Serogroup Conversion of Vibrio cholerae in Aquatic Reservoirs

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Abstract
The environmental reservoirs for Vibrio cholerae are natural aquatic habitats, where it colonizes the chitinous exoskeletons of copepod molts. Growth of V. cholerae on a chitin surface induces competence for natural transformation, a mechanism for intra-species gene exchange. The antigenically diverse O-serogroup determinants of V. cholerae are encoded by a genetically variable biosynthetic cluster of genes that is flanked on either side by chromosomal regions that are conserved between different serogroups. To determine whether this genomic motif and chitin-induced natural transformation might enable the exchange of serogroup-specific gene clusters between different O serogroups of V. cholerae, a strain of V. cholerae O1 El Tor was co-cultured with a strain of V. cholerae O139 Bengal within a biofilm on the same chitin surface immersed in seawater, and O1-to-O139 transformants were obtained. Serogroup conversion of the O1 recipient by the O139 donor was demonstrated by comparative genomic hybridization, biochemical and serological characterization of the O-antigenic determinant, and resistance of O1-to-O139 transformants to bacteriolysis by a virulent O1-specific phage. Serogroup conversion was shown to have occurred as a single-step exchange of large fragments of DNA. Crossovers were localized to regions of homology common to other V. cholerae serogroups that flank serogroup-specific encoding sequences. This result and the successful serogroup conversion of an O1 strain by O37 genomic DNA indicate that chitin-induced natural transformation might be a common mechanism for serogroup conversion in aquatic habitats and for the emergence of V. cholerae variants that are better adapted for survival in environmental niches or more pathogenic for humans. The reservoirs of Vibrio cholerae are aquatic environments, where it attaches to the chitin-containing shells of small crustaceans. Chitin serves as a nutrient for V. cholerae and it induces natural transformation, a process by which it acquires new genes from other microbes in the same habitat. The most compelling consequence of a V. cholerae gene acquisition event occurred in 1992 when a vast cholera epidemic erupted in India and Bangladesh and spread through Asia. Genetic analysis showed that this outbreak was due to the acquisition of a gene cluster that converted the ancestral V. cholerae O1 El Tor serogroup to an entirely new serogroup, designated O139 Bengal. This report shows that acquisition of the O139 gene cluster by an O1 El Tor strain can be mediated by natural transformation and that this can occur within a community of bacteria living on a chitin surface. The O139 derivatives of this transformation event were not killed by bacteriophages that attack O1 strains, explaining in part why O139 strains have replaced O1 strains in some Asian water sources. These results also illustrate how a combination of genetic and ecological factors can lead to the emergence of new pathogenic microbes in environmental reservoirs.