Coordinated regulation of virulence by quorum sensing and motility pathways during the initial stages of Vibrio cholerae infection

Abstract

Pathogenic bacteria, such as Vibrio cholerae, must be capable of adapting to diverse living conditions, especially when transitioning from life in environmental reservoirs to life in a host. The abilities to sense arrival at a site suitable for colonization or infection and to respond with appropriate alterations in gene expression are crucial for a pathogen's success. Recently, we have shown that V. cholerae is able to recognize that it has reached its colonization site in the small intestine by sensing breakage of its flagellum as it penetrates the mucosal layer overlaying the intestinal epithelium. Flagellar loss results in the release of the anti-sigma factor FlgM and subsequent activation of the alternative sigma-factor FliA. FliA represses the quorum sensing-controlled transcriptional regulator, HapR, allowing increased expression of virulence factors such as Cholera Toxin (CT) and the Toxin Coregulated Pilus (TCP). In this way, the de-repression of virulence factor expression coincides with the arrival of bacteria at the site of infection at the intestinal mucosa. Our work reveals an interesting interplay between motility and quorum sensing signaling pathways to precisely time virulence gene expression during colonization.

Keywords: FlgM; Vibrio cholerae; motility; mucosal penetration; quorum sensing; σ28.

Figure 1

Repression of HapR During Mucosal Penetration. Due to the harsh conditions of the upper digestive tract, V. cholerae reaches the small intestine in relatively low numbers. At low cell density, the quorum sensing response regulator, LuxO, is phosphorylated and represses HapR indirectly via small regulatory RNAs. As the bacteria penetrate the mucus covering the intestinal epithelial cells, their flagella break, FlgM is secreted, and FliA is activated, further repressing HapR. Since HapR represses virulence factor (CT and TCP) expression, these mechanisms of HapR repression during mucosal penetration serve to increase virulence gene expression.

Figure 2

Quorum Sensing Regulation in vivo. Approximately 10⁶ V. cholerae containing the hapR-Km^r chromosomal reporter were orally inoculated into infant CD-1 mice. At various time points, mouse intestines were homogenized, treated with or without kanamycin (500 µg/ml) for 10 min, and plated on LB plates. The number of colonized bacteria (dashed line) was calculated based on the number of colony-forming units (CFU) recovered from samples not treated with kanamycin. Expression of hapR (solid line) was defined as the percentage of Km-surviving cells out of total CFU. Results are means from experiments with three mice ± standard deviations.