Postharvest Biology and Technology 17 1999 189 – 199
Genetic diversity and biocontrol activity of Aureobasidium pullulans isolates against postharvest rots
Leonardo Schena
a
, Antonio Ippolito
a,
, Tirtza Zahavi
b
, Lea Cohen
b
, Franco Nigro
a
, Samir Droby
b
a
Dipartimento di Protezione delle Piante dalle Malattie, Uni6ersita` degli Studi di Bari, Via Amendola
165
A,
70126
Bari, Italy
b
Department of Posthar6est Science of Fresh Produce, Institute for Technology and Storage of Agricultural Products, ARO, The Volcani Center, P.O. Box
6
, Bet Dagan
50250
, Israel Received 10 February 1999; accepted 31 May 1999
Abstract
Aureobasidium pullulans, a cosmopolitan yeast-like fungus, colonises leaf surfaces and is a potential biocontrol agent for plant pathogens. Forty-one isolates of the fungus from the surface of several fruits and vegetables cultivated
in Southern Italy were compared by molecular analysis and biocontrol activity. Characterisation of the isolates by using arbitrarily primed PCR ap-PCR confirmed the presence of a high genetic variability within the species. All the
isolates were evaluated for their ability to control postharvest grey mold of apples, and two of them SL250 and SL236, plus a proven antagonist isolate L47, were able to control Penicillium digitatum on grapefruit, Botrytis
cinerea, Rhizopus stolonifer and Aspergillus niger on table grape and B. cinerea and R. stolonifer on cherry tomato. Furthermore, preharvest application of isolate L47 on table grape resulted in a significant reduction of grey mold
ranging from 27.1 to 49.5 compared to the untreated control. Random amplified polymorphic DNA technique RAPD was a useful method for the identification and evaluation of the survival rate of the applied antagonist.
© 1999 Elsevier Science B.V. All rights reserved.
Keywords
:
Aureobasidium pullulans; Postharvest; Biological control; RAPD-PCR; ap-PCR www.elsevier.comlocatepostharvbio
1. Introduction
Currently, fungicide treatments represent the primary method for the control of postharvest
diseases of fruits and vegetables Eckert and Ogawa, 1985. However, public concern about
fungicide residues, and development of fungicide- resistant isolates of postharvest pathogens have
promoted the search for alternative means, less harmful to human health and to the environment.
In recent years, considerable success has been achieved utilising microbial antagonists to control
postharvest diseases and a large body of informa- tion on the subject is now available Wilson and
Wisniewski, 1994. Because the infection of fruits by postharvest pathogens often occurs in the field
Corresponding author. Tel.: + 39-080-5443053; fax: + 39- 080-5442911.
E-mail address
:
antonio.ippolitoagr.uniba.it A. Ippolito 0925-521499 - see front matter © 1999 Elsevier Science B.V. All rights reserved.
PII: S 0 9 2 5 - 5 2 1 4 9 9 0 0 0 3 6 - 8
prior to harvest Biggs, 1995, it may be advanta- geous to apply antagonists before harvest. For
this approach to be successful, putative biocontrol strains must be able to tolerate low nutrient
availability, UV-B radiation, low temperatures, and climatic changes. The yeast-like fungus Au-
reobasidium pullulans de Bary Arnaud is one of the most widespread and well-adapted sapro-
phytes, both in the phyllosphere Blakeman and Fokkema, 1982 and in the carposphere. A. pullu-
lans has a high tolerance to desiccation and irradi- ation Elad et al., 1994; Kohl et al., 1995, and
has been considered as a possible biocontrol agent of postharvest diseases Bhatt and Vaughan, 1962;
Leibinger et al., 1997.
A. pullulans is characterised by an extreme vari- ability and genetic instability Cooke, 1959; Bulat
and Mironenko, 1992; Yurlova et al., 1995. Mor- phological and cultural characteristics alone are
not sufficient to assess interspecific variability and to differentiate closely related strains. Random
amplified polymorphic DNA RAPD and arbi- trary primed polymerase chain reaction ap-PCR
techniques can be valid means for characterising naturally occurring micro-organism populations
and to obtain preliminary information about the genetic complexity of a natural epiphytic popula-
tion Welsh and McClelland, 1990; Williams et al., 1990.
Biological control agents differ fundamentally from chemical fungicides in that, in order to be
effective, they must grow and proliferate on the plant surface. Therefore, effective antagonists
must become established in crop ecosystems and remain active against target pathogens during pe-
riods favourable to plant infection Lo et al., 1998. The survival ability of biocontrol agents, in
terms of population size, survival period, and distribution in crops, needs to be surveyed and
compared with biocontrol effects. The most fre- quently used methods for the detection and quan-
tification of microbes in ecosystems is the viable-cell count, typically reported as colony
forming units cfu per unit area or sample weight Li et al., 1996. However, the introduced strains
are
difficult to
distinguish from
indigenous strains. RAPD markers allow the unequivocal
recognition of the selected antagonistic strain af- ter its introduction into the phyllosphere Gullino
et al., 1995. The aims of this research were: i to obtain
preliminary information about the genetic diver- sity of a natural epiphytic population of A. pullu-
lans, isolated in Southern Italy, by using ap-PCR; ii to evaluate the potential of several isolates of
A. pullulans as biocontrol agents; iii to determine the effectiveness of an isolate of A. pullulans L47
previously tested in Southern Italy Lima et al., 1996, 1997; Nigro et al., 1997 in a new environ-
ment Israel and on a different table grape vari- ety; iv to identify and to monitor the population
of the antagonist L47 in the field using RAPD- PCR.
2. Materials and methods
