Genetic diversity and virulence potential of environmental Vibrio cholerae population in a cholera-endemic area

Abstract

To understand the evolutionary events and possible selection mechanisms involved in the emergence of pathogenic Vibrio cholerae, we analyzed diverse strains of V. cholerae isolated from environmental waters in Bangladesh by direct enrichment in the intestines of adult rabbits and by conventional laboratory culture. Strains isolated by conventional culture were mostly (99.2%) negative for the major virulence gene clusters encoding toxincoregulated pilus (TCP) and cholera toxin (CT) and were nonpathogenic in animal models. In contrast, all strains selected in rabbits were competent for colonizing infant mice, and 56.8% of these strains carried genes encoding TCP alone or both TCP and CT. Ribotypes of toxigenic O1 and O139 strains from the environment were similar to pandemic strains, whereas ribotypes of non-O1 non-O139 strains and TCP ؊ nontoxigenic O1 strains diverged widely from the seventh pandemic O1 and the O139 strains. Results of this study suggest that (i) the environmental V. cholerae population in a cholera-endemic area is highly heterogeneous, (ii) selection in the mammalian intestine can cause enrichment of environmental strains with virulence potential, (iii) pathogenicity of V. cholerae involves more virulence genes than currently appreciated, and (iv) most environmental V. cholerae strains are unlikely to attain a pandemic potential by acquisition of TCP and CT genes alone. Because most of the recorded cholera pandemics originated in the Ganges Delta region, this ecological setting presumably favors extensive genetic exchange among V. cholerae strains and thus promotes the rare, multiple-gene transfer events needed to assemble the critical combination of genes required for pandemic spread.

Vibrio cholerae Strain Typing and Phylogeny Study Based on Simple Sequence Repeats

“…The environmental V. cholerae strains from non-O1, non-O139 serogroups are a possible natural reservoir of potentially new emerging epidemic strains (67,73). This assumption is supported by the finding that some of these environmental strains harbor virulence genes (23) and thus are likely to evolve into novel pathogenic strains by horizontal gene transfer (24,25). The emergence of new pathogenic V. cholerae strains requires not only an efficient, rapid, and accurate identification tool but also a means for determining genetic relationships among environmental and clinical isolates.…”

“…Coding (hypothetical protein) 2 d 0.50 VC1457- (7) 4 (AAATCAA) 4 Noncoding (cholera enterotoxin, A subunit) 5 d 0.58 VC1650- (9) 7 (GATAATCCA) 7 Coding (collagenase) 8 0.84 VCA0171- (6) 23 (TGCTGT) 23 Coding (hypothetical protein) 13 0.91 VCA0283- (6) ing of the 15 O1 and O139 ctxA (ϩ) isolates to three ST and the 14 environmental and 0139 ctxA (Ϫ) strains to 10 ST (Fig. 2a).…”

“…Two cocktails of three markers were chosen. Mix 1 consisted of VC0437-(7) 7 , VC1650-(9) 7 , and VCA0283-(6) 14 primers labeled with Hex, Fam, and Hex, respectively, and mix 2 consisted of VC0147-(6) 9 , VCA0171-(6) 23 , and VC1457-(7) 4 primers labeled with Fam, Hex, and Fam, respec- tively. The same alleles were observed in both multiplex and separate amplifications, supporting the use of the multiplex assay for high-throughput typing.…”

“…Primer concentrations for each mix were as follows. Mix 1 included VC0437-(7) 7 , VC1650-(9) 7 , and VCA0283-(6) 14 at final concentrations of 0.8, 0.6, and 1.4 M, respectively; mix 2 included VC0147-(6) 9 , VCA0171-(6) 23 , and VC1457-(7) 4 at final concentrations of 0.4 M. The annealing temperatures were 60 and 62°C for mix 1 and mix 2, respectively. Detection of specific genes by PCR.…”

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).…”