MULTIPLE LOCI OF PSEUDOMONAS-SYRINGAE PV SYRINGAE ARE INVOLVED IN PATHOGENICITY ON BEAN - RESTORATION OF ONE LESION-DEFICIENT MUTANT REQUIRES 2 TRANSFER-RNA GENES

A mutational analysis of lesion-forming ability was undertaken in Pseu domonas syringae pv. syringae B728a, causal agent of bacterial bro cvn spot disease of bean. Following a screen of 6,401 Tn5-containing deri vatives of B728a on bean pods, 26 strains that did not form disease le sions were identified. Nine of the mutant strains were defective in th e ability to elicit the hypersensitive reaction (HR) and were shown to contain Tn5 insertions within the P. syringae pv. syringae hrp region . Ten HR(+) mutants were defective in the production of the toxin syri ngomycin, and a region of the chromosome implicated in the biosynthesi s of syringomycin was deleted in a subset of these mutants. The remain ing seven lesion-defective mutants retained the ability to produce pro tease and syringomycin. Marker exchange mutagenesis confirmed that the Tn5 insertion was causal to the mutant phenotype in several lesion-de fective, HR(+) strains. KW239, a lesion- and syringomycin-deficient mu tant, was characterized at the molecular level. Sequence analysis of t he chromosomal region flanking the Tn5 within KW239 revealed strong si milarities to a number of known Escherichia coil gene products and DNA sequences: the nusA operon, including the complete initiator tRNA(Met ) gene, metY; a tRNA(Leu) gene; the tpiA gene product; and the MrsA pr otein. Removal of sequences containing the two potential tRNA genes pr evented restoration of mutant KW239 in trans. The Tn5 insertions withi n the lesion-deficient strains examined, including KW239, were not clo sely linked to each other or to the lemA or gacA genes previously iden tified as involved in lesion formation by P. syringae pv. syringae.