H-041. Mutation of relA Disrupts Quorum Sensing Signal Production and Growth Characteristics in a Sphingomonas sp. Isolated from a Grape Crown Gall Tumor

H. Gan1, E. Szegedi2, M. Savka1;
1Rochester Inst. of Technology, Rochester, NY, 2Res. Inst. for Viticulture and Enology, Kecskemet, HUNGARY.

Bacterial isolates were collected from grape field crown gall tumors. One-hundred and twenty-eight isolates were purified and characterized for bacterial communication signals called N-acyl-homoserine lactone (acyl-HSL). An isolate, Rr-2-17, was found to produce significant amounts of acyl-HSLs and identified as a member of the Sphingomonas genus using 16S rRNA gene sequencing. Sphingomonas sp. isolate Rr2-17 was subjected to mutagenesis using transposon, Tn5. Approximately 800 kanamycin-resistant mutants were screened for disruption of acyl-HSL signal production. Mutant HX-699 was identified by its lack of acyl-HSL production in cross-streak bioassays using biosensor CV026. DNA sequence analysis of the mutated region in HX-699 showed homology to known relA/spoT genes. The relA/spoT are commonly involved in the regulation of genes in response to bacterial cell nutrient starvation, a response known as the stringent response. In this response bacteria acclimatize to a shortage of nutrients through the accumulation of a global signal called nucleotide alarmones guanosine tetraphosphate (ppGpp) and guanosine pentaphosphate (pppGpp), the products of the relA/spoT locus. The cell surface and growth characteristics were altered in mutant HX-699. Mutant phenotypes could not be restored by the addition of exogenous acyl-HSLs produced by wild type isolate Rr2-17. These findings suggest that relA affects quorum sensing signal production, cell surface and growth characteristics in members of the Sphingomonas genus.