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Six Novel O Genotypes from Shiga Toxin-Producing Escherichia coli.

ABSTRACT: Serotyping is one of the typing techniques used to classify strains within the same species. O-serogroup diversification shows a strong association with the genetic diversity of O-antigen biosynthesis genes. In a previous study, based on the O-antigen biosynthesis gene cluster (O-AGC) sequences of 184 known Escherichia coli O serogroups (from O1 to O187), we developed a comprehensive and practical molecular O serogrouping (O genotyping) platform using a polymerase chain reaction (PCR) method, named E. coli O-genotyping PCR. Although, the validation assay using the PCR system showed that most of the tested strains were successfully classified into one of the O genotypes, it was impossible to classify 6.1% (35/575) of the strains, suggesting the presence of novel O genotypes. In this study, we conducted sequence analysis of O-AGCs from O-genotype untypeable Shiga toxin-producing E. coli (STEC) strains and identified six novel O genotypes; OgN1, OgN8, OgN9, OgN10, OgN12 and OgN31, with unique wzx and/or wzy O-antigen processing gene sequences. Additionally, to identify these novel O-genotypes, we designed specific PCR primers. A screen of O genotypes using O-genotype untypeable strains showed 13 STEC strains were classified into five novel O genotypes. The O genotyping at the molecular level of the O-AGC would aid in the characterization of E. coli isolates and will assist future studies in STEC epidemiology and phylogeny.

SUBMITTER: Iguchi A

PROVIDER: S-EPMC4873512 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Six Novel O Genotypes from Shiga Toxin-Producing Escherichia coli.

Iguchi Atsushi A Iyoda Sunao S Seto Kazuko K Nishii Hironobu H Ohnishi Makoto M Mekata Hirohisa H Ogura Yoshitoshi Y Hayashi Tetsuya T

Frontiers in microbiology 20160520

Serotyping is one of the typing techniques used to classify strains within the same species. O-serogroup diversification shows a strong association with the genetic diversity of O-antigen biosynthesis genes. In a previous study, based on the O-antigen biosynthesis gene cluster (O-AGC) sequences of 184 known Escherichia coli O serogroups (from O1 to O187), we developed a comprehensive and practical molecular O serogrouping (O genotyping) platform using a polymerase chain reaction (PCR) method, na ...[more]

PMID: 27242776

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Project description:Many pathogens produce the ?-(1-6)-linked poly-N-acetylglucosamine (PNAG) surface polysaccharide that is being developed as a broadly protective antimicrobial vaccine. However, it is unknown whether systemically injected PNAG vaccines or antibodies would provide protective immunity against pathogens confined to the gastrointestinal tract such as Shiga toxin (Stx)-producing Escherichia coli (STEC), an important group of gastrointestinal (GI) pathogens for which effective immunotherapeutics are lacking. To ascertain whether systemic IgG antibody to PNAG impacts this infectious situation, a vaccine consisting of a synthetic nonamer of nonacetylated PNAG, 9GlcNH2, conjugated to the Shiga toxin 1b subunit (9GlcNH2-Stx1b) was produced. Rabbit antibodies raised to the conjugate vaccine were tested for bacterial killing and toxin neutralization in vitro and protection against infection in infant mice. Cell surface PNAG was detected on all 9 STEC isolates tested, representing 6 STEC serogroups, including E. coli O157:H7. Antibody to the 9GlcNH2-Stx1b conjugate neutralized Stx1 potently and Stx2 modestly. For O157:H7 and O104:H4 STEC strains, antibodies elicited by the 9GlcNH2-Stx1b conjugate possessed opsonic killing and bactericidal activity. Following intraperitoneal injection, antibodies to both PNAG and Stx were needed for infant mouse protection against O157 STEC. These antibodies also mediated protection against the Stx2-producing O104:H4 strain that was the cause of a recent outbreak in Germany, although sufficient doses of antibody to PNAG alone were protective against this strain in infant mice. Our observations suggest that vaccination against both PNAG and Stx, using a construct such as the 9GlcNH2-Stx1b conjugate vaccine, would be protective against a broad range of STEC serogroups. IMPORTANCE The presence of poly-N-acetylglucosamine (PNAG) on many pathogens presents an opportunity to target this one structure with a multispecies vaccine. Whether antibodies to PNAG can protect against pathogens confined to the gastrointestinal tract is not known. As Shiga toxin (Stx)-producing Escherichia coli (STEC) bacteria are serious causes of infection whose virulence is dependent on elaboration of Stx, we prepared a vaccine containing a synthetic nonamer of PNAG (9GlcNH2) conjugated to Shiga toxin 1b subunit (9GlcNH2-Stx1b) to evaluate bacterial killing, toxin neutralization, and protective efficacy in infant mice. All nine (100%) clinical strains of STEC from different serogroups expressed PNAG. Vaccine-induced antibody mediated in vitro killing of STEC and neutralization of both Stx1 and Stx2. Passive administration of antibody to the conjugate showed protection requiring immunity to both PNAG and Stx for O157 strains, although for an O104 strain, antibody to PNAG alone was protective. Immunity to PNAG may contribute to protection against STEC infections.

Dataset Information

Six Novel O Genotypes from Shiga Toxin-Producing Escherichia coli.

Publications

Six Novel O Genotypes from Shiga Toxin-Producing Escherichia coli.

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