Variation of the Pseudomonas community structure on oak leaf lettuce during storage detected by culture-dependent and -independent methods.

The genus Pseudomonas plays an important role in the lettuce leaf microbiota and certain species can induce spoilage. The aim of this study was to investigate the occurrence and diversity of Pseudomonas spp. on oak leaf lettuce and to follow their community shift during a six day cold storage with culture-dependent and culture-independent methods. In total, 21 analysed partial Pseudomonas 16S rRNA gene sequences matched closely (> 98.3%) to the different reference strain sequences, which were distributed among 13 different phylogenetic groups or subgroups within the genus Pseudomonas. It could be shown that all detected Pseudomonas species belonged to the P. fluorescens lineage. In the culture-dependent analysis, 73% of the isolates at day 0 and 79% of the isolates at day 6 belonged to the P. fluorescens subgroup. The second most frequent group, with 12% of the isolates, was the P. koreensis subgroup. This subgroup was only detected at day 0. In the culture-independent analysis the P. fluorescens subgroup and P. extremaustralis could not be differentiated by RFLP. Both groups were most abundant and amounted to approximately 46% at day 0 and 79% at day 6. The phytopathogenic species P. salmonii, P. viridiflava and P. marginalis increased during storage. Both approaches identified the P. fluorescens group as the main phylogenetic group. The results of the present study suggest that pseudomonads found by plating methods indeed represent the most abundant part of the Pseudomonas community on oak leaf lettuce.