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Background: The Twin-arginine translocation (Tat) system serves to translocate folded proteins, including periplasmic enzymes that bind redox cofactors in bacteria. The Tat system is also a determinant of virulence in some pathogenic bacteria, related to pleiotropic effects including growth, motility, and the secretion of some virulent factors. The contribution of the Tat pathway to Vibrio cholerae has not been explored. Here we investigated the functionality of the Tat system in V. cholerae, the etiologic agent of cholera. Results: In V. cholerae, the tatABC genes function in the translocation of TMAO reductase. Deletion of the tatABC genes led to a significant decrease in biofilm formation, the ability to attach to HT-29 cells, and the ability to colonize suckling mouse intestines. In addition, we observed a reduction in the output of cholera toxin, which may be due to the decreased transcription level of the toxin gene in tatABC mutants, suggesting an indirect effect of the mutation on toxin production. No obvious differences in flagellum biosynthesis and motility were found between the tatABC mutant and the parental strain, showing a variable effect of Tat in different bacteria. Conclusion: The Tat system contributes to the survival of V. cholerae in the environment and in vivo, and it may be associated with its virulence. [ABSTRACT FROM AUTHOR]

Receptor recognition is a key step in the initiation of phage infection. Previously, we found that VP3, the T7 family phage of the Vibrio cholerae serogroup O1 biotype El Tor, can adsorb the core oligosaccharide (OS) of lipopolysaccharides of V. cholerae. However, some wildtype strains of V. cholerae possessing the intact OS gene cluster still have VP3 binding but are resistant to VP3 infection. Moreover, an OS gene-deletion mutant still exhibits weak VP3 binding, suggesting multiple factors are possibly involved in VP3 binding to V. cholerae. Here, we report that the outer-membrane protein TolC of V. cholerae is involved in the host adsorption of VP3. We observed that TolC directly interacts with the VP3 tail fiber protein gp44 and its C-terminal domains, and we also found that three amino acid residues in the outside loops of TolC, at positions 78, 290, and 291, are critical for binding to gp44. Among the VP3-resistant wildtype V. cholerae strains, frequent amino acid residue mutations were observed in the loops around the sites 78, 290, and 291, which were predicted to be exposed to the cell surface. These findings reveal a co-receptor-binding mechanism for VP3 infection of V. cholerae and that both outer-membrane TolC and OS are necessary for successful VP3 infection of V. cholerae. We conclude that mutations on the outside loops of the receptor may confer V. cholerae strains with VP3 phage resistance, enabling these strains to survive in environments containing VP3 or related phages. [ABSTRACT FROM AUTHOR]

The cholera toxin genes of Vibrio cholerae are encoded by CTXΦ, a lysogenic bacteriophage. Infection with this phage plays a determinant role in toxigenicity conversion and the emergence of new clones of pathogenic V. cholerae. Multiple phage alleles, defined by sequence types of the repressor gene rstR, have been found, showing the divergence of phage genomes. Pre-CTXΦ, which is characterized by the absence of toxin genes, is predicted to be the precursor of CTXΦ. We have found a new pre-CTXΦ prophage genome (named pre-CTXZJΦ for its novel rstR allele) in nontoxigenic V. cholerae O1 isolates that were obtained during surveillance of the estuary water of the Zhujiang River. A novel hybrid genome of the helper phage RS1 was identified in an environmental strain carrying pre-CTXZJΦ in this study. The chromosomal integration and genomic arrangement of pre-CTXZJΦ and RS1 were determined. The RS2 of pre-CTXZJΦ was shown to have a function in replication, but it seemed to have lost its ability to integrate. The RstR of pre-CTXZJΦ exerted the highest repression of its own rstA promoter compared to other RstRs, suggesting rstR-specific phage superinfection immunity and potential coinfection with other pre-CTXΦ/CTXΦ alleles. The environmental strain carrying pre-CTXZJΦ could still be infected by CTXETΦ, the most common phage allele in the strains of the seventh cholera pandemic, suggesting that this nontoxigenic clone could potentially undergo toxigenicity conversion by CTXΦ infection and become a new toxigenic clone despite already containing the pre-CTXΦ prophage. [ABSTRACT FROM AUTHOR]

Background: Vibrio cholerae serogroup O1 has two major serotypes, Ogawa and Inaba, which may alternate among cholera epidemics. The rfbT gene is responsible for the conversion between the two serotypes. In this study, we surveyed the sequence variance of rfbT in the Ogawa and Inaba strains in China over a 48-year (1961-2008) period in which serotype shifts occurred among epidemic years. Results: Various mutation events including single nucleotide, short fragment insertions/deletions and transposases insertions, were found in the rfbT gene of the Inaba strains. Ectopically introducing an intact rfbT could overcome the mutations by converting the Inaba serotype to the Ogawa serotype, suggesting the effects of these mutations on the function of RfbT. Characteristic rfbT mutations were recognized in the Inaba strains among Inaba serotype dominant epidemic years which were separate from the Ogawa dominant epidemics. Three distinguishable mutation sites in rfbT between the classical and the El Tor biotype strains were identified and could serve as biotype-specific biomarkers. Conclusions: Our results provide a comprehensive picture of the rfbT gene mutations among the V. cholerae O1 strains in different epidemic periods, which could be further used as the tracing markers in clonality analysis and dissemination surveillance of the epidemic strains. [ABSTRACT FROM AUTHOR]

Bacteriophage VP4 is a lytic phage of the Vibrio cholerae serogroup O1, and it is used in phage subtyping of V. cholerae El Tor. Studies of phage infection mechanisms will promote the understanding of the basis of phage subtyping as well as the genetic differences between sensitive and resistant strains. In this study, we investigated the receptor that phage VP4 uses to bind to El Tor strains of V. cholerae and found that it infects strains through adsorbing the O antigen of V. cholerae O1. In some natural isolates that are resistant to VP4 infection, mutations were identified in the wb* cluster (O-antigen gene cluster), which is responsible for the biosynthesis of O antigen. Mutations in the manB, wbeE, and wbeU genes caused failure of adsorption of VP to these strains, whereas the observed amino acid residue mutations within wbeWand manChave no effect on VP4 infection. Additionally, although mutations in two resistant strains were found only in manB and wbeW, complementing both genes did not restore sensitivity to VP4 infection, suggesting that other resistance mechanisms may exist. Therefore, the mechanism of VP4 infection may provide a basis for subtyping the phage. Elaborate mutations of the O antigen may imbue V. cholerae strains with resistance to phage infection. [ABSTRACT FROM AUTHOR]

VP3 is a T7-like phage and was used as one of the typing phages in a phage-biotyping scheme that has been used for the typing of Vibrio cholerae O1 biotype El Tor. Here, we studied the receptor and other host genes of V. cholerae necessary for the lytic propagation of VP3. Six mutants resistant to VP3 infection were obtained from the random transposon insertion mutant bank of the sensitive strain N16961. The genes VC0229 and VC0231, which belong to the wav gene cluster encoding the core oligosaccharide (OS) region of lipopolysaccharide, were found to be interrupted by the transposon in five mutants, and the sixth mutant had the transposon inserted between the genes rhlB and trxA, which encode the ATP-dependent RNA helicase RhlB and thioredoxin, respectively. Gene complementation, transcription analysis, and the loss of VP3 sensitivity by the gene deletion mutants confirmed the relationship between VP3 resistance and VC0229, VC0231, and trxA mutation. The product of VP3 gene 44 (gp44) was predicted to be a tail fiber protein. gp44 could bind to the sensitive wild-type strain and the trxA mutant, but not to VC0229 and VC0231 mutants. The results showed that OS is a VP3 receptor on the surface of N16961, thioredoxin of the host strain is involved in the propagation of the phage, and gp44 is the tail fiber protein of VP3. This revealed the first step in the infection mechanism of the T7-like phage VP3 in V. cholerae. [ABSTRACT FROM AUTHOR]

Phage typing is used for the subtyping of clones of epidemic bacteria. In this study, we identified the outer membrane protein OmpW as the receptor for phage VP5, one of the typing phages for the Vibrio cholerae O1 El Tor biotype. A characteristic 11-bp deletion in ompW was observed in all epidemic strains resistant to VP5, suggesting that this mutation event can be used as a tracing marker in cholera surveillance. [ABSTRACT FROM AUTHOR]