Dataset Information

Multiplex real-time PCR method to identify Shiga toxin genes stx1 and stx2 and Escherichia coli O157:H7/H- serotype.

ABSTRACT: A multiplex real-time PCR method to simultaneously detect the stx1 and stx2 genes of Shiga toxin-producing Escherichia coli and a unique conserved single-nucleotide polymorphism in the E. coli O157:H7/H- uidA gene has been developed. There is more than 98.6% sensitivity and 100% specificity for all three gene targets based on a panel of 138 isolates. The PCR efficiencies were >/= 1.89, and as few as 6 CFU/reaction could be detected.

SUBMITTER: Jinneman KC

PROVIDER: S-EPMC201207 | biostudies-literature | 2003 Oct

REPOSITORIES: biostudies-literature

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Multiplex real-time PCR method to identify Shiga toxin genes stx1 and stx2 and Escherichia coli O157:H7/H- serotype.

Jinneman Karen C KC Yoshitomi Ken J KJ Weagant Stephen D SD

Applied and environmental microbiology 20031001 10

A multiplex real-time PCR method to simultaneously detect the stx1 and stx2 genes of Shiga toxin-producing Escherichia coli and a unique conserved single-nucleotide polymorphism in the E. coli O157:H7/H- uidA gene has been developed. There is more than 98.6% sensitivity and 100% specificity for all three gene targets based on a panel of 138 isolates. The PCR efficiencies were >/= 1.89, and as few as 6 CFU/reaction could be detected. ...[more]

PMID: 14532101

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Project description:Escherichia coli O157:H7 is a foodborne pathogen implicated in various multistate outbreaks. It encodes Shiga toxin on a prophage, and Shiga toxin production is linked to phage induction. An E. coli strain, designated 0.1229, that amplified Stx2a production when cocultured with E. coli O157:H7 strain PA2 was identified. Growth of PA2 in 0.1229 cell-free supernatants had a similar effect, even when supernatants were heated to 100°C for 10?min, but not after treatment with proteinase K. The secreted molecule was shown to use TolC for export and the TonB system for import. The genes sufficient for production of this molecule were localized to a 5.2-kb region of a 12.8-kb plasmid. This region was annotated, identifying hypothetical proteins, a predicted ABC transporter, and a cupin superfamily protein. These genes were identified and shown to be functional in two other E. coli strains, and bioinformatic analyses identified related gene clusters in similar and distinct bacterial species. These data collectively suggest that E. coli 0.1229 and other E. coli strains produce a microcin that induces the SOS response in target bacteria. Besides adding to the limited number of microcins known to be produced by E. coli, this study provides an additional mechanism by which stx 2a expression is increased in response to the gut microflora.IMPORTANCE How the gut microflora influences the progression of bacterial infections is only beginning to be understood. Antibiotics are counterindicated for E. coli O157:H7 infections, limiting treatment options. An increased understanding of how the gut microflora directs O157:H7 virulence gene expression may lead to additional treatment options. This work identified E. coli strains that enhance the production of Shiga toxin by O157:H7 through the secretion of a proposed microcin. Microcins are natural antimicrobial peptides that target specific species, can act as alternatives to antibiotics, and mediate microbial competition. This work demonstrates another mechanism by which non-O157 E. coli strains may increase Shiga toxin production and adds to our understanding of microcins, a group of antimicrobials less well understood than colicins.

Dataset Information

Multiplex real-time PCR method to identify Shiga toxin genes stx1 and stx2 and Escherichia coli O157:H7/H- serotype.

Publications

Multiplex real-time PCR method to identify Shiga toxin genes stx1 and stx2 and Escherichia coli O157:H7/H- serotype.

Similar Datasets

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