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Biofilm formation enhances the survival and persistence of the facultative human pathogen Vibrio cholerae in natural ecosystems and its transmission during seasonal cholera outbreaks. A major component of the V. cholerae biofilm matrix is the Vibrio polysaccharide (VPS), which is essential for development of three-dimensional biofilm structures. The vps genes are clustered in two regions, the vps-I cluster (vpsU, vpsA-K, VC0916-27) and the vps-II cluster (vpsL-Q, VC0934-39), separated by an intergenic region containing the rbm gene cluster that encodes biofilm matrix proteins. In-frame deletions of the vps clusters and genes encoding matrix proteins drastically altered biofilm formation phenotypes. To determine which genes within the vps gene clusters are required for biofilm formation and VPS synthesis, we generated in-frame deletion mutants for all the vps genes. Many of these mutants exhibited reduced capacity to produce VPS and biofilms. Infant mouse colonization assays revealed that mutants lacking either vps clusters or rbmA (encoding secreted matrix protein RbmA) exhibited a defect in intestinal colonization compared to the wild-type. Understanding the roles of the various vps gene products will aid in the biochemical characterization of the VPS biosynthetic pathway and elucidate how vps gene products contribute to VPS biosynthesis, biofilm formation and virulence in V. cholerae. DOI 10.1099/mic.0.040196-0

Cholera is an acute intestinal infection caused by the bacterium Vibrio cholerae. In order for V. cholerae to cause disease, it must produce two virulence factors, the toxin-coregulated pilus (TCP) and cholera toxin (CT), whose expression is controlled by a transcriptional cascade culminating with the expression of the AraC-family regulator, ToxT. We have solved the 1.9 [Angstrom] resolution crystal structure of ToxT, which reveals folds in the N- and C-terminal domains that share a number of features in common with AraC, MarA, and Rob as well as the unexpected presence of a buried 16-carbon fatty acid, cis-palmitoleate. The finding that cis-palmitoleic acid reduces TCP and CT expression in V. cholerae and prevents ToxT from binding to DNA in vitro provides a direct link between the host environment of V. cholerae and regulation of virulence gene expression. AraC | crystal structure | pathogenesis | oleic acid | palmitoleic acid www.pnas.org/cgi/doi/10.1073/pnas.0915021107

Defined mutant libraries allow for efficient genome-scale screening and provide a convenient collection of mutations in almost any nonessential gene of interest. Here, we present a near-saturating transposon insertion library in Vibrio cholerae strain C6706, a clinical isolate belonging to the O1 El Tor biotype responsible for the current cholera pandemic. Automated sequencing analysis of 23,312 mutants allowed us to build a 3,156-member subset library containing a representative insertion in every disrupted ORF. Because uncharacterized mutations that affect motility have shown utility in attenuating V. cholerae live vaccines, we used this genome-wide subset library to define all genes required for motility and to further assess the accuracy and purity of the library. In this screen, we identified the hypothetical gene VC2208 (flgT) as essential for motility. Flagellated cells were very rare in a flgT mutant, and transcriptional analysis showed it was specifically stalled at the class Ill/IV assembly checkpoint of the V. cholerae flagellar regulatory system. Because FIgT is predicted to have structural homology to TolB, a protein involved in determining outer membrane architecture, and the sheath of the V. cholerae flagellum appears to be derived from the cell's outer membrane, FIgT may play a direct role in flagellar sheath formation. cholera | flagella | flgT | mariner

A study of epidemiology and ecology of Vibrio cholerae O1 and O139 was carried out in two coastal areas, Bakerganj and Mathbaria, Bangladesh where cholera occurs seasonally. The results of bi-weekly clinical study employing culture methods, direct and enrichment culture, colony blot hybridization and direct fluorescent-antibody methods showed that cholera is endemic in both Bakerganj and Mathbaria and that V. cholerae O1, O139, and nonO1/nonO139 are autochthonous to aquatic environment.

The emergence of Vibrio cholerae O139 Bengal during 1992-1993 was associated with large epidemics of cholera in India and Bangladesh and, initially, with a total displacement of the existing V. cholerae 01 strains. However, the 01 strains reemerged in 1994 and initiated a series of disappearance and reemergence of either of the two serogroups that was associated with temporal genetic and phenotypic changes sustained by the strains. Since the initial emergence of the 0139 vibrios, new variants of the pathogen derived from multiple progenitors have been isolated and characterized. The clinical and epidemiological characteristics of these strains have been studied. Rapid genetic reassortment in 0139 strains appears to be a response to the changing epidemiology of V. cholerae 01 and also a strategy for persistence in competition with strains of the 01 serogroup. The emergence of V. cholerae 0139 has provided a unique opportunity to witness genetic changes in V. cholerae that may be associated with displacement of an existing serogroup by a newly emerging one and, thus, provide new insights into the epidemiology of cholera. The genetic changes and natural selection involving both environmental and host factors are likely to influence profoundly the genetics, epidemiology, and evolution of toxigenic V. cholerae, not only in the Ganges Delta region of India and Bangladesh, but also in other areas of endemic and epidemic cholera.

Based on results of ecological studies demonstrating that Vibrio cholerae, the etiological agent of epidemic cholera, is commensal to zooplankton, notably copepods, a simple filtration procedure was developed whereby zooplankton, most phytoplankton, and particulates >20 [micro]m were removed from water before use. Effective deployment of this filtration procedure, from September 1999 through July 2002 in 65 villages of rural Bangladesh, of which the total population for the entire study comprised [approximately equal to] 133,000 individuals, yielded a 48% reduction in cholera (P < 0.005) compared with the control.

Results demonstrate that the new CTX Calcutta prophage is capable of giving rise to infectious cholera virus and it is differentiated from the other already existing prophage in the nucleotide sequences of the repressor genes.