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Genomic and phenotypic characterization of Vibrio cholerae non-O1 isolates from a US Gulf Coast cholera outbreak.

Haley BJ, Choi SY, Grim CJ, Onifade TJ, Cinar HN, Tall BD, Taviani E, Hasan NA, Abdullah AH, Carter L, Sahu SN, Kothary MH, Chen A, Baker R, Hutchinson R, Blackmore C, Cebula TA, Huq A, Colwell RR - PLoS ONE (2014)

Bottom Line: From the 11 diagnosed cases, eight isolates of V. cholerae were isolated and their genomes were sequenced.Comparative genomics, phenotypic analyses, and a Caenorhabditis elegans model of infection for the isolates were conducted.This analysis coupled with isolation data of V. cholerae O75 and O141 suggests these strains may represent an underappreciated clade of cholera-causing strains responsible for significant disease burden globally.

View Article: PubMed Central - PubMed

Affiliation: Maryland Pathogen Research Institute, University of Maryland, College Park, Maryland, United States of America.

ABSTRACT
Between November 2010, and May 2011, eleven cases of cholera, unrelated to a concurrent outbreak on the island of Hispaniola, were recorded, and the causative agent, Vibrio cholerae serogroup O75, was traced to oysters harvested from Apalachicola Bay, Florida. From the 11 diagnosed cases, eight isolates of V. cholerae were isolated and their genomes were sequenced. Genomic analysis demonstrated the presence of a suite of mobile elements previously shown to be involved in the disease process of cholera (ctxAB, VPI-1 and -2, and a VSP-II like variant) and a phylogenomic analysis showed the isolates to be sister taxa to toxigenic V. cholerae V51 serogroup O141, a clinical strain isolated 23 years earlier. Toxigenic V. cholerae O75 has been repeatedly isolated from clinical cases in the southeastern United States and toxigenic V. cholerae O141 isolates have been isolated globally from clinical cases over several decades. Comparative genomics, phenotypic analyses, and a Caenorhabditis elegans model of infection for the isolates were conducted. This analysis coupled with isolation data of V. cholerae O75 and O141 suggests these strains may represent an underappreciated clade of cholera-causing strains responsible for significant disease burden globally.

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rstR sequences from V. cholerae CP1110.Sequences are aligned to their most similar homologs extracted from NCBI Genbank database. Nulceotide sequence identity is shown to the right of the last nucleotide aligned for each allele.
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pone-0086264-g002: rstR sequences from V. cholerae CP1110.Sequences are aligned to their most similar homologs extracted from NCBI Genbank database. Nulceotide sequence identity is shown to the right of the last nucleotide aligned for each allele.

Mentions: The V. cholerae FL Group isolates were determined to contain the full CTX phage encoding the cholera toxin, but the structure of this region was unresolved due to the limitations of assembly since ORFs were found on multiple contigs. For similar reasons, CTX phage copy number could not be resolved. A BLASTN analysis with V. cholerae N16961 and O395 as reference demonstrated the presence of regions homologous to VC1456 to VC1463 (VC0395_0512 to VC0395_0505 and VC0395_A1060 and VC0395_A1067 of V. cholerae O395) of the CTX phage (ctxB, ctxA, zot, ace, orfU,cep, rstB, rstA, and rstRClassical). To infer the biotype of the cholera toxin, PCR targeting the ctxB gene was employed and resulted in an amplicon for primers of targeting ctxBClassical. These PCR results are consistent with profiles of other clinically isolated V. cholerae strains on a global scale that suggest this cholera toxin biotype is the predominant biotype currently causing the majority of disease [34], [35]. Based on the genome sequence data, the CTX phage of the V. cholerae FL Group genomes were lacking the rstR gene of V. cholerae N16961 El Tor (VC1464), but did encode the rstR gene homologous to the one encoded in V. cholerae O395 Classical. To further investigate and confirm these in silico results, PCR targeting the rstR region was done and resulted in amplicons for the Calcutta, Environmental, and Classical biotypes, but not the El Tor biotype, an as-to-date uncommon combination. The rstR amplicons of CP1110 were subjected to Sanger sequencing and the resulting sequences were compared by BLASTN to the NCBI Genbank database for better interpretation of these results and each showed ≥99% nucleotide sequence similarity to Calcutta, Environmental, and Classical sequences (Figure 2). These amplicon sequences were compared with V. cholerae CP1110 reads by BLASTN to re-confirm their presence in the genome sequences. The rstR sequences from the V. cholerae FL Group were confirmed as Calcutta, Environmental, and Classical biotypes (Figure 2). The prototypical V. cholerae O1 El Tor strains encode rstREl Tor and ctxBEl Tor while Classical strains encode rstRClassical and ctxBClassical. Altered V. cholerae O1 El Tor strains which differ from prototypical El Tor strains in their rstR/ctxB types have recently been identified [24]. Data from this study further demonstrates the diversity of the CTX phage outside of the more frequently studied V. cholerae O1 strains and suggests many alleles of this phage can be associated with cholera. Cholera toxin expression was not assayed in this study.


Genomic and phenotypic characterization of Vibrio cholerae non-O1 isolates from a US Gulf Coast cholera outbreak.

Haley BJ, Choi SY, Grim CJ, Onifade TJ, Cinar HN, Tall BD, Taviani E, Hasan NA, Abdullah AH, Carter L, Sahu SN, Kothary MH, Chen A, Baker R, Hutchinson R, Blackmore C, Cebula TA, Huq A, Colwell RR - PLoS ONE (2014)

rstR sequences from V. cholerae CP1110.Sequences are aligned to their most similar homologs extracted from NCBI Genbank database. Nulceotide sequence identity is shown to the right of the last nucleotide aligned for each allele.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3974666&req=5

pone-0086264-g002: rstR sequences from V. cholerae CP1110.Sequences are aligned to their most similar homologs extracted from NCBI Genbank database. Nulceotide sequence identity is shown to the right of the last nucleotide aligned for each allele.
Mentions: The V. cholerae FL Group isolates were determined to contain the full CTX phage encoding the cholera toxin, but the structure of this region was unresolved due to the limitations of assembly since ORFs were found on multiple contigs. For similar reasons, CTX phage copy number could not be resolved. A BLASTN analysis with V. cholerae N16961 and O395 as reference demonstrated the presence of regions homologous to VC1456 to VC1463 (VC0395_0512 to VC0395_0505 and VC0395_A1060 and VC0395_A1067 of V. cholerae O395) of the CTX phage (ctxB, ctxA, zot, ace, orfU,cep, rstB, rstA, and rstRClassical). To infer the biotype of the cholera toxin, PCR targeting the ctxB gene was employed and resulted in an amplicon for primers of targeting ctxBClassical. These PCR results are consistent with profiles of other clinically isolated V. cholerae strains on a global scale that suggest this cholera toxin biotype is the predominant biotype currently causing the majority of disease [34], [35]. Based on the genome sequence data, the CTX phage of the V. cholerae FL Group genomes were lacking the rstR gene of V. cholerae N16961 El Tor (VC1464), but did encode the rstR gene homologous to the one encoded in V. cholerae O395 Classical. To further investigate and confirm these in silico results, PCR targeting the rstR region was done and resulted in amplicons for the Calcutta, Environmental, and Classical biotypes, but not the El Tor biotype, an as-to-date uncommon combination. The rstR amplicons of CP1110 were subjected to Sanger sequencing and the resulting sequences were compared by BLASTN to the NCBI Genbank database for better interpretation of these results and each showed ≥99% nucleotide sequence similarity to Calcutta, Environmental, and Classical sequences (Figure 2). These amplicon sequences were compared with V. cholerae CP1110 reads by BLASTN to re-confirm their presence in the genome sequences. The rstR sequences from the V. cholerae FL Group were confirmed as Calcutta, Environmental, and Classical biotypes (Figure 2). The prototypical V. cholerae O1 El Tor strains encode rstREl Tor and ctxBEl Tor while Classical strains encode rstRClassical and ctxBClassical. Altered V. cholerae O1 El Tor strains which differ from prototypical El Tor strains in their rstR/ctxB types have recently been identified [24]. Data from this study further demonstrates the diversity of the CTX phage outside of the more frequently studied V. cholerae O1 strains and suggests many alleles of this phage can be associated with cholera. Cholera toxin expression was not assayed in this study.

Bottom Line: From the 11 diagnosed cases, eight isolates of V. cholerae were isolated and their genomes were sequenced.Comparative genomics, phenotypic analyses, and a Caenorhabditis elegans model of infection for the isolates were conducted.This analysis coupled with isolation data of V. cholerae O75 and O141 suggests these strains may represent an underappreciated clade of cholera-causing strains responsible for significant disease burden globally.

View Article: PubMed Central - PubMed

Affiliation: Maryland Pathogen Research Institute, University of Maryland, College Park, Maryland, United States of America.

ABSTRACT
Between November 2010, and May 2011, eleven cases of cholera, unrelated to a concurrent outbreak on the island of Hispaniola, were recorded, and the causative agent, Vibrio cholerae serogroup O75, was traced to oysters harvested from Apalachicola Bay, Florida. From the 11 diagnosed cases, eight isolates of V. cholerae were isolated and their genomes were sequenced. Genomic analysis demonstrated the presence of a suite of mobile elements previously shown to be involved in the disease process of cholera (ctxAB, VPI-1 and -2, and a VSP-II like variant) and a phylogenomic analysis showed the isolates to be sister taxa to toxigenic V. cholerae V51 serogroup O141, a clinical strain isolated 23 years earlier. Toxigenic V. cholerae O75 has been repeatedly isolated from clinical cases in the southeastern United States and toxigenic V. cholerae O141 isolates have been isolated globally from clinical cases over several decades. Comparative genomics, phenotypic analyses, and a Caenorhabditis elegans model of infection for the isolates were conducted. This analysis coupled with isolation data of V. cholerae O75 and O141 suggests these strains may represent an underappreciated clade of cholera-causing strains responsible for significant disease burden globally.

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Related in: MedlinePlus