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Molecular basis of the Competitive Exclusion of Vibrio Cholerae Classical Biotype by the el Tor Biotype

IR@IICB: CSIR-Indian Institute of Chemical Biology, Kolkata


Field	Value

Title	Molecular basis of the Competitive Exclusion of Vibrio Cholerae Classical Biotype by the el Tor Biotype

Creator	Pradhan, Subhra

Subject	Infectious Diseases and Immunology

Description	Vibrio cholerae is the causal organism of the diarrheal disease cholera. The first six pandemics of cholera occurred between 1817 and 1961 and were caused by classical biotype. In 1961, the El Tor biotype emerged in Indonesia to cause the ongoing seventh cholera pandemic and within a decade displaced the classical biotype as the causal organism of epidemic cholera. Such a rapid displacement of an existing pathogen by a newly emerged one was a unique event in bacterial epidemiology. Despite the accumulation of several facts about the molecular differences between the El Tor and the classical biotypes, the genetic basis of this phenomenon is still unknown. In this present study attempts have been made to elucidate the nature of fitness gained by the El Tor biotype over the classical biotype in cocultures and the molecular basis of exclusion of the classical biotype in cocultures with the El Tor biotype. We demonstrate that when strains of the V. cholerae El Tor and classical biotype were cocultured in standard LB medium, the El Tor strains had a competitive growth advantage over the classical biotype starting from the late stationary phase and could eventually take over the population. The possibility of nutrients being present in the stationary phase culture filtrates of the classical biotype stationary phase cultures were next examined. So, when the El Tor biotype cells were grown in the conditioned medium from the stationary phase classical biotype cultures it derived a marked growth advantage. Next, it was observed that indeed the classical biotype produces extracellular protease(s) in the stationary phase, and the amounts of amino acids and small peptides in the late stationary phase culture filtrates of the classical biotype were higher than those in the corresponding culture filtrates of the El Tor biotype. The El Tor biotype cells could utilize the amino acids more efficiently than the classical biotype cells under the alkaline pH of the stationary phase cultures. The growth advantage of the El Tor biotype was also observed in vivo using the ligated rabbit ileal loop and infant mouse animal models. Subsequently the nature of competitive exclusion of the classical biotype cells in the cocultures with El Tor biotype was investigated. It was observed that when the El Tor and classical biotypes were cocultured in standard laboratory medium a precipitous decline in colony forming units (CFU) of the classical biotype occurred in a contact dependent manner. Several lines of evidence including DNA release, microscopy and flow cytometric analysis indicated that the drastic reduction in CFU of the classical biotype in cocultures was not accompanied by lysis, although when the classical biotype was grown individually in monocultures, lysis of the cells occurred concomitant with decrease in CFU starting from late stationary phase. Furthermore, uptake of a membrane potential sensitive dye and protection of genomic DNA from extracellular DNases strongly suggested that the classical biotype cells in cocultures retained viability. These results suggest that coculturing the classical biotype with the El Tor biotype protects the former from lysis allowing the cells to remain viable in spite of the loss of culturability. The stationary phase sigma factor RpoS may have a role in the loss of culturability of the classical biotype in cocultures. Although competitive exclusion of closely related strains has been reported for several bacterial species, conversion of the target bacterial population to the viable non-culturable state has not been demonstrated previously and may have important implications in the evolution and epidemiology of bacterial strain.

Date	2013

Type	Thesis NonPeerReviewed

Format	application/pdf

Identifier	http://www.eprints.iicb.res.in/2002/1/Thesis_Subhra_Pradhan.pdf Pradhan, Subhra (2013) Molecular basis of the Competitive Exclusion of Vibrio Cholerae Classical Biotype by the el Tor Biotype. PhD thesis, Calcutta University.

Relation	http://www.eprints.iicb.res.in/2002/