Multi-pathogen disease caused by Didymella bryoniae and bacteria on Styrian oil pumpkin: microbial ecology and biocontrol

The Styrian oil pumpkin, Cucurbita pepo L. subsp. pepo var. styriaca Greb., is a crop of cultural, medical and commercial importance in Austria but also in other regions of the world, e.g. in China. In the recent years, fruit rot and black rot caused by the ascomycete Didymella bryoniae (Fuckel) Rehm has lead to dramatic yield losses. In the field, the fungal disease was usually associated with characteristic symptoms of bacteriosis. Bacterial pathogens include Erwinia carotovora, Pseudomonas viridiflava, Pseudomonas syringae and Xanthomonas cucurbitae. The high coincidence of fungal and bacterial disease suggests mutualistic effects in pathogenesis. In this study, Styrian oil pumpkin-associated microbial communities with focus on bacterial endophytes were analyzed by microbial fingerprints performed by PCR-Single Strand Conformation Polymorphism analysis (SSCP) and Fluorescence in situ hybridisation (FISH) in combination with confocal laser scanning microscopy. Computer-assisted comparisons of community profiles revealed microhabitat-dependent community structures for Pseudomonas, whereas Bacillus communities were more influenced by the plant development stage. By cultivation dependent methods, the fraction of Styrian oil pumpkin inhabiting in vitro antagonists against D. bryoniae and bacterial pathogens was determined: 9% (=199 strains) of bacterial and fungal isolates showed an antagonistic potential against the fungus. From these in vitro Didymella antagonists, 43 bacterial strains inhibited growth of at least two of the three tested bacterial pathogens. Based on genotypic characterization of these isolates, five potential broad-spectrum antagonists were identified: strains of Lysobacter spp., Pseudomonas chlororaphis, Paenibacillus polymyxa and Serratia plymuthica. They were successfully evaluated in field trials. On their basis, a biological product to protect the Styrian oil pumpkin against multi-pathogen disease is currently under development.