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Appl. Environ. Microbiol. doi:10.1128/AEM.00029-08
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Putative Virulence Traits and Pathogenicity of Vibrio cholerae non-O1, non-O139 Isolated from Surface Waters in Kolkata, India

Department of Biochemistry, University of Calcutta, 35 Ballygunge Circular Road, Kolkata 700 019, India; and National Institute of Cholera and Enteric Diseases, P-33 C. I. T. Road, Scheme-XM, Beliaghata, Kolkata 700 010, India

^* To whom correspondence should be addressed. Email: pkbbioc{at}caluniv.ac.in.

	Abstract

V. cholerae non-O1, non-O139 was isolated from natural surface waters from different sites sampled in diarrhea endemic zones in Kolkata, India. Twenty-one of these isolates were randomly selected and included for the characterization. The multi serogroup isolates were compared by their virulence traits with a group of clinical non-O1, non-O139 isolates from the same geographic area. Of the 21 environmental isolates, 6 and 14 strains belonged to Heiberg groups I and II respectively. Three of the environmental isolates showed resistance to O/129. All of the non-O1, non-O139 strains were positive for toxR, and except one environmental isolate, none of them were positive for tcpA in the PCR assay. None of the isolates were positive for genes encoding cholera toxin (ctxA), heat-stable toxin (est), heat-labile toxin (elt), and shiga toxin variants (stx) of Escherichia coli. Additionally, except for one environmental isolate (PC32), all were positive for the gene encoding El Tor hemolysin (hly). Culture supernatants of 86% (18 of 21) of environmental isolates showed a distinct cytotoxic effect to HeLa cells and some of these strains also produced cell rounding factor. Lipase, protease and cell-associated hemagglutination activities, and serum resistance properties of the environmental and clinical isolates did not differ much. However, 7 environmental isolates exhibited very high hemolytic activity (80 to 100%), while none of the clinical strains belonged to this group. Environmental isolates manifested three adherence patterns namely carpet-like, diffuse and aggregative adherence, and clinical isolates showed diffuse adherence on HeLa cells. Of the 11 environmental isolates tested for their enteropathogenic potential, eight (73%) induced positive fluid accumulation (FA 100) in mouse model and the reactivities of these isolates were comparable to those of clinical strains of non-O1, non-O139 and toxigenic O139. Comparison in counts of the colonized environmental and clinical strains in mouse intestine showed that the organism of both groups had similar colonizing efficiency. These findings indicate the presence of potentially pathogenic V. cholerae non-O1, non-O139 strains in surface waters of the studied sites in Kolkata.