Novel multiple mutations in the topoisomerase gene of Haitian variant Vibrio cholerae O1.
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ABSTRACT:
SUBMITTER: Divya MP
PROVIDER: S-EPMC4136007 | biostudies-literature | 2014 Aug
REPOSITORIES: biostudies-literature
Publications
Novel multiple mutations in the topoisomerase gene of Haitian variant Vibrio cholerae O1.
Divya M P MP Sivakumar K C KC Sarada Devi K L KL Remadevi S S Thomas Sabu S
Antimicrobial agents and chemotherapy 20140527 8
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Project description:Cholera continues to be an important public health concern in developing countries where proper hygiene and sanitation are compromised. This severe diarrheal disease is caused by the Gram-negative pathogen Vibrio cholerae belonging to serogroups O1 and O139. Cholera toxin (CT) is the prime virulence factor and is directly responsible for the disease manifestation. The ctxB gene encodes cholera toxin B subunit (CTB) whereas the A subunit (CTA) is the product of ctxA gene. Enzymatic action of CT depends on binding of B pentamers to the lipid-based receptor ganglioside GM1. In recent years, emergence of V. cholerae Haitian variant strains with ctxB7 allele and their rapid spread throughout the globe has been linked to various cholera outbreaks in Africa and Asia. These strains produce classical type (WT) CTB except for an additional mutation in the signal sequence region where an asparagine (N) residue replaces a histidine (H) at the 20th amino acid position (H20N) of CTB precursor (pre-CTB). Here we report that Haitian variant V. cholerae O1 strains isolated in Kolkata produced higher amount of CT compared to contemporary O1 El Tor variant strains under in vitro virulence inducing conditions. We observed that the ctxB7 allele, itself plays a pivotal role in higher CT production. Based on our in silico analysis, we hypothesized that higher accumulation of toxin subunits from ctxB7 allele might be attributed to the structural alteration at the CTB signal peptide region of pre-H20N CTB. Overall, this study provides plausible explanation regarding the hypertoxigenic phenotype of the Haitian variant strains which have spread globally, possibly through positive selection for increased pathogenic traits.
| S-EPMC7112172 | biostudies-literature
Project description:The world's worst cholera epidemic in Haiti (2010) coerced to trace the origin and dissemination of the causative agent Vibrio cholerae O1 for proper management of cholera. Sequence analysis of the Haitian strain showed several variations in the genes encoding cholera toxin B subunit (ctxB); toxin-co-regulated pilus (tcpA), repeat in toxins (rtxA), quinolone resistance-determining region (QRDR) of gyrase A (gyrA), rstB of RS element along with the change in the number of repeat sequences at the promoter region of ctxAB. Our earlier studies showed that variant tcpA (tcpA CIRS) and ctxB (ctxB7) first appeared in Kolkata during 2003 and 2006, respectively. The present study revealed that a variant rtxA was first isolated in Kolkata during 2004 and probably formed the genetic background for the emergence of the ctxB7 allele as we were unable to detect a single strain with the combination of El Tor rtxA and ctxB7. The variant gyrA was first time detected in Kolkata during 1994. The Kolkata strains contained four heptad repeats (TTTTGAT) in their CT promoter regions whereas Haitian strains carried 5 heptad repeats. Haitian strains had 3 nucleotide deletions at the rstB gene, which is a unique feature of the classical biotype strains. But the Kolkata strains did not have such deletion mutations in the rstB. Our study demonstrated the existence of some Haitian genetic traits in Kolkata isolates along with the dissimilarities in genomic content with respect to rstB and ctxAB promoter region. Finally, we conclude that Haitian variant strain may be evolved due to sequential event in the Indian subcontinent strain with some cryptic modification in the genome.
| S-EPMC4240540 | biostudies-literature
Project description:BackgroundIncrease in the number of multidrug resistant pathogens and the accompanied rise in case fatality rates has hampered the treatment of many infectious diseases including cholera. Unraveling the mechanisms responsible for multidrug resistance in the clinical isolates of Vibrio cholerae would help in understanding evolution of these pathogenic bacteria and their epidemic potential. This study was carried out to identify genetic factors responsible for multiple drug resistance in clinical isolates of Vibrio cholerae O1, serotype Ogawa, biotype El Tor isolated from the patients admitted to the Infectious Diseases Hospital, Kolkata, India, in 2009.Methodology/principal findingsOne hundred and nineteen clinical isolates of V. cholerae were analysed for their antibiotic resistance phenotypes. Antibiogram analysis revealed that majority of the isolates showed resistance to co-trimoxazole, nalidixic acid, polymixin B and streptomycin. In PCR, SXT integrase was detected in 117 isolates and its sequence showed 99% identity notably to ICEVchInd5 from Sevagram, India, ICEVchBan5 from Bangladesh and VC1786ICE sequence from Haiti outbreak among others. Antibiotic resistance traits corresponding to SXT element were transferred from the parent Vibrio isolate to the recipient E. coli XL-1 Blue cells during conjugation. Double-mismatch-amplification mutation assay (DMAMA) revealed the presence of Haitian type ctxB allele of genotype 7 in 55 isolates and the classical ctxB allele of genotype 1 in 59 isolates. Analysis of topoisomerase sequences revealed the presence of mutation Ser83 → Ile in gyrA and Ser85→ Leu in parC. This clearly showed the circulation of SXT-containing V. cholerae as causative agent for cholera in Kolkata.ConclusionsThere was predominance of SXT element in these clinical isolates from Kolkata region which also accounted for their antibiotic resistance phenotype typical of this element. DMAMA PCR showed them to be a mixture of isolates with different ctxB alleles like classical, El Tor and Haitian variants.
| S-EPMC3576390 | biostudies-literature
Project description:The presence of SXT/R391-related integrating conjugative elements (ICEs) in Vibrio cholerae O1 and non-O1/non-O139 isolated from clinical and environmental samples in Haiti in 2010 was studied. The main finding of this work was the identification of the novel ICEVchHai2 among closely related V. cholerae non-O1/non-O139 clinical strains. The mosaic structure of this element confirms the role of ICEs as efficient recombination systems whereby new genetic material can be acquired and exchanged, according V. cholerae strains new accessory functions.
| S-EPMC3999591 | biostudies-other
Project description: Not available
| S-EPMC3713811 | biostudies-literature
Project description:A PCR-based assay was developed to discriminate the classical, El Tor, and Haitian types of ctxB alleles. Our retrospective study using this newly developed PCR showed that Haitian ctxB first appeared in Kolkata during April 2006, and 93.3% of strains isolated during 2011 carried the new allele. Dendrogram analysis showed a pulsed-field gel electrophoresis (PFGE) pattern of the new variant strains isolated recently that was distinct from the PFGE pattern of the strains carrying classical ctxB that closely matched the 2006 to 2007 variant strains.
| S-EPMC3347119 | biostudies-literature
Project description:Vibrio cholerae O1 exists as two major serotypes, Inaba and Ogawa, which are associated with the O antigen of the lipopolysaccharide and are capable of unequal reciprocal interconversion. The 20-kilobase rfb regions encoding O-antigen biosynthesis in strains 569B (Inaba) and O17 (Ogawa) have been cloned in Escherichia coli K-12 and the nucleotide sequences have been determined. Besides several base substitutions and a small deletion in the 569B sequence relative to O17, there is a single nucleotide change resulting in a TGA stop codon within the gene for the 32-kDa RfbT protein. We have demonstrated that rfbT is responsible for serotype conversion (Inaba to Ogawa). The construction of a specific rfbT mutation in the Ogawa strain O17, and the ability of the gene from O17 to complement Inaba strains to Ogawa, confirmed rfbT as the gene required for the Ogawa serotype. By Southern hybridization and sequencing of PCR products of a number of strains, we have shown that the changes observed in one Inaba strain (569B) are not conserved in other Inaba strains. This may explain why some Inaba strains are able to convert to Ogawa whereas others are not. The protein encoded by rfbT has been identified and expressed in E. coli K-12 using a phage T7 expression system. Amino-terminal analysis of partially purified protein has identified the translational start of the protein. Primer extension studies have enabled the 5' end of the mRNA to be defined. It exists as a separate transcript from the rest of the rfb region, and the distinctive G + C content of rfbT suggests that it has been acquired from a non-Vibrio source.
| S-EPMC48702 | biostudies-other
Project description:A novel qnr determinant emerged in ciprofloxacin-resistant Vibrio cholerae O1 from the Amazon region of Brazil. This qnrVC1 was in a typical class 1 integron. Its attC showed 89% identity with V. parahaemolyticus superintegron repeats. Analysis showed V. cholerae O1 carrying qnrVC2 associated with a V. cholerae superintegron repeat.
| S-EPMC2600354 | biostudies-literature
Project description: Not available
| S-EPMC5088018 | biostudies-literature
Project description:The draft genome sequence of a non-O1 Vibrio cholerae strain, PS15, organized into 3,512 open reading frames within a 3.9-Mb genome, was determined. The PS15 genome sequence will allow for the study of the evolution of virulence and environmental adaptation in V. cholerae.
| S-EPMC3569316 | biostudies-literature