Pseudomonas syringae genes induced during colonization of leaf surfaces

The foliar pathogen and ice nucleator, Pseudomonas syringae pv. syringae B728a, demonstrates a high level of epiphytic fitness on plants. Using a promoter-trapping strategy termed habitat-inducible rescue of survival (HIRS), we identified genes of this organism that are induced during colonization of healthy bean leaf surfaces. These plant-inducible genes (pigs) encode diverse cellular functions including virulence, transcription regulation, transport, nutrient acquisition and other known and unknown loci, some of which may result in antisense transcripts to annotated P. syringae genes. Prominent among the pigs was ssuE, a gene in the sulfate-starvation regulon, indicating that sulfate is not abundant on leaf surfaces. inaZ reporter gene fusion assays of the plant-inducible loci revealed up to 300-fold higher levels of pig transcriptional activity on plant leaves compared with minimal medium. However, the maximum levels of pig transcriptional activity were typically too weak to be measured using a gfp reporter gene. One exception was orf6 in the hrp/hrc pathogenicity island which was highly induced in epiphytic P. syringae cells. Four pigs were disrupted by insertional mutagenesis. While growth of the ssuE mutant was impaired under certain conditions in laboratory medium, the epiphytic and virulence properties of the mutants on bean plants were identical to wild-type P. syringae. Our results demonstrate the utility of HIRS to identify genes expressed on leaves and provide new insight into the leaf surface environment.