Fis Connects Two Sensory Pathways, Quorum Sensing and Surface Sensing, to Control Motility in Vibrio parahaemolyticus

Abstract

Factor for inversion stimulation is a global regulator that is highly expressed during exponential phase growth and undetectable in stationary phase growth. Quorum sensing (QS) is a global regulatory mechanism that controls gene expression in response to changes in cell density and growth phase. In Vibrio parahaemolyticus, a marine species and a significant human pathogen, the QS regulatory sRNAs, Qrr1 to Qrr5, are expressed during exponential growth and negatively regulate the high cell density QS master regulator OpaR. OpaR is a positive regulator of capsule polysaccharide (CPS) formation, which is required for biofilm formation, and is a repressor of lateral flagella required for swarming motility. In V. parahaemolyticus, we show that Fis is a positive regulator of the qrr sRNAs expression. In an in-frame fis deletion mutant, qrr expression was repressed and opaR expression was induced. The Δfis mutant produced CPS and biofilm, but swarming motility was abolished. Also, the fis deletion mutant was more sensitive to polymyxin B. Swarming motility requires expression of both the surface sensing scrABC operon and lateral flagella laf operon. Our data showed that in the Δfis mutant both laf and scrABC genes were repressed. Fis controlled swarming motility indirectly through the QS pathway and directly through the surface sensing pathway. To determine the effects of Fis on cellular metabolism, we performed in vitro growth competition assays, and found that Δfis was outcompeted by wild type in minimal media supplemented with intestinal mucus as a sole nutrient source. The data showed that Fis positively modulated mucus components L-arabinose, D-gluconate and N-acetyl-D-glucosamine catabolism gene expression. In an in vivo colonization competition assay, Δfis was outcompeted by wild type, indicating Fis is required for fitness. Overall, these data demonstrate a global regulatory role for Fis in V. parahaemolyticus that includes QS, motility, and metabolism.

A class IV adenylate cyclase CyaB is required for capsule polysaccharide production and biofilm formation inVibrio parahaemolyticus

“…The Δ cyaA and Δ cyaB mutants grew similar to wild type producing typical budding swarming cells as previously shown for this species ( Fig. 6B ) (43–45). The WT, Δ cyaA and Δ cyaB swarming colonies were examined further to show that the swarming colony structural differences were not due to mutants forming.…”

“…Biofilm assays were performed as previously described (39, 40, 57). Briefly, overnight cultures were used to inoculate strip wells in a 1:40 dilution in LBS.…”

“…Capsule polysaccharide (CPS) formation assays were conducted as previously described (43)(44)(45). Wild type and mutant strains were grown on HI media containing 1.5% agar, 2% NaCl, 2.5 mM CaCl2, and 0.25% Congo red dye for 48 h at 30°C.…”

Gene Regulatory Network of the Noncoding RNA Qrr5 Involved in the Cytotoxicity of Vibrio parahaemolyticus during Infection

“…It has been found that Qrr1-5 negatively regulate the high cell density quorum-sensing regulator OpaR , which is required for biofilm formation and acts as a repressor of swarming motility [ 15 , 33 ]. However, we found that the cytotoxicity of Caco-2 cells increased with increasing post-infection time, and Qrr5 was significantly upregulated at different post-infection time points compared to Qrr1 – 4 ( Figure 1 a,b).…”

“…Fis is a regulator that negatively regulates the expression of the quorum-sensing master regulator OpaR and inhibits capsule polysaccharide (CPS) and biofilm formation, implying that Fis plays a direct role in the quorum sensing, biofilm formation, and metabolism of V. parahaemolyticus . Interestingly, when Fis binds directly to the regulatory regions of Qrr genes, it acts as a positive regulator [ 15 ].…”