RprR is a plant-responsive regulator of exopolysaccharide production, biofilm formation, and virulence in <i>Ralstonia pseudosolanacearum</i>

<i>Ralstonia pseudosolanacearum</i> (<i>Rp</i>s), which causes bacterial wilt disease of many crops, must integrate environmental signals to successfully transition from soil to its pathogenic niche in host plant xylem tissue. Mutating a gene encoding a putative sensing/signaling protein had little transcriptomic effect on <i>Rps</i> strain GMI1000 in culture. However, when the mutant grew in tomato, over 180 genes were differentially expressed relative to the wild type. The gene was therefore named <i>rprR</i> for <i><u>R</u>alstonia</i> <u>p</u>lant-<u>r</u>esponsive <u>r</u>egulator. <i>In planta</i>, the ∆<i>rprR</i> mutant dysregulated genes for diverse traits, including stress response, degradation of phenolic compounds, motility, attachment, and production of extracellular polysaccharide (EPS), which is a key bacterial wilt virulence factor. Quantifying <i>Rps</i> EPS by ELISA found increased levels in stems of plants infected with ∆<i>rprR</i> as compared to the wild type. Functional assays showed that ∆<i>rprR</i> is defective in attachment to tomato roots, colonization of tomato stems, and bacterial wilt virulence. In a rich medium, ∆<i>rprR</i> formed biofilm normally, but the mutant formed less biofilm in tomato stem homogenate and in tomato xylem sap under flow. This phenotype correlates with the mutant's altered expression of EPS biosynthetic genes and aberrant extracellular matrix. When grown in tomato stem homogenate, ∆<i>rprR</i> produced 57% more of the bacterial signal cyclic di-GMP (c-di-GMP) than the wild type. This is consistent with the presence, in RprR, of predicted c-di-GMP-modulating domains. Together, these findings reveal that RprR, which is highly conserved across plant pathogenic <i>Ralstonia</i>, modulates several bacterial wilt virulence traits in response to the plant host.IMPORTANCEMembers of the <i>Ralstonia solanacearum</i> species complex (RSSC) cause bacterial wilt, a globally destructive disease of market and subsistence crops. Like other plant-associated microbes, bacteria in the RSSC must integrate a complex array of biotic and abiotic signals to successfully infect plant hosts. All RSSC genomes encode an unusual protein, termed RprR, that contains multiple sensing and signaling domains, including two putative modulators of the secondary messenger c-di-GMP. Deleting RprR in <i>Ralstonia pseudosolanacearum</i> affected many virulence properties, including production of biofilm and exopolysaccharide, and increased intracellular c-di-GMP levels, all in a strictly plant-dependent fashion. While c-di-GMP has been investigated in other plant pathogenic bacteria, this is the first report of its role in the RSSC. Most importantly, <i>rprR</i> was required for <i>Ralstonia</i> to effectively colonize plants and cause wilt disease. Thus, RprR is a plant-responsive sensor-regulator that controls pathogen adaptation to the host environment and virulence.