roridum
from Memordica charantia, A. terrans and Cylindrocarpon radicicola
from Musa paradisiaca, A. a l ternate , £ . cladosporiodes and £ .
expansum from Physalis peruviana; Curvularia pallescens from
Pyrus communis; A. asculeatus from Spondias mangifera and
Penicillium chrysogenum from Vitis vinifera are found to be
pathogens.
-19 -
The per ishables undergo dras t ic changes in biochemical
status of fruits and vegetables as a result of infection.
Considerable studies have been made both in the favour on the
changes in carbohydrates , proteins , protein-bound and free amino
acid due to pathogens on large number of fruits and vegetables
both in t ropics and temperate areas (basgupta 6 Mandal, , 1989).
It i s difficult to review the entire l i te ra ture . Therefore»some
of important studies carried out on changes in biochemical status
of fruits and vegetables l ike tomato, banana, brinjal , guava, have
been summarised in the t ab les .
CARBOHYDRATE :
Drastic changes in sugar contents was observed in
pineapple, banana, mango, sapota and musambi fruits infected with
Botryodiplodia theobromae (Bhargava, 1962); banana with Fusarium
oxysporum (Chandra S Tandon, 1963); cucumber by Pythium
aphanidermatum (McCombs 8 Winstead, 1964). .
Banana infected with B. theobromae showed decrease in
fructose and glucose content (Williamson 8 Tandon, 1S65|.
Oligosaccharides were not detected in fruits infected with
Pestalotia sapota and B. theobromae on sapota, Gloeosporium psidi i
and Phoma ps id i i on guava (Tandon, 1967). However,
oligosaccharides were detected in mango fruits infected by
Colletotrichum gloeosporioides (Ghosh et^ a^., 1965a), while in
-20 -
musambi fruits infected with B. theobromae the oligosaccharide
developed on the 4th day after infection (SrivastavaS Taridon,1966b)
A significant decrease in sucrose, glucose and fructose
was found in guava fruits infected with Machrophomina
a l lahabadensis (Kapoor S Tandon, 1967)*, pomegranate, guava,
musambi by A. niger (Singh, 1968); banana and papaya with R.
stolonifer; tomato with Phoma destructiva [Aulakh et_ al . 1970b);
ch i l l i f rui ts with Choanephora cucurbitarum (Chahal 8 Grover,
1972); banana with Cochliobolus spici fer 6 Altemaria altemata
(P rasad , 1974), Papaya with Phomopsis caricae (Dhingra S Khare,-
1975); tomato with A. solani (Mehta et_ al_., 1975b) and musambi
frui ts with B. theobromae (Ali, 1976). Such changes were not
detected in healthy vegetables and f ru i t s .
Qualitative and quantitative difference in the sugar
contents in healthy and diseased fruits have been worked out by
many workers (Chahal and Grover, 1972; Ghosh et^ al^., 1964;
Kapoor 6 Tandon, 1971; Singh S Sinha, 1983, 1984). The decrease
in carbohydra te in the fruit t issue has been at t r ibuted to
i ) breakdown of carbohydrates by the enzymes produced
by the pathogens,
i i ) increased respiration of the infected t issue
i i i ) utilization of carbohydrates as substrate for the
production of metabolites.
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-26 -
According to Sharma (1981), qual i tat ive analysis of the
heal thy and diseased fruits and vegetables juices for the presence
of different carbohydrates showed that nearly al l the healthy
fruits and vegetables contained glucose, fructose and sucrose but
for banana where starch was present and grapes where only glucose
was detected. However, in some infected fruits and vegetables
these sugars disappeared simultaneously with the appearance of
an unknown sugar when infected with £ . digitatum, IJ. chartarum,
A. e r ibo t ryae , A. terreus , £ . c ladosporioides , £ . expansum and
£ . pallescens infected frui ts . Total sugar content was lower in Aspergillus
s p . infected tissues (Verma, 1991).
During pathogenesiis t he re i s an increase in the loss
of complex carbohydrate and in i t ia l decrease in single sugars
follows by decrease (Table No. 3 ) .
PROTEIN AND AMINO ACIDS :
Infection of the fruits causes many changes in the amino
acid contents (Kapoor 6 Tandon, 1969a, 1970, 1971; Chahal fi
Grover, 1972). It i s considered that absence or decrease in the
concentration of amino acids in infected fruits may possibly be
a t t r ibuted to the i r preferential uti l ization by the fungus or to
the i r degradation by enzymes or simultaneous utilization in
synthes is of protein.
In cucumber fruits infected with Pythium aphanidermatum
(McCombS S Winstead, 1964); blue ber ry with Glomerella cingulata
-27 -
(Strech 6 Cappellini, 1965); guava fruits with B. theo bromae,
Phoma ps id i i ; banana frui ts with Gloeosporium musarum (Tendon,
1967). Pomegranate, mango and guava fruits infected with
Aspergillus niger (Singh, 1968); Tomato with Drechslera
australiensa (Kapoor 6 Tandon, 1969a) and citrus fruits with B.
theobromae (Srivastava, 1966a); ch i l l i fruits with Choanephora
cucurbitarum (Chahal S Grover, 1972), and tomato fruits with
Altemaria solani (Mehta et^ al^., 1975 a 6 b) decrease in amino
acids contents have been r epo r t ed .
Giorbelidze (1987) worked on effect of the fungus
Macrophoma mantazziana on the composition and content of free
amino acids in Lemon. When the susceptible variety Gruizinskii
and the relat ively res is tant Meyer and Dioskuriya were artificially
infected with the fungus^ content of free amino acid fall in all
va r i e t i e s , having been h ighes t in healthy tissues of the resistant
va r i e t i e s . At the ear ly stage of infection, the quantitative
composition of amino acids fell in all var ie t ies , but at the more
advanced infection stage the range (composition) of amino acids
increased even beyond that in heal thy tissue in the susceptible
var ie ty but not in the r e s i s t an t .
According to Sharaf et ^ . (1989) 14 amino acids can
be detected in apricot and 12 in mango and percentage of each
var ied according to the degree of r ipening.
Both free and protein-bound fractions increase during
pathogenesis, probably due to proteolysis of the host-protein.
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There a r e evidence to show reduction or disappearance of some
amino acid l ike valine in infected t issue (Table No. 4 ) . In some
cases enhanced synthesis of protein (in sweet potato infection with
Ceratocytis fimbriata; in s t rawberry infection with R. stolonifer
and Mucor piriformes (Thompson S Eribo, 1984). Increased protein
content of affected tissues due to the associaticai of fungal mycelium
with h o s t t i ssues . The total free amino acid content of infected
t issues was reduced by infection of Aspergillus s p . (Verma et a l . ,
1991).
ORGANIC ACIDS :
Organic acids are important constituents of the fruits
Drastic changes were found in t a r t a r i c , fumaric succinic, oxalic,
malic a c id s which a re constituents of many fruits and vegetables.
Sr ivas tava 6 Tandon (1966) observed disappearance of organic acids
in guava, mango, musambi and sapota fruits infected with B.
theobromae. Changes in organic acid content has been reported
in c h i l l i infected with R. stolonifer (Tandon et^ al^., 1974); and
tomato f rui ts infected with Altemaria solani and A. tenuis (Mehta
et a l . . 1975 a 6 b) and in peach fruit during infection by A. flavus
(Jamaluddin, 1979).
During pathogenesis, the concentration of some organic
acids increase while in others there is a decrease (Dasgupta 8
Mandal, 1989) c i t r ic , fumaric, malic, malonic, oxalic, succinic acid
decrease probably because these a re consumed during respirat ion.
However in early phases of pathogenesis, there could be an increase
(McElroy, 1961).
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CO
a CO
E 3 x: u
i H
u 4-*
o 4-1 03
i -H i-H O
u
E 3
• F ^
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u o u •c c 1 H
^ H
>) u
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1 - 1
U CO a o o CO
E 3
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XJ c CD u
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4-1
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E 3
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U
o u E 3
• i - i
o 03 x: • J o (H
> s
CD
o a CO
o x: *-» c •^ E »—1 03 X
E s (D
<i-i i H
U 03 Q CO
CD
E o E-
co
c •1-1
u PQ
CD
03
-38-
a •o c CO
CO CO
CO
E (D O U
x:
CD
(0
a 3 00 CO a
a o <
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u 1 - 1
CO
E
"O 0} 00 c CO
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g c o u
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u CO
u i H E u o
00
>
-39-
In tomato infected by D. aus t ra l ience , succinic acid is
produced by host which is reduced by the fungus (Kapoor S
Tandon, 1969; Tandon, 1970). This fungus i s not able to produce
the acid in v i t ro (Table No. 5 ) .
ASCORBIC ACID :
Ghosh £t aa. (1965); Srivastava and Tandon (1966);
Kapoor and Tandon (1969b); Tandon and Jamaluddin (1973),* Thind
et al_. (1977): Kapoor (1982)', Prasad and Bilgrami (1979) have
repor ted that infection by pathogen induced marked changes in
the ascorb ic acid (vitamin C) content in different fruits.
According to Tandon (1970) the loss of vitamin C during
pathogenesis may be due to production of suitable ascorbic acid
degenerating enzymes either by the pathogen alcne or by the host-
pathogen complex. Increased resp i ra t ion in diseased tissue may
also be responsible for a decline of vi tamin.
Reduction in ascorbic acid contents has been observed
in guava infected with Phoma ps id i i (Ghosh et^ al_., 1965b), mango
fruits with Aspergillus niger and papaya frui ts with B. theobromae
(Ghosh et^ al^., 1966).
Rate of decrease of ascorbic acid was faster in mango
fruits infected with B. theobromae only 10.8% ascorbic acid
remained in apple coloured var ie ty of guava while none was left
in safeda var ie ty , both infected with B. theobromae and no
CO
to z u o o s H < QU
z I—I OS
s Q
CO
u • J CO
K U OU
b O
I
TJ C CD
T3 CO CO CO
OH
C^
r Oi
CO CO CD
cu
£35
CO
c
CO
CO CD • M
a 3 DO CO CO
Q o
4 ^
• u o <
r [^ 05
CO 00 CO
u cu
- 4 0
C CO
£ 00
c 1 - 1 CO
-
' ^ rO 00 CJ) rH
CO
n-l cn
T3 C CO
£ C30
5 en
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c o
• o
c CD H
T3 C CO
JS DO
c 1 - 1 CO
,.^ CO 00 C3>
CD
1 H
CO
c CD
CO > CO
•>-> (0 CO
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u en
CD CO CJJ
c O
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CO
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CO n c CO
S do
a 3 00 CO
5
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(J) H
1 00 o C3
s g -• H CO
te S t^ ffi^ (0 w A O
O)
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CO
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CD
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-42-
ascorbic acid was observed in musambi fruits infected with B,
theobromae (Srivastava 8 Tandon, 1966b). In papaya fruits
infected with A. niger, there was 81.3% loss of ascorbic acid
(Singh, 1968), however, there was 92.8% loss after 12 days of
incubation in musambi fruits infected with B. theobromae. Kapoor
and Tandon (1969) found reduction in tomato infected with
Drechslera austral iense, guava infected with A. niger (TandonS Jamalu-
ddin, 1973) chi l l i fruits with Choanephora cucurbitarum (Chahal
6 Grover, 1972). Chilli fruits infected with R. stolonifer (Tandon
et a l , 1974). Papaya fruits infected with A. tenuis (Prasad 6
Verma, 1976) and banana fruit infected with H. specifer (Prasad,
1977).
Tropical fruits are important sources of vitamin C.
Ascorbic acid content normally diminishes during storage, which f
i s enhanced by an injury or infection (Dasgupta a Mandal , 1989),
fungi differ in the ra te and degree of reduction of ascorbic acid
(Table No. 6 ) .
ENZYME :
Chesson (1980) has reviewed the ro le of pectolytic
.microorganisms or cell free pectic enzymes in relatirai to tissue
maceration during post harves t handling and processing. Abdetal
et al^. (198 d) studied enzyme ac t iv i t ies during development of
certain ci trus fruit rot d iseases . Cellulase act ivi ty increased
-43-
in falvedo, a lbedo S pulm of naval organes as a resul t of infection
by Geotrichum candidum, Phytophthora ci t rophthora and Diplodia
natal ens i s , polygalactxoronase was detected at h igher levels in
diseased flavedo and abledo tissues while i t s act ivi ty in the pulp
was l e s s than in controls . Catalase was also p resa i t following
infection, especial in the flavedo. Ascorbic acid oxidase actively
was great ly reduced in . all fruit p a r t s . Per-oxidase activity in
the a lbedo and pulp was highest after inoculation with £ .
citropthora and IB. theobromae; in the flavedo, it was not affected.
Polyphenol oxidase act iv i ty in albedo and pulp tissues remained
unchanged as a resul t of P . ci trophthora infection and decreased
in f lavedo.
Kantaishreede et al^. (1987) observed the effect of
coUetotrichum infection on the oxidizing enzymes of different citrus
frui ts . Changes in act ivi ty of polyphenol oxidase, peroxidase,
ascorbic acid oxidase and catalase were determined in fruits of
3 species inoculated with £ . gleosporioides (Glomerella cingulata).
There was a tendency for activity to be greatest in citrus medica,
which i s l e s s suscept ible than £ . sinencis or £ . re t iculata .
Gautam et^ al^. (1987) observed changes in oxidative
enzyme ac t iv i ty for 15 days at 3 days in tervals in healthy,
incised and diseased peel and pulp of ci trus medica fruits infected
by Thielaviopsis (Ceratocystis) paradoxa. The enzyme studied
were : peroxidase , polyphenol oxidiase , catalase and ascorbic
acid ox idase . Lit t le difference was found in polyphenol oxidase
03 0) U
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-48-
activity in incised and healthy tissues, but more in peroxidase,
catalase and ascorbic acid oxidase activity. Low _gt aJL (1989)
found the reduction in glucose content in potatoes with glucose
oxidase.
Table No, 7 shows the invivo production of enzymes by
post harvest pathogens of perishables.
- 4 9 -
PLAN OF WORK
1. Isolation and identification of fungi and bacteria from fruits
and vegetables (Cruickshank, _et al_., 1973; Riker a Riker, 1936)
from warehouses and market.
2. Pathogenicity of different microorganisms on respective fruits
and changes in ascrobic acid , amino acid, proteins and
"carbohydrate contents.
3 . Inoculation of fruits with two or more microorganisms
simultaneously, sequentially and consequential biochemical
changes.
4 . Inoculation of fruits with pathogenic microorganism and with
microorganisms antagonistic to pathogenic ones and development
of resultant disease and biochemical changes.
5 . Growths of pathogenic microorganism in the presence of
antagonistic microorganisms in v i t ro . Effect of culture f i l trate
on growth of pathogenic microorganisms.
MATERIALS
AND
METHODS
- 50 -
COLLECTION OF SAMPLES
Fresh vegetables and fruits purchased will be collected
and kept in s te r i l i sed containers. Great care will be taken during
transportat ion and handling.
ISOLATION OF MICROORGANISMS
A s t e r i l e normal saline solution dipped swab will be
passed through various area of the resultant inoculum will be
prepared and s treaked upon the Nutrient agar, Eosine methylene
blue agar , Mac conkey agar . Potato dextrose agar and Sabroud's
dextrose medium contained in pe t r i p la tes . The inoculum will be
placed d i rec t ly in the plates or different dilutions will be made.
The pla tes will be incubated for 24 to 48 h r s . for bacter ial
infection and 5-10 days for fungal infection. Bacterial flora will
be identified on the basis of morphology, staining and
physiological ac t iv i t ies (Cruickshank et^ al^., 1973). Fungi will
be isolated in pure culture by hypha l t ip /s ingle spore isolation
and will be identified by using k e y s .
The frequency of fungi/bacteria will be calculated a s :
No. of plates containing a particular fungus/ bacteria Total plates poured
Frequency = bacteria ' x 100
Total no. of colonies of a fungus/bacteria and Relative = ; x 100 Abundance Total no. of colonies of al l the fungi/
bacter ia
- 5 1 -
PREPARATION OF SWABS (Cruickshank et^al^., 1973)
A swab usually consisted of cotton t ight ly wrapped round
one end of a wooden stick 16 cm. long. I t was fitted to a tube
and was autoclave at 15 ps i for 15 miiutes These swabs will be
rubbed aseptical ly on the surface of vegetable and frui ts .
For Isolation of pathogens, the diseased fruits will be
thoroughly washed with tap water and then the surface will be
s te r i l i zed with 0.1% HgCl™ solution for 1 to 2 minutes. After
repeated washing with s ter i le d i s t i l l ed water, diseased lesions
will be removed and will be t ransferred in s ter i l ized pet r id ishes
containing PDA and nutrient agar . Before transfer of the diseased
t issue the petr idishes will be incubated at 28±1°C. The bacterial
colony and fungal hyphae growing out of the diseased tissues will
be t ransferred to fresh pe t r id i shes containing PDA and NA. The
pure cultures will be maintained on PDA and NA s lants .
CONSTITUENTS OF MEDIA TO BE USED FOR IDENTIFICATION OF
BACTERIA
1 . NUTRIENT BROTH : (Sydney and William. 1982)
Broth peptone - 5 g
NaCl - 5 g
Yeast extract - 1-5 g
Beef extract - 1-5 g
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DLstilled water - 1 l i t r e
pH - 7.2 ± 0.2
The above ingredients will be mixed together, adjusted
to pH 7.2
2 . NUTRIENT AGAR : j l . i L o c . c i t )
Peptone - 5 g
Yeast extract - 3 g
NaCl - 5 g
Agar - 15 g
Distilled water - 1 l i t r e
pH - 6.8
The above ingredients will be mixed together and
autoclaved.
3 . MAC CONKEY AGAR (Loc.ci t )
Peptone - 14 g
Protease peptone - 03 g
Lactose - 10 g
Bile sa l t s - 1.5 g
Sodium chloride - 05 g
Agar - 13.5 g
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Neutral r ed - O.OSg
Crystal violet - 0.001 g
Disti l led water - 1 l i t re
pH ± 7.1
4. POTATO DEXTROSE AGAR MEDIUM (Finegold 6 Martin, 1982)
Potato (peeled a sl iced) - 250 g
Dextrose - 020 g
Agar - 015 g
Rose bengal - 0.32 g
Disti l led water - 1 l i t re
The potatoes will be boiled in water for 15 minutes^
filtered through cotton cloth and made upto volume with water .
Added dry ingredients and agar will be dissolved. No pH
adjustment will be required ,
5. SABOURAUD DEXTROSE AGAR (Larone, 1976)
Dextrose - 20 g
Peptone - 10 g
Agar - 17 g
Disti l led water - 01. l i t r e
pH - 5.6
6. RICHARDS MEDIUM (Riker and Riker, 19 36)
Potassium ni t ra te - 10 g
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Potassium dihydrogen
phosphate
Magnesium sulphate
Fer r ic chloride
Sucrose
Distilled water
CZAPEK'S MEDIUM
Sucrose
Sodium ni t rate
Potassium dihydrogen
phosphate
Magnesium sulphate
TH^O
Potassium chloride
Ferrous sulphate
Distilled water
•
^
^
^
^
05 g
2.5 g
0.02 g
50 g
01 l i t r e
(Loc.ci t )
•
^
^
^
^
^
^
30 g
02 g
01 g
0.5 g
0.5 g
0.01 g
01 l itre
8. CITRATE AGAR (Simmon's, 1962)
Agar ^ 20 g
NaCl ^ 05 g
Magnesium sulphate ^ 0.2 g
Ammonium dihydrogen
phosphate ^ 01 g
-55-
Dipotassium phosphate - 01 g
Sodium citrate - 0? 8
Bromethymol blue - 0.08 g
pH - 6.9
9. METHYL RED-VOCES PROSKAUR MEDIUM (Sydney 8 William, 1982)
Buffered peptone
Glucose
Dipotassium phosphate
Distilled water
pH
- 7 g
- .5 g
- 5 g
- 1 litre
- 6.9 ± 0.2
ID. MOTILITY MEDIUM
Beef extract
Peptone
NaCl
Agar
Distilled water
(Loc.ci t )
- 03 g
- 10 g
- 05 g
- 04 g
- 01 l i t r e
In al l the above media, ingredi^i ts will be mixed in
water and boiled. The media will be autoclaved at 15 lbs
pressure for 15 minutes. pH will be adjusted to ± 6.00.
-56-
REAGENTS/SOLUTIONS TO BE USED FOR IDENTIFICATION
1. GRAM'S STAINING SOLUTION (Mucker's Modification)
(A) ( i ) Stock Crystal Violet Solution :
Crystal violet - 20 g
Ethanol (95%) - 100 g
( i i ) Stock ammcnjiiin oxalate solution :
Ammonium oxalate - 1 g
Distilled water - 100 ml.
The two solutions will be mixed and allowed to stand
for 24 hrs and then will be stored in a glass stoppered bot t le .
(B) Grain's iodine solution :
Iodine crysta ls - 1 g
Potassium iodide - 2 g
The above chemicals will be mixed in 5 ml of dist i l led
water, and to th is solution the following will be added .
Distilled water - 240 ml.
5% a q . soln. of sod.
carbonate - 60 ml
The contents will be mixed well and will be stored in
glass stoppered bot t les .
(C) Decoloriser :
Ethanol (95%) and acetone will be mixed in a mixture
of equal volume.
-57-
(D) Counterstaln (Stock safranins)
Safranin - 2.5 g
Ethanol - 100 ml
Working solution : The stock safranin will be diluted
as 1:5 or 1:10 with dis t i l led water and be stored in glass
s toppered bo t t l e .
The bacteria will be stained as per technique suggested
by Sydney and William (1982).
INDOLE PRODUCTION TEST (Sydney a William, 1982)
Reagent :
Paradimethyl amino i
benzaldehyde - 10 g
Ethyl alcohol - 150 ml
Hydrochloric acid - 50 ml.
p-dimethylamino benzaldehyde will be dissolved in
alcohol . To th i s will be added hydrochlor ic acid slowly. The
organism in the nutrient broth will be tested to indole production
by adding 5 drops of kovac 's reagent. A deep red colour will
indicate the production of indole.
2. METHYL RED TEST FOR ACID PRODUCTION (Clark a Lubs, 1975)
Methyl red (0.1 g) will be dissolved in 300 ml of ethyl
alcohol (95%) and diluted with d is t i l led water.
-58-
Appearance of red colour in culture broth by adding 5
drops of methyl red solution will indicate the presence of acid
(pH 4 . 3 ) .
3 . VOCES PROSKAUR TEST (Sydney and William, 1982)
Reagent :
The reagent will be prepared by adding alpha naphthol
(5% in absolute ethyl alcohol) in KOH (40% containing 0.3%
creat inine. To culture grown in broth will be added 0.6 ml of
5% «< -naphthol in a absolute e thyl alcohol together with 0.2 ml
of 40% Potassium hydroxide creatinine solution. After 10-20
minutes the production of bright orange red colour will indicate
the presence of orgeinic acid .
METHODS FOR ESTIMATION OF BIOCHEMICAL CONTENTS
Fresh , s tored, preserved and inoculated fruits and
vegetables shal l be analysed for carbohydrate , protein, amino
acid and ascorbic acid content.
The dr ied pulp will be macerated in ground-glass
homogeniser containing acid washed sand and 20 ml of 80% ethanol.
Later, the pulp will be boiled in 20 ml of 80% ethanol on a water
bath, t h r i ce in order to obtain las t t races of organic compounds
present . The solution, obtained will be kept at temperature
overnight. The solution obtained will be centrifuged at 2,000
- 5 9 -
rpm for 30 minutes. These studies will be init iated after 3 days
of inoculation and will be continued upto 12 days of inoculatioi.
ESTIMATION OF TOTAL PROTEIN
Proteins present in ex t rac t s have to be separated from
other interfering substances pr ior to their estimation.
Proteins will be estimated following the methods of Lowry
et ^ . (1951) by using Fol in ' s reagent .
Reagent A : 2% sodium ions carbonate in 0.1 N NaOH.
Reagent B : 0.5% CuSO.. 5 H O in 1% sodium tartarate tp r tpare
f resh) .
Reagent C : 50 ml of reagent A and 1 ml of reagent B.
Reagent D : Folin-ciocalteu reagent in the Laboratory,
Reagent E : 1 N NaOH.
Standard curve will be obtained with egg albumin.
Albumin (10 mg) will be dissolved In 100 ml of 1 N. NaOH (1
ml) will be diluted to obtain 10 ml by adding more 1 N NaOH.
From th is 0 . 1 , 0 .2 , 0 .3 , 0 .4 , 0 . 5 , 0 .6, 0 .7 , 0 .8 , 0.9 and 1.0
ml will be transferred to test tubes and the volume will be made
upto 1 ml. To th is 5 ml of the reagent_wil l be incorporated.
The mixture will be allowed to stand at room temperature for
about 10 minutes. To this will be added 0.5 ml of Folin's reagent
and mixed immediately.
-60-
After 30 minutes the optical densit ies will be measured
in Bausch and Lomb Spectronic-20 calorimeter at 660 nm against
a reagent blank. The amount of protein in the samples will be
estimated by matching, the optical density with standard curve.
ESTIMATION OF TOTAL FREE AMINO ACIDS .
Amino acids will be estimated by Moore and Stein (1954 J
Method.
Ninhydrin Reagent : To 750 ml of methyl cellulose will be added
20 g of ninhydrin and 3 g of hydr indant in . To this solution 250
ml of sodium acetate buffer CpH 5.5) will be added. .At soon
as the resultant redish reagent becomes yellow, i t will be
transferred into a dark glass bo t t l e . The reagent will be used
within a week.
Standard curve will be obtained by estimating a .mino
acid in a solution of leucine. Ini t ia l ly leucine (10 mg) will be
dissolved in 100 ml of d i s t i l l ed water . Aliquotes of 0.1 to 1
ml in different concentrations will be used . The volume of each,
if l e s s , will be made to 1 ml. To th i s will be added 1 ml of
ninhydrine reagent. This will be boiled for 15 minutes in water
ba th . The absorbance will be determined at 570 nm for all the
amino acid except for proline and hydroxyprol ine which will be
measured at 440 nm in spectrophotometer.
- 6 1 -
ESTIMATION OF CARBOHYDRATE
Carbohydrate would be extracted and estimated by using
the methods of Yin and Clark (1965) and Dubois et^ al^. (1956)
respec t ive ly .
Glucose (Analar , 10 ml) will be dissolved in 100 ml
of d is t i l led water and 1 ml of th i s will again diluted to 10 ml
Ettfferent quantities of the di luted solution i . e . 0.1 to 1.0 ml
would be transferred to test tubes and the volume will be made
upto 1 ml by adding dis t i l led water . To these solutions, 1 ml
of 5% phenol and 5 ml of concentrated sulphuric acid (Analar)
will be added . After 10 minutes optical densities will be
measured in Bausch and Lomb Spectronic-20 colorimeter at 490 nm
against a reagent blank. Graph will be plotted between optical
densit ies and different concentration of the glucose solution. This
will serve as a ' s tandard curve* which will be used for estimation
of carbohydrate in solution.
ESTIMATION OF ASCORBIC ACID
Ascorbic acid is estimated by the visual titration method
based on the reduction of 2, 6-dichlorophenol dye (Tandon et al^.
1974).
The sample will be prepared by blending vegetable/fruit
pulp (10 g) in 100 ml of 2% oxalic acid. The pulp will be sieved
and transferred to 250 ml volumetric flask giving several washing
-52-
2% oxalic acid and the volume will be made to 250 ml, the aliquot
will be filtered through the muslin cloth.
Indophenol reagent (Dye: 2-6, Dich-lorophenol Indophenol)
Sodium 2, 6-dichlorophenol indophenol (50 mg) will be
added to 150 ml glass disti l led water. I t will be warmed to
dissolved the dye NaHCO- (40 mg) will be added . The volume
will be made to 200 ml by adding glass d i s t i l l ed water. The
reagent will be stored in dark glass bottle at 2°C.
Ascorbic acid standard solutLon :
To 50 ml of 2% oxalic acid will be added . 50 ml of
ascorbic acid in a 250 ml volumetric flask and volume will be
made to 250 ml with oxalic acid. Thus giving 0.2 mg of ascorbic
acid in 1 ml.
Standardization :
Indophenol reagent will be s tandardized before use.
Ascorbic acid solution (5 ml) will be pipetted to a white porcelain
dish and will be t i t rated against indophenol d y e . The first
appearance of the pink colour will be taken as an end point.
Similar procedure will be used for the estimation of the
unknown ascorbic ac id .
-63-
Calculatlan :
The ascorbic acid content of the extract will be
calculated as follows :
IxSxD/AxlOO/W = mg of ascorbic acid/100 g of tissue
where.
S = mg of ascorbic acid reacting with 1 ml of the
reagent.
I = ml of Indophenol reagent used in t i t ra t ion.
D = volume of this extract in ml.
A = the aliquot t i t ra ted in ml and
W = the weight of the sample in g.
INOCULATION OF FRUITS AND VEGETABLES :
Healthy r i p e fruits and vegetables will be inoculated
with tes t fungus and bacteria separately and In different
combinations to ascertain the antagonistic behaviour of the two
microorganism. Inoculation of fruits and vegetables by fungus will
be done by pin~prick method while with bacterium by spraying
the fruits with bacter ial suspension after imparting some injury
to the hos t .
Inoculation levels of fungi and bacterium water suspension
will be prepared and number of spores per ml of suspension will
be counted by haematocytometer. Where the sporulating fungus
- 6 4 -
i s not available, mycelial suspension will be made by with known
amount of mycelium. The fruits so inoculated will be incubated
in s t e r i l e des ica tors , the bottom of which wil l be filled with
s te r i l e water.
After different intervals the extent of rotting will be
calculated by measuring the rotted area.
The pulp will be used for estimating the amino-acid,
proteins, carbohydrates , ascorbic acid and enzyme ac t iv i ty .
Appropriate controls will be maintained throughout.
The data will be subject to s ta t is t ica l ana lys i s .
The interaction of different pathogens will also be studied
in v i t r o . The two microorganism will be grown on sterilized
czapecks medium contained in s ter i l ized p e t r i d i s h e s . These plates
will be inoculated with 2 or 3 or 4 microorganism in different
combinations on different places, so that al l the microorganisms
appear to be located at equal distance from the centre of
pe t r i d i sh . The growth of the fungi/bacteria will be measured
and inhibition zone if any, indicating inhibit ion of the
microorganisms will be noted.
The microorganism indicating inhib i tory zones will be
tested for inhibi tory propert ies in the i r culture f i l t ra tes . The
microorganism will be grown in the media supporting the optimum
growth contained in 250 ml Earlemeyer f lask. After 15 days for
-65-
fungi and after 7 days of growth of bacteria the culture fil trate
will be obtained by passing the contents of the flask through
filter paper . The f i l ter paper s t r ips dipped in the culture
f i l t rate will be kept in pet r id ishes containing the media on one
side and on the opposite s ide the microorganism will be grown.
The inhibition zone if any will be determined.
BIBLIOGRAPHY
-66-
Abbot, E.V. (1931). Diseases of economic p lants . Phytopatholog\i
19: 645.
Abdelal, M.R.; M.A. Shatta 6 A.B.H. Abdelaziz (1986). Activities f
of enzymes during development of certain ci trus fruit rot diseases.
Egyptian Journal of Phytopathology, 16(1-2): 11-21.
Abdel-Rehim, M.A.; H. El-Arosi 6 M.S. Hassouna (1974). In:
Post-harvest Pathology of Per ishables . Eds. Dasgupta, M.K. fi
N.C. Mandal: 277-286. Oxford 6 IBH Publishing Co. Pvt . Ltd. ,
New Delhi .
Adam, D.B. (1923). Experiments in storage of f ru i t s . J . Dept.
Agr. Vic tor ia . , 2 1 : 307-317.
All, S.S. (1976). Changes in sugar content of musambi fruits
infected with Botryodiplodia theobromae. Acta Bot. Indica, 4:
63-65.
Ali, S.S. 8 A. Shukla (1981). Pathogenesis of Rhizopus oryzae
in fruit rot d isease of br injal . Curr. Sci . (1981), 50(5): 232-
233.
Alvizatos, A.S. (1986). Tomato pith necrosis caused by
Pseudomonas v i r i d i f l ava . Annales de I ' Ins t i tu t Phytopathologue
Benaki. 15(1): 40-47.
-67-
Aulakh, K.S. ; R.K. Grover 6 S. Malhotra (1970a). Utilization
of free amino acids in vivo and in v i t ro by isolates of Phoma
destructiva Plower. Phytopath . Medit, 9: 9-12.
Aulakh, K.S.; R.K. Grover 6 S. Malhotra (1970b). Changes in
vitamin C and carbohydrate content of tomato fruits inoculated
with isolates of Phoma destruct iva. Phytopath. Medit, 9: 91-94.
Aulakh, K.S. (1977). Abstr . All Indian Symp. on ^Physiology of
Host-pathogen Interaction, Madras (1977): 61 .
Babovic, M.; M. Per i s ic ; S, Markovie 5 S. Stojanovic (1980).
Investigation on rot of apple fruits In cold s tore house. Zastita
Bilia , 30(147): 83-87. Fac. Agric. Belgrade. Zemui.
Yugoslavia.
Bai, D.H. (1979). Survery on the fruit rot occurrence and
damages of shipping mandarin, Korean Journal of Plant Protection,
, 16: 245-247.
Bartz, J .A. (1972). Studies on the causal agent of black fungal
lesions on stored tomato frui t . Proc. Fla St. h o r t . Soc., 84: 117-
119.
Bartz, J .A. (1980). Causes of post harvest losses in a Florida
tomato shipment. Plant Disease, 64: 934-937.
-68-
Beraha, L. 6 W.F. Kwolek (1975). Prevalence and extent of eight
market disorders of Western-grown head lettuce during 1973 and
1974 in The Greater Chicago, Ill inois Area. P I . P i s , Reptr. .
59: 1001-1004.
Bhargava, S.N. (1962), Pathological and Physiological studies
of some fungi causing fruit r o t . D.Phi l . Thes is , Allahabad
University, India.
Bhargava, S.N.; D.N. Shukla a N. Sinha (1979). Some studies
on Botryodiplodia rot of app le . Proc. Nat. Acad. Sci. (India).
49(B): 43-45.
Bilgrami, R.S. 6 R. Verma (1978). All India Symp. on ^Physiology
of ParasitLsmt Abstr . . . 54.
Bobbin, P . a J .D. Geeson (1977). Effects of benzimidazole
fungicide drenches on cabbage storage d i so rde r s . In Experiments
and Development in the Eastern Region : 151-154. Agricultural
Development and Advisory Service.
Boyd, A.E.W. (1972). Potato storage d iseases . Rev. PI. Pathol . ,
51 : 297-321.
Brocklehurst, T.F. a B.M. Lund (1982). Isolation and properties
of psychrotrophic and psychrophi l ic , pectolyt ic strains of
Clostridium. J . Appl. Bact. , 53: 355-361.
-69-
Brown, G.E. a C R . Barmore (1977). The effect of Ethylene on
Succeptibility of Robinson Tangerines to Anthracnose. Phytopath.
^ . , 6 7 : 120-123.
Burkholder, W.H. (1930). The bacterial diseases of the bean.
Mem. Cornell Univ. Agric. Exp. St.* 127.
Calzolari , A. (1986). Bacterial Diseases of tomatoes: Symptoms,
Epidemiology, Diagnosis, Control. Informatore Fitopat .ologico^
36(5): 11-17.
Campos, E . ; E.A. Maher 6 A. Kelman (1982). Relationship of
pectolytic Clostridia and Erwinia carotovora s t ra ins to decay of
potato tubers in s torage . Plant Diseases, 66: 543-546.
Ceponis, M.J . (1970). Diseases of California head Lettuce on
the New York market during the spring and summer months. PI .
DLS. Rept r . , 54: 964-966.
Chahal, A.S. 6 R.K. Grover (1972). Changes in ascorbic acid,
amino acids and carbohydrate content of ch i l l i fruits with
Choanephora cucurbitarum. Indian Phytopath. , 25: 257-260.
Chandra, S. 8 R.N. Tandon (1963). Studies on rotting of apple,
banana, lemon and tomato fruits by fungi. Phyton, 26: 41-47.
Chandra, S. (1986). Post-harvest microbial spoilage of fruits.
Rev. T rop . PI . Path. , 2(1985/86); 365-388.
- 70 -
Chastangnr, G.A. 6 J .M. Ogawa (1979). A fungicide-wax treatment
to suppress Botrytis cinerea and protect fresh market tomatoes.
PhytopatholoRy. 69: 59-63.
Chesson, A. (1980). Maceration in relation to the post-harvest
handling and processing of plant material . J . APPl« Bact., 48:
1-45.
Chowdhury, S.R. a S.K. Hasija (1981). Pathological studies on
Phomopsis vexans causing soft rot of brinjal f rui ts . Indian
Phytopath. 32(3)^ 495-496.
Chupp, C. 6 A.F. Sherf (1960). In: Vegetable Diseases and their
control. The Ronald P re s s , New York.
Clark 6 Lubs (1975). Methyl red t e s t . J . Infect. P i s . , 17:
160.
Cruickshank, R.; J . P . Duguid a R.H.A. Swain (1973).In: Medical
Microbiology.fi)6-1043. Longman Group Limited.
Daradhiyar, P.K. (1979). Presence of pectic and cellulolytic
enzymes in tomato frui ts infected by three pathogenic fungi. In'.
Physiology of Parasi t ism:. Eds . G.P. Agarwal,6 K.S.Bilgrarai, . •
147-154. Today 6 Tommorrow's, New Delhi.
Dasgupta, M.K. a N.C. Mandal (1989). In:Post harvest Pathology
of Per ishables: 623. Oxford a IBH Publishing Co. Pvt . Ltd.
New Delhi.
- 7 1 -
Datar. V.V. (1981). Brinjal fruit r o t caused by Fusarium
monlliforme - a new record from India. Curr . Sci.
49(14): 555.
Davis, R .P . a C. Dennis (1977). The fungal flora of loganberries
in relat ion to storage and spoilage. Ann. Appl. Biol. ^ 85; 301-
304.
Dennis, C. 6 J . Mountford (1975). The fungal flora of soft fruits
in relat ion to storage and spoilage, Ann. Appl. Biol . , 79; 141-147.
Dennis, C. 5 R .P . Davis (1977). Susceptibility of strawberry
var ie t ies to post harves t fungal spoilage. J . Appl . Bact . , 42:
197-206.
Dennis, C. Q R.P. Davis (1980). Fungal spoilage of stored
tomatoes. Bienn. Rep. Fd. Res. Inst . : 44-45.
Dhingra, O.D. 6 M.N. Khare (1975). Post infection changes in
sugar contents of papaya fruits due to attack of Phomopsis caricae»
JNKW Res. J . , 8 : 260.
Dhingra, R. S R.S. Mehrotra (1980). A few unrecorded post
harves t diseases of fruits and vegetables. Indian Phytopath.
(1980)^ 33(3): 475-476.
Doudoroff, M. 8 N.J . Pelleroni (1974). Pseudomonas. In:
Se rgey ' s Manual of Determinative Bacteriology, Eds . R.E. Buchman
a S.E. Gibbons. 8th Edn: 217-243, Williams and Wilkins. Baltimore.
-72-
Dowson, W.J. (1941). Soft-rots due to green fluorescent bacteria.
Trans. Br. My col . Soc.«25: 215-216.
Dubois, M.; K.A. Gilles 6 J .K. Hamilton (1956). Calorimetric
method for determination of sugars and related substances. Anal.
Chem., 28: 350-356.
Eckert , J . W. (1975). . I n : Post harvest
Physiology, handling and utilization of tropical and subtropical
fruits and vegetables: Ed. Pantastico: 550, AVI, Connecticut.
Eckert , J.W. (1978). Pathological diseases of fresh fruits and
vegetables . In: Post harvest Biology and Biotechnologyv^si Food
and Nutrition Press , Westport, Connecticut.
Edney, K.L. 6 D.A. Chambers (1981). Pos t -harves t treatments
for the control of Phytophthora syringae storage rot of apples .
Ann. Appl . Biol . , 97: 237-241.
Feld, S . J . ; J .A. Mange (1979). Brown rot of c i t rus : a review
of the d i sease . Citrograph (1979), 64(5): 101-106.
Finegold, S.M. 6 W.J. Martin (1982). Diagnostic Microbiology.
VI e d i t . Published by C.V. Mosby Company.
Fraz ier , W.C. 6 D.C. Westhoff (1989). In: Food Microbiology.
14: 194-218. Tata McGraw Hill Publishing Company Limited, New
Delhi.
-73-
Friedman, B.A. (1951). Pseudomonas marginalis as the cause of
soft rot of imported wiltloof chicory. Phytopathology, 41 : 880-
881.
Garcha, Hamek Singh 5 Vapinder Singh (1980). Post harvest
diseases of fruits in Punjab. Indian Phytopath. , 33(1):
42-47.
Gautam, S.P. (1987). Studies on oxidizing enzymes of citrus
fruits infected with Thielaviopsis paradoxa. National Academy
Science Le t te rs , 10(1): 9 -11 .
Ghosh, A.K.; R.N, Tandon; K.S. Bilgrami 6 M.P. Srivastava
(1964). I I . Post infection changes in the sugar contents of some
frui ts . Phytopathology, 50: 250-261.
Ghosh, A.K. 6 R.N. Tandon (1965a). Formation of a new
oligosaccharide in mango frui ts under pathogenesis. Curr. Sci . ,
34 : 465.
Ghosh, A.K.; R.N. Tandon; S.N. Bhargava 6 M.P. Srivastava
(1965b). Vitamin C content of guava fruits after fungal infection.
Naturwissenschaften, 52: 478.
Ghosh, A.K.J S.N. Bhargava 6 R.N. Tandon (1966). Studies on
fungal diseases of some t ropical f rui ts . VI. Post infection change
in ascorbic acid contents of mango and papaya. Indian Phy topath. ,
19: 262-268.
-74-
Gibson, T. 6 R.E. Gordon (1974). Bacillus. In: Bergey's Manual
of Determinative Bacteriology, 8th edn . : 529-550, Williams and
Wilkins Baltimore.
Giorbelidze, A.A. (1987). Effect of the fungus Macrophoma
mantegazziana causing gummosis in c i t rus crops, on the composition
and content of free amino acids in lemon. Subtropicheskie
ku l ' t u ry . 2: 113-115.
Gitai t is , R.D.; M . j . Sasser 8 R.W. Beaver (1987). Pectolytic
Xanthomonads in mixed infections with Pseudomonas syringae and
Xanthomonas campestris in tomato and pepper transplants.
Phytopathology. 77(4): 611-615.
Grewal, J .S . (1954). Cultural and Pathological studies of some
fungi causing diseases of f ru i t s . D.Phi l . Thesis , University of
Allahabad, Allahabad.
Grierson, W. (1969). Some random thought on CA research . Mich.
State Univ. Hortic. Kept. , 9: 77-79.
Grogan, R.G.; I . J . Misaghi; A.K. Kimble 6 R. Bardin (1977).
Varnish spot , dest ruct ive d isease of lettuce in California caused
by Pseudomonas ch icor i i . Phytopathology, 67: 957-960.
Gupta, P .C . a R.L. Madaan (1979). Fruit rot disease of bery
from Haryana. Indian Phytopath.« 30(4) : 554-555.
-75-
Hall, C .B. ; R.E. Stall S H.W. Burdine C1971). Association of
Pseudomonas marginalis with pink r ib of le t tuce . Proc. Fla» St.
hort Soc.« 84: 163-165.
Harvery, J .M. (1978). Reduction of losses in fresh market fruits
and vegetables . Ann Rev. Phytopathol. (1978), 16: 321-341.
Harrison, L. S B. Barlow (1904). Some bacter ia l diseases of
plants prevalent in Ontario. Bull. Agrlc. Coll. Exp. Farm, 136:
1-20.
Hasija, S.K. Q S. Batra (1981). Pathological s tudies on Phoma
destructiva causing rot of tomato f ru i t s . Indian Phytopath.^
32: 327-328.
Hedge, R.K. S S. Kulkami (1985). List of diseases of grapes
recorded in India. In: Review of Tropical Plant Pathology. Eds»
Raychaudhuri a J . P . Verma. I I . 141-175. Today a Tomorrows,N.Delhi. Horsfall, J .G. fi A.E. Dimond (1957). Interaction of tissue sugar,
growth substances and disease s u s c e p t i b i l i t y . Z. Pflanzenkranth,
Pflanzenthol Pflanzensuchutz, 64: 415-421.
Howard, Charles , M. (1972). A s t rawberry fruit rot caused by
Colletotrichum fragarie. Phytopathology, 62(6): 600-602.
Howard, C.M. 6 E .E . Albregts (1974). A s t rawberry fruit rot
caused by Alternaria tenuisslana. Phytopathology. 63(7): 938-
939.
-76-
Jainini, J .R . (1923). The chief fungus diseases of the orange
and lemon groves in Spain. Int. Rev, of Sci. 6 Pract of Agr.
N.S. , 1: 61-73.
Jamaluddin, M.P. Srivastava 6 R.N. Tandon (1972). A storage
rot of f rui ts of apple (Mylus s y l v a s t r i s ) . Indian Phytopath. 25:
593-595.
Jamaluddin; M.P. Srivastava 6 R.N. Tandon (1974). Influence
of Myrothecium roridum on the ascorbic acid content of tomato
frui ts . Indian Phytopathol . , 27: 44.
Jamaluddin; M.P. Tandon a R.N. Tandon (1975). Rot of fruits
of l i t c h i (Litchi chinensis) in marketing processes . Indian
Phy t o p a t h . . 28: 530-531.
Jamaluddin (1979). Organic acid composition of peach fruits
during infection by Aspergillus f lavus. Indian Phytopath . , 32:
490.
Kamal; R . P . Singh 6 K.S. Bhargava (1978). Sour rot of lemon
new to India . Indian Phytopathological Society. 31(1): 76-77.
Kantishreede (1987). Influence of Colletotrichum infection on the
oxidizing enzymes of different c i t rus frui ts and the i r importance
in r e s i s t ance . In perspectives in mycological research . I . Ed .
Hasija, S.K., R.C. Rajak 6 S.M. Singh: 127-130. Today and
Toraarrow's Pr inters and Publishers , New Delhi.
ACC No.
-77- \ y~^^^^^'^^...,?.
Kao, C.W. a L.S. Leu (1981). Strawberry fruit rot caused
by Phytophthora cactorum and £ . c i t rophthora . Plant Protection
Bulletin Taiwan , 21(2): 239-243.
Kapoor, I . J . a R.N. Tandon (1967). Formation of a new
oligosaccharide in guava fruits under pathogenesis. Curr. Sci.«
38: 444.
Kapoor, I . J . (1968). Pathological studies of the fruits rot
diseases . D.Phi l . Thesis , Allahabad University, Allahabad,
India.
Kapoor, I . J . 6 R.N. Tandon (1969a). Changes in the free amino
acids of tomato fruit caused by Drechslera austral iense. Indian
Phy topa th . ,22 ; 408-410.
Kapoor, I . J . 6 R.N. Tandon (1969b). Post infection changes in
ascorbic acid content of tomato fruit caused by Drechslera
austral iense. Curr. Sci.« 38: 397-398.
Kapoor, I . J . 6 R.N. Tandon (1970). A new species of Macrophoma
causing frui t-rot of guava, Indian Phytopath , 23: 122-125.
Kapoor, I . J . 6 R.N. Tandon (1971). Biochemical and Pathological
studies on Macrophoma rot of guava. Phytopathology, 70: 137-
144.
Kapoor, I . J . (1982). Changes in ascorbic acid content of
Macrophoma infected guava f rui ts . Indian Phytopath . , 35: 699-
700.
-78-
Kapur, S .P . a J . S . Chohan (1974). Factors affecting infection
and development of fruit rot in papaya by Macrophomina phaseol i .
Indian Phytopath . , 27; 252-253.
Karkun, Deepak a S.S. Ali (1979). The cel l wall degrading
enzymes produced in-vivo by the tomato fruit rot pa thogens^ in l
Physiology of Parasi t ism; 163-169. Todays and tomorrows, New
Delhi.
Kaul, J . L . and R.L. Munjal (1981). Post infection biochemical
changes in apple frui ts due to rot causing fungal pathogens.
Garten bauwissenschaf ten , 45(4); 185-187.
Keller, H. a D. Knosel (1980). Untersuchungeniiber d ie
bakter ie l le Strunkfaule des Lagerkohls. Nachr Bl. d t . Pflschutz.j
32; 161-163.
Khanna, K.K. a S. Chandra (1976). Control of tomato fruit ro t
caused by Fusarium roseum with homeopathic d rugs . Indian
Phytopath . , 29; 269-272.
Kogan, E.D. (1979). The pathogen of black rot of tomato fruits
in Maldavia . Referativnyi Zhumal ^(1977), 9(55); 1248.
Kurozawa, C. (1984). Bacterial Canker of tomato. Biologico,
50: 105-113. Dep. PI . Health, UNESP, 18600 Bohicatu, SP,
Brazil ,
-79 -
Lal, B . ; R.N. Rai 8 A. Arya (1980). Colletotrlchum rot of
papaya. National Academy Science L e t t e r s , , , 2(5) : 174.
Lantican, M.T. 5 T.H. Quimo (1978). Pathogenecity and cultural
character is t ics of Botryodiplodia s p p . causing fruit ro t s .
Phil l ippine Phytopathol . . , 12(1 /2) : 66-74.
Larone, D.H. (1976). Medically important fungi. Harpet and
Row Publ isher Hagerstown Maryland, New York.
Laxminarayana, P. 6 S.M. Reddy (1975). A new post harves t
disease of mango. Indian Phytopath.« 28: 529-530.
Lim, W.H. (1974). The etiology of fruit collapse and bacterial
heart rot of pineapple. MARDI Res. Bull . , 2: 11-16.
Lim, W.H. and Lo wings, P.H. (1983). Pathogenic comparison of
Erwinia chrysanthemi s trains from pineapple with strains from
other h o s t s . MARDI Res. Bul l . , 1 1 : 342-350.
Liu, M.A. 6 P .C. Ma (1983). Post harvest problems of
vegetables and fruits in t ropics and subtropics , Asian Veg. Res.
and Dev. Centre, Taiwan.
Loerakkar, W.M.; M. Lobo 6 B. Navarro (1986). Phoma andina
va r . nov, A new pathogen of tomato and potato in the Andes.
Fitopatologia^ 21(2): 99-102.
Low, N. ; Z. Jiang S M.M. Palcic (1989). Reduction in glucose
content in potatoes with glucose oxidase . J . Food Sci . , 54(1):
118-121.
-80-
Lowry, O.H.; Rose Brou; A.L. Farr [1951). Protein measurement
with the Folin-phenol reagent . J . Biol. Chem,, 193: 265-275.
Lund, B.M.; T .F . Brocklehurst 6 G.M. Wyatt (1981).
Clostridium puniceum, a pink pigmented, pectolytic bacterium.
J . gen. Microbiol.. 122: 17-26.
Lund, B.M. (1982a). The effect of bacteria on post harvest
quality of vegetables and f rui ts , with part icular reference to
spoilage. In: Bacteria and plants : 133-153. Eds^ Rhodes . Roberts,
M.E. a F.A. Skinner, 13-153. , Academic Press London
and New York.
Lund, B.M. (1982b). Clostr idia and plant disease: new pa^..-.,,.
In: Phytopathogenic Prokaryotes . Eds. Mount, M.S. Q G.f ' o c v .
I : 263-283. Academic P r e s s , New York.
Mac clellan W.D. a William B. Hewitt (1973). Early Botrytis
rot of grapes: Time of infection of latency of Botrytis cinerea
Pers in Vitis vineifera. PhytopatholORV. 63(9): 1151-1157.
McCombs, C.L. a N.N. Winstead (1964). Changes in sugars and
amino acids of cucumber fruits infected with Pythium
aphanidermatum, Phytopathology, 54(1): 233-234.
McElroy, W.D. (1961). Cellular Physiology and Biochemistry,
Prentice Hall, Englewood Cliffs, New Jersey .
Madhok, M.R. a Fazal-ud-Din (1943). Bacterial soft rot of
tomatoes caused by a spore forming organism. Indian J . Agric.
Sci . , 13: 129^33 .
- 8 1 -
Mandal, N.C. (1981). Post harves t diseases of fruits and
vegetables in West Baigal . Ph .D. Thesis , Visva-Bharati .
Mandal, N.C. a M.K. Dasgupta (1981). In: Post harves t diseases
of pe r i shab les . By Dasgupta, M.K. a N.C. Mandal, 1989. Oxford
6 IBH Publishing Company Pvt . L td . , New Delhi.
Mandal, K.C. 6 M.K. Dasgupta (1982). Post harvest diseases
of per ishables in West Bengal, Indian Phytopath . , 35; 645-649.
Mandal, N.C. 6 Dasgupta, M.K. (1983). National Seminar on t ^ Constraints in Crop Production of Subhumid Zones: 28.
Mason, D.T. a C. Dennis (1978). Post harves t spoilage of
Scottish raspber r ies in relation to pre-harvest fungicide s p r a y s .
Hort. R e s . . 18: 41-53.
Matsuzaki, M.; M. Khan a A. Kise (1981). Phytophthora rot of
strawberry in kyushu Japan. Annuals of the Phytopathological
Society of Japan. , 46(2): 179-184.
Mehta, P . ; K.M. Vyas a S.B. Saksena (1974). Hindustan b
Antibiotic Bull . , 16: 247-252. In: Post harves t Microbiology of Perisha^es.
Eds. Dasgupta, M.K. a N.C. Mandal.
Mehta, P . ; K.M. Vyas a S.B. Saksena (1975a). Pathological
studies on fruit rot of tomato caused by Altemaria solani and
A. tenuis . Indian Phytopa th . , 28: 247-252.
Mehta, P . ; K.M. Vyas a S.B. Saksena (1975b). Metabolic changes
during pathogenesis of fruit rot diseases of tomato. Indian
Phytopath. . 28: 253-255.
-82-
Mehta, P . ; K.M. Vyas 5 S.B. Saxena (1977). Metabolic changes
during pathogenesis of fruit ro t diseases of tomato. Indian
Phytopath , (1977), 13(1/2) : 14-23.
Midha, S.K. a J . S . Chohan (1967). Factors affecting the
production of pectino ly t ic Kizymes by Gloeosporium ps id i i , the
causal agent of fruit rot of Guava. Indian Phytopath . , 20: 215-
219.
Moore, H. a W.H. Stein (1954). Modified Ninhydrin reagent for
the spectrophotometric determination of amino ac ids . J . Biol.
Chem., 24: 904-913.
Nayudu, M.V. (1972). Pseudomonas vit icola s p . Incitant of a new
bacterial disease of grapevine. Phytopathology, 73: 183-186.
Nedumaran, S. a Vidhyasekaran, P. (1981). Techniques to detect
Corynebacterium michiganese p v . michiganense infection in tomato
seed. Indian Phytopath . , 35: 143-144.
Olsson, K. (1986). A disease of tomato new for Sweden, caused
by Pseudomonas conugata .. Vaxtskydtsnotiser,
50(1): 20-27.
Ooke, J . J . (1981). Guava fruit rot caused by. Rhizopus stolonifer
in Hawaii. Plant Diseases« . , 64(4): 412-413.
Pantastico, E.B. (1975). Post harves t physiology, handling and
utilization of tropical and subtropical fruits and vegetables.
Westport, Conn AVI P u b l . ; 560.
-83-
Pathak, V.N. (1972). Essential plant pathology: 448.
Prakash Publ i shers , Ja ipur .
Pearson, R.C. 5 D.H. Hall (1974). Ripe fruit rot of tomato
caused by Pythium ultimum and Pythium aphanidermatum. Plant
P i s . Rep . . ' 7 ( 1 2 ) : 1066-1069.
Pearson, R.C. & D.H. Hall (1975). Factors affecting the
occurrence and sever i ty of black mould on r ipe tomato fruit caused
by Alternaria a l t ema ta . Phytopathology, 6: 1352-1359.
Perombelon, M.C.M. 6 A. Kelman (1979). Population dynamics
of Erwinia carotovora and pecto lyt ic Clostridium s p p .
PhytopatholoRv. 69: 167-173.
Perombelon, M.C.M. a A. Kelman (1980). Ecology of the soft
rot Erwinia. A Rev. Phytopath . , 18: 361-367.
Ph i l ip , S. (1981). Aspergillus rot of pomegranate f ru i t s . Indian
Phytopath. 32: 332.
Phocas, C.L. (1986).- Plant Pathology Virology. Annual
Report of the Dept. of Agr., Cyprus, for the year 1985. Nicosia,
Cyprus; Ministry of Agr. and Natural Resources: 90-92.
Pimentel, D. (1983). Environmental aspects of pest management.
In: Chemistry and World Supplies. The New Front iers , Chemrawn
II , Ed. Shemitt, L.W. Peragamon Press , Oxford.
-84-
Prakash, O.M.; B. Mishra 5 A .P . Mishra (1976). Greeneria
iuliginea Scribner and viala causing b i t t e r rot of grapes in India.
Indian Phytopath . . 27(4): 603-606.
Prasad, M.M. (1974). Post infection changes in sugar content
of banana fruits (Alternaria a l t emata and Cochliobolus specifer) .
Curr. S c i . . 43: 802.
Prasad, J .S . 6 R.A.B. Verma (1970). Fungal diseases of papaya
fruits in Bihar. Indian Phytopa th . , 23: 722.
Prasad, J . S . 6 R.A.B. Verma (1976). Investigations on the
diseases of papaya. IV. Post infection changes in ascorbic acid
content. Indian Phytopath . , 29: 84-85.
Prasad, M.M. (1977). Post infection changes in vitamin C contents
of banana fruit. Curr. Sc i . , 46: 197.
Prasad, M.M. -^ K.D. Poddar (1977). Physiological changes in
vegetables during pathogenesis. In: Physiology of Microorganisms..
Ed. K.S. Bilgrami, 275-284. Tbday,- and Tomorrow's, New Delhi.
Prasad, S.S. a S.K. Sinha (1979). Post infection changes in
ascorbic acid content of l i tch i fruits caused by certain pathogenic
fungi. Proceedings of the National Academy of Sciences, India
B (1975), 45(1): 43-44.
Prasad, S.S. 5 R.S. Bilgrami (1979). Investigations on diseases
of l i t c h i . VII. Changes in ascorbic acid content of fruits during
pathogenesis of Aspergillus. Indian Phytopath . , 23: 722-724. ,
-85-
Preece, T .F . (1967). Losses of Cox's Orange Pippin apples
during refrigerated storage in England, . PI. Pa th . , 16:
176-180.
Quimio, T.H. 8 A.J . Quimo (1977). A new fruit rot disease of
papaya in the Ph i l l ipp ines . Phi l l ipp ARric. 58(7/8) : 330-331.
Rai, R.N. (1982). Pathological and Physiological studies of
certain fungi causing fruit rot d iseases . D.Phi l . Thes is , Allahabad
University, Allahabad.
Rai, R.N. 8 B. Lai (1984). A new variety of Chaetosphaeropsis
truncata causing fruit rot of banana. Indian Phy topa th . , 37: 566-
568.
Ramesh, C. 8 Ram Kishun (1991). Studies on bacter ia l blight
(Xanthomonas campestris) of pomegranate. Indian Phytopath . ,
44(3): 370-372.
Ramsey, G.B.; B.A. Friedman, 6 M.A. Smith (1959). Market
Diseases of Beets, Chicory, Endive, Lettuce, Spinach and Sweet
Potatoes. Agr. Handbook No. 155, United States Dept. of
Agriculture, Washington, D.C.
Ratnam, C.V. 6 K.G. Nema (1967). Studies on market diseases
of fruits and vegetables . Andhra Agr. J . , 14: 60-65.
Reddy, B.C. ; P.V. Reddy 8 D.G. Raju (1984). Post-infection
changes in acid lime fruits caused by Aspergillus niger . Indian
Phytopath . , 37: 185-187.
-86-
Riker, A . J . 6 R.S. Riker C1936). Introduction to research on
plant disease»John Swift and Co. St. Louis MO: 117.
Rohrbach, K.G. 6 J . B . Pfeiffer (1976). The interaction of four
bacteria causing pink disease of pineapple with several pineapple
cu l t ivars . Phytopathology, 66: 396-399.
Sasaki, T. S M. Uraekawa (1986). Ecological studies on bacterial
canker of tomat6 (The occurrence of bacterial canker of tomato).
Annual Report of the Society of Plant Protection of North Japan,
37: 65-67.
Sattar , M.H.A. 8 M.N. Haithami (1986). Diseases of major crops
in democratic yemen and the i r economic importance. FAQ plant
protection Bulletin. 34(2): 73-76.
Saxena, A.K. 6 S.K. Jain (1981). A new fruit rot disease of
papaya (Trichothecium roseum on papaya) . Curr. Sci. (1981),
50(2): 96.
Schiffmann-Nadel, M.; J . Waks 8 E. Chalutz (1975). Frost injury
Predisposes Grape fruit to storage r o t s . Phytopathology, 65: 630.
Serrano, F .B . (1928). Bacterial fruitlet brown rot of pineapple
in the Phi l l ippine . P h i l l i p p . J . Sc i . , 36: 271-305.
Sharaf, A. ; F.A. Ahmad 8 S.S. El-Saadany (1989). Biochemical
changes in some fruits at different ripening stages. Food Chem.,
31(1): 19-28.
-87-
StSrma, N. (1981). Storage rot of vegetables and frui ts . Ph.D.
Thesis, A.M.U. Aligarh, India.
Sharma, N. 6 A.M. Khan (1981). Two hi ther to unrecorded disease
of banana frui ts . National Academy Science Letter (1978), 1(3) :
85,
Simmono, S. (1962). Citrate ut i l izat ion. J . infect. P i s . , 39:
209-214.
Singh, R.H. (1968). Pathological and Physiological studies of
some fruit ro t s , D.Phi l . Thes is . Allahabad University, Allahabad.
Singh, A. 8 K.K. Sinha (1983). Biochemical changes and aflatoxin
production in guava fruits by Aspergillus flavus and A.
parasi t icus. Indian Phytopa th . , 36: 365-366.
Sinjgh, J . P . 6 R.L. Kainsa (1983). Microbial Flora of Grapes in
relation to storage and spoilage. Indian Phytopa th . , 36: 72-76.
Singh, S.; D.P. Thakur 6 J . P . Singh (1983). Post harvest decay
of r i p e tomato fruits caused by Cladosporium oxysporum. Indian
Phytopath . , 36: 723-724.
Singh, -A. a K.K. Sinha (1984). A new variety of
Chaetosphaeropsis truncata causing fruit rot , of banana. Indian
Phytopath . , 37: 566-568.
Smith, M.A. 8 G.B. Ramsey (1956). Bacterial zonate spot of
cabbage. Phytopathology, 46: 210-213.
-88-
Smith, W.L.; R.D. Rassott 6 R.K. Anderson (1964). Reduction
of post ha rves t decay of peaches and nactarines with heat
treatment. US Dep. Agric. Mork. Res. Rept. 643: 24.
Smith, M.A.; L .P . McColloch a B.A. Friedman (1966). Market
Diseases of Asparagus, Carrots, Pea, Bean and re la ted Vegetables,
Agricultural Handbook No. 303. US Dept. of Agr., Washington,
B.C.
Smith, W.L. 5 J . B . Wilson (1978). Market Diseases of Potatoes,
Agr. Handbook No. 479. US Dept. of Agr., Washington, D.C.
Sommer, N . F . ; J .R . Buchanan a R . J . Fortlage (1974). Appl .
Microbiol. , 28: 589-593.
Srivastava, M.P. a R.N. Tandon (1966a). Free amino acid
spectrum of healthy and infected musambi fruit . Naturwissenchaftan,
19: 508-509.
Sr ivastava, M.P. 8 R.N. Tandon (1966b). Effect of Botryodiplodia
on infection of vitamin C content of mango frui ts . Curr. Sc i . ,
35: 419.
Srivastava, M.P. a R.N. Tandon (1966c). Aspergillus ro t of
pomegranate. Indian Phytopath . , 19: 172.
Sr ivastava, M.P. a R.N. Tandon (1969). Studies on Botryodiplodia
of guava. Indian Phytopath . . 22: 268-269.
-89-
Srivastava, O.P. (1968). Soft rot of desh i mango fruit and the
pathogenicity of the isolated microorganisms. J . Indian Bot. Soc.,
47: 328-329.
Srivastava, M.P.a R.N. Tandon (1971). Aspergillus rot of
pomegranate. Indian Phytopath. , 24: 172.
Strech, A.W. 6 R.A. Capellini (1965). Changes in free amino
acids and reducing sugars in High Bush Blue-berry fruits infected
with Glomerella cinulata. Phytopathology, 55: 302-303.
Sumbali, G. Q R.S. Mehrotra (1980). An unrecorded post harvest
rot of app le . Curr. Sci. (1979), 48(22): 996.
Swinburne, T.R. (1970). Fungal rotting of app le s . I . A survey
of the extent and cause of current fruit losses in Northern Ireland.
Rec. Agric. Res. North. I r e . , 18: 15-19.
Sydney, M.F. 6 J .M. William (1982). Diagnostic Microbiology.
VI,C.V. Mosby comp. Missouri.
Tandon, R.N. (1967). Post harves t diseases of tropical and
subtropical f ru i t s . PL 480. Final Technical Report FG: 133,
University of Allahabad, Allahabad.
TandcHi, R.N. (1970). Certain problems of post harvest diseases
of fruits and vegetables, Indian Phytopath . , 23: 1-15.
-90 -
Tandon, M.P. 5 Jamaluddin (1973). Post infection changes in
ascorbic acid content in fruits of tomato induced by
Cylindrocladium scoparim Margan. Curr. Sc i . , 42: 400-401.
Tandon, R.N.; M.P. Srivastava S Jamaluddin (1974). Studies on
post harves t diseases of fruits and vegetables. Curr. Sci. Trends
PI. Pa th . . : 209-220.
Tandon, M.P. a N. Singh (1981). A new fruit rot of tomato
(Myrothecium carmichae l i ) . Proceedings of the Nat. Aca. of Sci .
India 47(2): 139.
Teviotdale, B.L. a M.N. Schroth (1981). Crown gal l . In: Grape
Pest Management, Publ . No. 4105. Univ. Calif., Berkeley, U.S.A.:
79-80.
Thakur, D.P . ; V.V. Chenulu (1970). Survey of Delhi market for
fungal diseases of frui ts and vegetables in Plant Diseases Problems:
301-308. Indian Phytopathol . S o c , New Delhi.
Thakur, D.P. (1972). Factors influencing storage rot of certain
fruits and vegetables caused by three species of Rhizopus. Indian
Phytopath. , 23: 58-61.
Thind, T .S . ; S.B. Saksena a S.C. Agrawal (1977). Post infection
changes in amino a c i d s , sugars , phenolic substances and organic
acids of apple fruits incited by Clathridium corticola. Indian
Phytopath. , 30: 323-325.
- 9 1 -
Thompson, D .P . 6 E .B. Eribo (1984). Changes in nucleic ac ids ,
protein and phenols in s trawberry frui ts infected with Rhizopus
and Mucor spec ies . Indian Phytopath. . 37: 498-500.
Thorn; son, S.V. 5 M.N. Schroth (1981). Identification and
nutritional differentiation of the Erwinia sugar beet pathogen from
members of E. carotovora and ^ . chrysanthemi, Phytopathology,
71: 1037-1042.
Tikoo, S.K.; N. Anand 6 Ramkishun (1984). Development of tomato
lines with varying degrees of res is tance to bacterial wilt
(Pseudomonas solanacearum). Report of tomato cooperative, 34:
20.
Tomkins, R.G. (1951). The microbiological problems in the
preservat ion of fresh fruits and vegetables . J . Sci. Food Agr. ,
2: 381-386.
Towner, D.B. 6 L. Beraha (1976). Core rot : a bacterial disease
of c a r r o t s . P I . P i s . Rep t r . . 60; 357-359.
Ullasa, B.A. (1986). Identity of Phytophthora s p p . Associated
with Post ha rves t rotting of ci t rus f ru i t s . Indian Phytopath . ,
39(3): 477-478.
Vaughn, R.H. (1953). Microbiological spoilage problems of fresh
and refr igerated foods. In: Microbiological quality of foods. Eds.
Slantz, L.W. Academic Press , Inc . , New York.
-92-
Verma, G.S. 6 M. Kamal (1951). Rot of Manglfera indica Linn,
caused by Aspergillus. Curr . Sc i . , 20: 68-69.
Verma, S.; S. Gupta; R.V. Singh a A.B. Abidi (1991), Changes
in biochemical constituents of fruits infected with Aspergillus s p . ,
Indian Phytopathol . , 44: 405-406.
Victoria, J . I . Q^^'t^anada (1983). A bacterial soft rot of tomatoes
induced by Erwinia chrysanthemi in the cauca val ley.
Fitopatologia. 18(1): 31-33.
Vyas, S.C.; D. Singh 6 N.D. Sharma (1976). Some new fungi
causing post harvest d iseases of apple . PI . P i s . Rep t r . , 60:
988-990.
Wilkie, J . P . ; D.W. Dye Q D.R.W. Watson (1973). Further hosts
of Pseudomonas v i r id i f l ava . N.Z.J . Agric. Res . , 16: 315-323.
Williamson, D. a R.N, Tandon (1965). Naturwissen chaften, 7:
166. In : Post ha rves t pathology of per ishables Eds.Dasgupta,
M.K. a N.C, Mandal, 1989. Oxford IBH Publishing Co. Pvt . Ltd.
New Delhi.
Winkler, A .J . ; J . A. Cook a L.A. Lider (1974). General
vi t icul ture. Univ. Calif. P ress , Berkeley, U.S.A.
Work» P . a J . Carew (1955). Vegetable production and Marketing.
J537, Wiley Eastern Pr iva te Limited, New Delhi.
- 9 3 -
Yin, R.V. a H.E. Clark (1965). Carbohydrate and protein contents
of boron deficient tomato root t i p s . Plant P h y s i o l . , 40: 312.
Young, J .M. ; D.W. Dye; J . F . Bradbury; C.G. Panagopoulos a C.F.
Robbs (1978). A proposed nomenclature and classification for
plant pathogenic bac ter ia . J . Agric. Res . , 21 : 153-177.
