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Applied and Environmental Microbiology, September 2000, p. 4022-4028, Vol. 66, No. 9
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Virulence Genes in Environmental Strains of Vibrio cholerae

Soumen Chakraborty,1 Asish K. Mukhopadhyay,1 Rupak Kumar Bhadra,2 Amar Nath Ghosh,1 Rupak Mitra,1 Toshio Shimada,3 Shinji Yamasaki,1,4 Shah M. Faruque,5 Yoshifumi Takeda,3 Rita R. Colwell,6,7,* and G. Balakrish Nair1

National Institute of Cholera and Enteric Diseases, Beliaghata, Calcutta 700 010,1 and Indian Institute of Chemical Biology, Calcutta 700 032,2 India; National Institute of Infectious Diseases, 1-23-1 Toyama,3 and Research Institute, International Medical Center of Japan,4 Shinjuku-ku, Tokyo 162, Japan; International Centre for Diarrhoeal Disease Research, Bangladesh, Dhaka-1212, Bangladesh5; and Center of Marine Biotechnology, University of Maryland Biotechnology Institute, Baltimore, Maryland 21202,6 and Department of Cell and Molecular Biology, University of Maryland, College Park, Maryland 207427

Received 9 March 2000/Accepted 21 June 2000

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.

* Corresponding author. Mailing address: Center of Marine Biotechnology, University of Maryland Biotechnology Institute, 701 East Pratt Street, Baltimore, MD 21202. Phone: (703) 306-1000. Fax: (703) 306-0109. E-mail: colwell{at}umbi.umd.edu.

Applied and Environmental Microbiology, September 2000, p. 4022-4028, Vol. 66, No. 9
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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