Environmental prevalence of toxigenic Vibrio cholerae O1 in Bangladesh coincides with V. cholerae non-O1 non-O139 genetic variants which overproduce autoinducer-2

Abstract

Prevalence of toxigenic Vibrio cholerae O1 in aquatic reservoirs in Bangladesh apparently increases coinciding with the occurrence of seasonal cholera epidemics. In between epidemics, these bacteria persist in water mostly as dormant cells, known as viable but non-culturable cells (VBNC), or conditionally viable environmental cells (CVEC), that fail to grow in routine culture. CVEC resuscitate to active cells when enriched in culture medium supplemented with quorum sensing autoinducers CAI-1 or AI-2 which are signal molecules that regulate gene expression dependent on cell density. V. cholerae O1 mutant strains with inactivated cqsS gene encoding the CAI-1 receptor has been shown to overproduce AI-2 that enhance CVEC resuscitation in water samples. Since V. cholerae non-O1 non-O139 (non-cholera-vibrios) are abundant in aquatic ecosystems, we identified and characterized naturally occurring variant strains of V. cholerae non-O1 non-O139 which overproduce AI-2, and monitored their co-occurrence with V. cholerae O1 in water samples. The nucleotide sequence and predicted protein products of the cqsS gene carried by AI-2 overproducing variant strains showed divergence from that of typical V. cholerae O1 or non-O1 strains, and their culture supernatants enhanced resuscitation of CVEC in water samples. Furthermore, prevalence of V. cholerae O1 in the aquatic environment was found to coincide with an increase in AI-2 overproducing non-O1 non-O139 strains. These results suggest a possible role of non-cholera vibrios in the environmental biology of the cholera pathogen, in which non-O1 non-O139 variant strains overproducing AI-2 presumably contribute in resuscitation of the latent pathogen, leading to seasonal cholera epidemics. Importance. Toxigenic Vibrio cholerae which causes seasonal epidemics of cholera persists in aquatic reservoirs in endemic areas. The bacteria mostly exist in a dormant state during inter-epidemic periods, but periodically resuscitate to the active form. The resuscitation is enhanced by signal molecules called autoinducers (AIs). Toxigenic V. cholerae can be recovered from water samples that normally test negative for the organism in conventional culture, by supplementing the culture medium with exogenous AIs. V. cholerae belonging to the non-O1 non-O139 serogroups which do not cause cholera are also abundant in natural waters, and they are capable of producing AIs. In this study we characterized V. cholerae non-O1 non-O139 variant strains which overproduce an autoinducer called AI-2, and found that the abundance of the cholera pathogen in aquatic reservoirs correlates with an increase in the AI-2 overproducing strains. Our results suggest a probable role of these variant strains in the environmental biology and epidemiology of toxigenic V. cholerae, and may lead to novel means for surveillance, prevention and control of cholera.