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Cholix toxin (ChxA) is a recently discovered exotoxin inVibrio cholerae which has been characterized as a third member of the eukaryotic elongation factor 2-specific ADP-ribosyltransferase toxins, in addition to exotoxin A ofPseudomonas aeruginosa and diphtheria toxin ofCorynebacterium diphtheriae . These toxins consist of three characteristic domains for receptor binding, translocation, and catalysis. However, there is little information about the prevalence ofchxA and its genetic variations and pathogenic mechanisms. In this study, we screened thechxA gene in a large number (n = 765) ofV. cholerae strains and observed its presence exclusively in non-O1/non-O139 strains (27.0%; 53 of 196) and not in O1 (n = 485) or O139 (n = 84). Sequencing of these 53chxA genes generated 29 subtypes which were grouped into three clusters designatedchxA I,chxA II, andchxA III.chxA I belongs to the prototype, whilechxA II andchxA III are newly discovered variants. ChxA II and ChxA III had unique receptor binding and catalytic domains, respectively, in comparison to ChxA I. Recombinant ChxA I (rChxA I) and rChxA II but not rChxA III showed variable cytotoxic effects on different eukaryotic cells. Although rChxA II was more lethal to mice than rChxA I when injected intravenously, no enterotoxicity of any rChxA was observed in a rabbit ileal loop test. Hepatocytes showed coagulation necrosis in rChxA I- or rChxA II-treated mice, seemingly the major target for ChxA. The present study illustrates the potential of ChxA as an important virulence factor in non-O1/non-O139V. cholerae , which may be associated with extraintestinal infections rather than enterotoxicity.

Novel Cholix Toxin Variants, ADP-Ribosylating Toxins in Vibrio cholerae Non-O1/Non-O139 Strains, and Their Pathogenicity