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.
Highlights
Serotyping is a standard method for subtyping of Escherichia coli strains in taxonomical and epidemiological studies (Orskov and Orskov, 1984)
Six novel O genotypes were revealed from Shiga toxin-producing E. coli (STEC) strains isolated from human patients, and the prevalence of these O genotype strains was confirmed in STECs
The O1-serogroup strain is often seen in extra-intestinal pathogenic E. coli from patients with urinary tract infections (Abe et al, 2008; Mora et al, 2009) and septicemic disease (Mora et al, FIGURE 3 | Comparison of O-antigen biosynthesis gene cluster (O-AGC). (A) The pair of O-AGCs, OgN31 and O116. (B) Four pairs that were serologically agglutinated with the same O antisera carried different types of O-AGCs
Summary
Serotyping is a standard method for subtyping of Escherichia coli strains in taxonomical and epidemiological studies (Orskov and Orskov, 1984). The World Health Organization Collaborating Centre for Reference and Research on Escherichia and Klebsiella, which is based at the Statens Serum Institut (SSI) in Denmark, has recognized 185 E. coli O serogroups. These are designated O1 to O188 (publication of O182 to O188 is pending) and include three pairs of subgroups, O18ab/ac, O28ab/ac, and O112ab/ac; and six missing numbers, O31, O47, O67, O72, O93, and O122 (Orskov and Orskov, 1992; Scheutz et al, 2004). Sequences from O-antigen processing genes (wzx/wzy and wzm/wzt) located on the O-AGCs are highly variable and can be used as gene markers for the identification of O serogroups via molecular approaches. We presented a comprehensive molecular O-typing scheme: an E. coli O-genotyping polymerase chain reaction (ECOG-PCR) system using 20 multiplex PCR sets containing 162 O-genotype-specific PCR primers (Iguchi et al, 2015b)
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