Molecular Characterization of Vibrio cholerae O139 Bengal Isolated from Water and the Aquatic Plant Eichhornia crassipes in the River Ganga, Varanasi, India

Abstract

A collection of ten strains of Vibrio cholerae O139, comprising six isolates from Eichhornia crassipes, two from water of the River Ganga, and one each from a well and a hand pump, were characterized. All the strains carried the CTX genetic element (ctxA, zot, and ace) except for the st gene and carried structural and regulatory genes for toxin-coregulated pilus (tcpA, tcpI, and toxR), adherence factor (ompU), and accessory colonization factor (acfB); all produced cholera toxin (CT). These strains were resistant to trimethoprim, sulfamethoxazole, streptomycin, and to the vibriostatic agent pteridine. Results obtained by ribotyping and enterobacterial repetitive intergenic consensus sequence-PCR fingerprint analysis indicate that multiple clones of toxigenicpathogenic V. cholerae O139 were present in the aquatic environment.Until 1992, Vibrio cholerae belonging to serogroup O1 was considered to be the only causative agent of epidemic cholera (2). However, V. cholerae non-O1 serogroups were associated mostly with sporadic cases of diarrhea and extraintestinal infections (4, 24). It was found recently that a highly epidemic form of cholera-like disease on the Indian subcontinent was strongly associated with non-O1 strains of V. cholerae designated V. cholerae O139 Bengal (1, 21, 25), which subsequently spread to the different parts of the world (1, 6, 11). Studies of V. cholerae from environmental surface water indicated that O139 Bengal, like O1 vibrios and non-O1, non-O139 vibrios, may survive better in the aquatic environment in association with aquatic plants (13,14,26) and that environmental water may be a reservoir for infectious V. cholerae O139 (14, 29).In the present study, we used molecular techniques to analyze V. cholerae O139 Bengal isolated between January 1992 and September 1994 from water and the aquatic plant Eichhornia crassipes (water hyacinth) from the river Ganga, Varanasi, India, to study genomic diversity and to examine the presence of virulence and regulatory genes, pathogenicity islands, and antibiotic resistance. Results obtained by ribotyping, enterobacterial repetitive intergenic consensus sequence (ERIC)-PCR, PCR analysis, and antibiotic susceptibility testing indicated that the cholera outbreaks caused by O139 vibrios between 1992 and 1994 were caused by toxigenic-pathogenic and antibiotic-resistant V. cholerae O139 strains identical to that found in the aquatic environment.Bacterial strains. A total of 10 strains of V. cholerae O139 were included in this study. Six isolates (EC1, EC2, EC3, EC6, EC7, and EC8) were from the aquatic plant Eichhornia crassipes and were collected between 28 January and 6 April 1992; two (GW3 and GW4) were from surface water of the River Ganga and were collected between 3 August and 29 August 1993 from different ghats on the bank of the River Ganga in Varanasi; and one each was from a well (WO4, collected on 10 September 1994 from Bhadaini, Varanasi) and a hand pump (HP11, collected on 5 August 1992 from Assi, Varanasi). The samples from the aquatic plants ...

Adhesion of Vibrio cholerae to Granular Starches

“…It has been speculated that the appearance of high numbers of V. cholerae is correlated to algal blooms (25), yet algae are known to contain large quantities of starch. In addition, V. cholerae could be isolated from plants (Eichhornia crassipes) found in river water (5). Therefore, the ability to utilize starch as well as the ability to adhere to it can contribute to the utilization of this food source in the aquatic environments.…”

Application of duplex-PCR in rapid and reliable detection of toxigenic Vibrio cholerae in water samples in Thailand

“…The conventional culture method, which is routinely used for the isolation of V. cholerae, is time-consuming and may not be suitable for cells that are VBNC (Huq et al, 1995;Islam et al, 1993bIslam et al, , 1994bKapley and Purohit, 2001). Recent molecular techniques such as polymerase chain reaction (PCR) offer both a rapid and reliable method for detecting toxigenic V. cholerae occurring in the aquatic environment (Bhanumathi et al, 2003;Chakraborty et al, 1999;Kapley and Purohit, 2001). The ctxAB and tcpA are the genes known to play key roles in the virulence of V. cholerae especially of serogroups O1 and O139 (Faruque et al, 1997;Herrington et al, 1988;Kovach et al, 1994).…”

Multiplex PCR for detection of antibiotic resistance genes and the SXT element: application in the characterization of Vibrio cholerae

“…As the SXT genetic element plays a role in the acquisition of antibiotic resistance, it is important to look for the presence of the sulII (encoding Sul resistance), dfrA1 (O1-specific Tmp resistance), dfr18 (O139-specific Tmp resistance) and strB (Str B resistance) genes and the SXT element (Bhanumathi et al, 2003;Falbo et al, 1999;Hochhut et al, 2001) in V. cholerae strains. To trace the changes in antibiotic resistance, we developed a multiplex PCR for the detection of antibiotic resistance genes and the SXT element and used it to characterize V. cholerae strains.…”