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Sequence of Shiga toxin 2 phage 933W from Escherichia coli O157:H7: Shiga toxin as a phage late-gene product.

ABSTRACT: Lysogenic bacteriophages are major vehicles for the transfer of genetic information between bacteria, including pathogenicity and/or virulence determinants. In the enteric pathogen Escherichia coli O157:H7, which causes hemorrhagic colitis and hemolytic-uremic syndrome, Shiga toxins 1 and 2 (Stx1 and Stx2) are phage encoded. The sequence and analysis of the Stx2 phage 933W is presented here. We find evidence that the toxin genes are part of a late-phage transcript, suggesting that toxin production may be coupled with, if not dependent upon, phage release during lytic growth. Another phage gene, stk, encodes a product resembling eukaryotic serine/threonine protein kinases. Based on its position in the sequence, Stk may be produced by the prophage in the lysogenic state, and, like the YpkA protein of Yersinia species, it may interfere with the signal transduction pathway of the mammalian host. Three novel tRNA genes present in the phage genome may serve to increase the availability of rare tRNA species associated with efficient expression of pathogenicity determinants: both the Shiga toxin and serine/threonine kinase genes contain rare isoleucine and arginine codons. 933W also has homology to lom, encoding a member of a family of outer membrane proteins associated with virulence by conferring the ability to survive in macrophages, and bor, implicated in serum resistance.

SUBMITTER: Plunkett G

PROVIDER: S-EPMC93574 | biostudies-literature | 1999 Mar

REPOSITORIES: biostudies-literature

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Sequence of Shiga toxin 2 phage 933W from Escherichia coli O157:H7: Shiga toxin as a phage late-gene product.

Plunkett G G Rose D J DJ Durfee T J TJ Blattner F R FR

Journal of bacteriology 19990301 6

Lysogenic bacteriophages are major vehicles for the transfer of genetic information between bacteria, including pathogenicity and/or virulence determinants. In the enteric pathogen Escherichia coli O157:H7, which causes hemorrhagic colitis and hemolytic-uremic syndrome, Shiga toxins 1 and 2 (Stx1 and Stx2) are phage encoded. The sequence and analysis of the Stx2 phage 933W is presented here. We find evidence that the toxin genes are part of a late-phage transcript, suggesting that toxin producti ...[more]

PMID: 10074068

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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.

Project description:BACKGROUND:Shiga toxin-producing Escherichia coli O157:H7 is a foodborne pathogen that causes severe human diseases including hemolytic uremic syndrome (HUS). The virulence factor that mediates HUS, Shiga toxin (Stx), is encoded within the genome of a lambdoid prophage. Although draft sequences are publicly available for a large number of E. coli O157:H7 strains, the high sequence similarity of stx-converting bacteriophages with other lambdoid prophages poses challenges to accurately assess the organization and plasticity among stx-converting phages due to assembly difficulties. METHODS:To further explore genome plasticity of stx-converting prophages, we enriched phage DNA from 45 ciprofloxacin-induced cultures for subsequent 454 pyrosequencing to facilitate assembly of the complete phage genomes. In total, 22 stx2a-converting phage genomes were closed. RESULTS:Comparison of the genomes distinguished nine distinct phage sequence types (PSTs) delineated by variation in obtained sequences, such as single nucleotide polymorphisms (SNPs) and insertion sequence element prevalence and location. These nine PSTs formed three distinct clusters, designated as PST1, PST2 and PST3. The PST2 cluster, identified in two clade 8 strains, was related to stx2a-converting phages previously identified in non-O157 Shiga-toxin producing E. coli (STEC) strains associated with a high incidence of HUS. The PST1 cluster contained phages related to those from E. coli O157:H7 strain Sakai (lineage I, clade 1), and PST3 contained a single phage that was distinct from the rest but most related to the phage from E. coli O157:H7 strain EC4115 (lineage I/II, clade 8). Five strains carried identical stx2a-converting phages (PST1-1) integrated at the same chromosomal locus, but these strains produced different levels of Stx2. CONCLUSION:The stx2a-converting phages of E. coli O157:H7 can be categorized into at least three phage types. Diversification within a phage type is mainly driven by IS629 and by a small number of SNPs. Polymorphisms between phage genomes may help explain differences in Stx2a production between strains, however our data indicates that genes encoded external to the phage affect toxin production as well.

Dataset Information

Sequence of Shiga toxin 2 phage 933W from Escherichia coli O157:H7: Shiga toxin as a phage late-gene product.

Publications

Sequence of Shiga toxin 2 phage 933W from Escherichia coli O157:H7: Shiga toxin as a phage late-gene product.

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