Virulence Genes in Environmental Strains of Vibrio cholerae

Abstract

The virulence of a pathogen is dependent on a discrete set of genetic determinants and their well-regulated expression. The ctxAB and tcpA genes are known to play a cardinal role in maintaining virulence in Vibrio cholerae, and these genes are believed to be exclusively associated with clinical strains of O1 and O139 serogroups. In this study, we examined the presence of five virulence genes, including ctxAB and tcpA, as well as toxR and toxT, which are involved in the regulation of virulence, in environmental strains of V. cholerae cultured from three different freshwater lakes and ponds in the eastern part of Calcutta, India. PCR analysis revealed the presence of these virulence genes or their homologues among diverse serotypes and ribotypes of environmental V. cholerae strains. Sequencing of a part of the tcpA gene carried by an environmental strain showed 97.7% homology to the tcpA gene of the classical biotype of V. cholerae O1. Strains carrying the tcpA gene expressed the toxin-coregulated pilus (TCP), demonstrated by both autoagglutination analysis and electron microscopy of the TCP pili. Strains carrying ctxAB genes also produced cholera toxin, determined by monosialoganglioside enzyme-linked immunosorbent assay and by passage in the ileal loops of rabbits. Thus, this study demonstrates the presence and expression of critical virulence genes or their homologues in diverse environmental strains of V. cholerae, which appear to constitute an environmental reservoir of virulence genes, thereby providing new insights into the ecology of V. cholerae.Vibrio cholerae is known to be an autochthonous inhabitant of brackish waters and estuarine systems (4, 13). Among the 193 currently recognized O serogroups of V. cholerae (43), only serogroups O1 and O139 have caused epidemics of cholera. The other serogroups of V. cholerae, collectively referred to as non-O1 non-O139 serogroups, have not been associated with epidemics but can cause sporadic diarrhea (30) and are ubiquitously distributed in the aquatic environment (22,26). This sharp distinction between serogroups which can cause cholera and those which are not associated with cholera is related to the observation that more than 95% of the strains belonging to serogroups O1 and O139 produce cholera toxin (CT), which is central to the disease process. In contrast, more than 95% of the strains belonging to non-O1 non-O139 serogroups do not produce CT (15). Another important virulence factor of V. cholerae is the toxin-coregulated pilus (TCP), which is an adhesin that is coordinately regulated with CT production (39). TCP is the only V. cholerae pilus that has been demonstrated to date to have a role in colonization of the gut mucosa of humans (9) and of infant mice (39), the latter being an experimental cholera model.It has been presumed that CT and TCP are exclusively associated with clinical strains of V. cholerae, notably those belonging to serogroups O1 and O139, whereas reports on the incidence of CT among environmental strains of V. cholerae are rare (24). Sim...

Characterization of pathogenicity island prophage in clinical and environmental strains of Vibrio cholerae

“…2/40 (5) Vibrio cholerae pathogenicity island EVC4 was considered as being of tcpA 2 /ctxB + genotype supporting the suggestion that not all CT positive V. cholera strains are TCP positive, since TCP is known to be receptor for CTX phage (Chakraborty et al, 2000). The findings reported here are supported by those of Karaolis et al (2001) who demonstrated that the central region of the VPI of tcpA contains most of the divergence between the VPIs of the 6th (classic) and the 7th (El Tor) pandemic strains.…”

“…For autoagglutination, all strains were grown in LB (pH 6.8) supplemented with 1 % (w/v) NaCl and incubated at 37 uC for 18 h (Chakraborty et al, 2000).…”

Detection of ctx gene positive non-O1/non-O139 V. cholerae in shrimp aquaculture environments

“…A DNA probe study showed that a small percentage of environmental strains of V. cholerae non-O1 have the ctx gene (Nair et al 1988). Virulence genes including ctxAB were found among environmental strains from Calcutta, India (Chakraborty et al 2000). Occurrence of ctxA was found among 10% on non-O1/non-O139 environmental isolates from Brazil (Rivera et al 2001).…”

Environmental Surveillance for Toxigenic Vibrio cholerae in Surface Waters of Haiti

“…1,36 In other countries, virulence genes, including tcpA, have been found in V. cholerae non-O1/O139 from the environment, notably in cholera-endemic areas. 37,38 Although initial PCR results suggested these strains carried El Tor and classical tcpA alleles, sequence determination of the tcpA gene and phylogenetic analysis showed that the alleles were related to (but different from) the prototypical alleles (Supplemental Figure S2). The ctxA-negative isolates were also positive by PCR for different alleles of rstR, suggesting they could each possess a different variant of pre-CTX prophage (the CTX prophage devoid of the ctxAB genes).…”