Quorum Sensing Regulators Are Required for Metabolic Fitness in Vibrio parahaemolyticus.

Quorum sensing (QS) is a process by which bacteria alter gene expression in response to cell density changes. In Vibrio species, at low cell density, the sigma 54-dependent response regulator LuxO is active and regulates the two QS master regulators AphA, which is induced, and OpaR, which is repressed. At high cell density the opposite occurs: LuxO is inactive, and therefore OpaR is induced while AphA is repressed. In Vibrio parahaemolyticus, a significant enteric pathogen of humans, the roles of these regulators in pathogenesis are less known. We examined deletion mutants of luxO, opaR, and aphA for in vivo fitness using an adult mouse model. We found that the luxO and aphA mutants were defective in colonization compared to levels in the wild type. The opaR mutant did not show any defect in vivo Colonization was restored to wild-type levels in a luxO opaR double mutant and was also increased in an opaR aphA double mutant. These data suggest that AphA is important and that overexpression of opaR is detrimental to in vivo fitness. Transcriptome sequencing (RNA-Seq) analysis of the wild type and luxO mutant grown in mouse intestinal mucus showed that 60% of the genes that were downregulated in the luxO mutant were involved in amino acid and sugar transport and metabolism. These data suggest that the luxO mutant has a metabolic disadvantage, which was confirmed by growth pattern analysis using phenotype microarrays. Bioinformatics analysis revealed OpaR binding sites in the regulatory region of 55 carbon transporter and metabolism genes. Biochemical analysis of five representatives of these regulatory regions demonstrated direct binding of OpaR in all five tested. These data demonstrate the role of OpaR in carbon utilization and metabolic fitness, an overlooked role in the QS regulon.

High-salt preadaptation of Vibrio parahaemolyticus enhances survival in response to lethal environmental stresses.

Adaptation to changing environmental conditions is an important strategy for survival of foodborne bacterial pathogens. Vibrio parahaemolyticus is a gram-negative seafoodborne enteric pathogen found in the marine environment both free living and associated with oysters. This pathogen is a moderate halophile, with optimal growth at 3% NaCl. Among the several stresses imposed upon enteric bacteria, acid stress is perhaps one of the most important. V. parahaemolyticus has a lysine decarboxylase system responsible for decarboxylation of lysine to the basic product cadaverine, an important acid stress response system in bacteria. Preadaptation to mild acid conditions, i.e., the acid tolerance response, enhances survival under lethal acid conditions. Because of the variety of conditions encountered by V. parahaemolyticus in the marine environment and in oyster postharvest facilities, we examined the nature of the V. parahaemolyticus acid tolerance response under high-salinity conditions. Short preadaptation to a 6% salt concentration increased survival of the wild-type strain but not that of a cadA mutant under lethal acid conditions. However, prolonged exposure to high salinity (16 h) increased survival of both the wild-type and the cadA mutant strains. This phenotype was not dependent on the stress response sigma factor RpoS. Although this preadaptation response is much more pronounced in V. parahaemolyticus, this characteristic is not limited to this species. Both Vibrio cholerae and Vibrio vulnificus also survive better under lethal acid stress conditions when preadapted to high-salinity conditions. High salt both protected the organism against acid stress and increased survival under -20°C cold stress conditions. High-salt adaptation of V. parahaemolyticus strains significantly increases survival under environmental stresses that would otherwise be lethal to these bacteria.