2.10.14
ISOLATION AND CHARACTERIZATION OF EPIPHYTIC ERWINIA HERBICOLA AND PSEUDOMONAS FLUORESCENS WITH POTENTIAL APPLICATION AS BIOLOGICAL CONTROL AGENTS

A BONATERRA, E BADOSA and E MONTESINOS

Institute for Food and Agricultural Technology-CERTA, University of Girona, Avda. Luis Santala, s/n. 17071 Girona, Spain

Background and objectives
Recent progress in biological control has generated interest in the characterization of new biocontrol agents. Some bacteria, particularly Pseudomonas fluorescens and Erwinia herbicola are effective as biological control agents against several bacterial and fungal plant pathogens. The aim of this study was the isolation of E. ;herbicola and P. ;fluorescens strains and their characterization for the ability as biological control agents of fruit tree diseases including production and post harvest.

Materials and methods Over 410 strains of E. ;herbicola and P. ;fluorescens were isolated from either aerial parts or roots of several crops. in vitro antagonism against the pathogens E. ;amylovora, P. ;syringae and Stemphylium vesicarium was studied in Luria Bertani, King's, glucose asparagine and potato dextrose (agar. In vivo antagonism was determined using a detached leaf assay for S. ;vesicarium, an inmature pear disc assay for E. ;amylovora and an apple assay for Penicillium expansum. The most interesting antagonistic strains were phenotipically characterized for in vitro antagonism against eight fungal and three bacterial plant pathogens and were studied in relation to the utilization of 95 organic carbon sources. The strains were also genotipically characterized by macrorestriction fragments of DNA using pulsed-field gel electrophoresis (PFGE) with low frequency target restriction enzymes.

Results and conclusions
The group of 410 isolates was composed of 217 P. ;fluorescens and 193 E. ;herbicola. Most of the P. ;fluorescens isolated from roots belonged to biovar V, whereas those isolated from aerial plant parts were classified in biovar V and biovar I. Most of the E. ;herbicola isolates belonged to biogroup I. A correspondence analysis between antagonism and several strain characteristics, origin, and media conditions for antagonism expression indicated that antagonism depended on the growth medium and on the pathogen used as indicator microorganism. The GA medium showed the highest frequency of antagonistic strains: 40% of the strains were antagonistic to E. ;amyiovora, 16% to Pseudomonas syringae and 7% to Stemphylium vesicarium. The percentage of antagonists was higher among isolates of P. ;fluorescens than E. ;herbicola and among isolates from roots than from aerial plant parts. A more detailed study was performed with P. ;fluorescens strains, some isolates significantly inhibited infection of pear tissue by E. ;amylovora, other isolates were able to reduce infection produced by P. ;expansum on apples and some others inhibited leaf infection caused by S. ;vesicarium, only a few inhibited the three pathogens. However there was no relationship between in vitro and in vivo antagonism. Several groups of strains were obtained according to their antagonistic spectrum against several bacterial and fungal plant pathogens. Some strains had strong antifungal and antibacterial activity and others showed only antibacterial activity or only antifungal activity. The strains also differed in carbon source utilization patterns and genetic characterization. No relationship was observed between antagonism pattern, carbon source utilization profiles and genomic fingerprinting.