Escherichia coli O-Genotyping PCR: a Comprehensive and Practical Platform for Molecular O Serogrouping

The O serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In particular, the identification of strains of the same O serogroup is essential in outbreak investigations and surveillance. In a previous st...

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Published inJournal of clinical microbiology Vol. 53; no. 8; pp. 2427 - 2432
Main Authors Iguchi, Atsushi, Iyoda, Sunao, Seto, Kazuko, Morita-Ishihara, Tomoko, Scheutz, Flemming, Ohnishi, Makoto
Format Journal Article
LanguageEnglish
Published United States American Society for Microbiology 01.08.2015
Subjects
Animals
Bacteriology
DNA Primers - genetics
Escherichia coli
Escherichia coli - classification
Escherichia coli - genetics
Escherichia coli - isolation & purification
Escherichia coli Infections - microbiology
Escherichia coli Infections - veterinary
Genotyping Techniques - methods
Humans
Multiplex Polymerase Chain Reaction - methods
O Antigens - genetics
Sensitivity and Specificity
Serogroup
Serotyping - methods
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Abstract The O serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In particular, the identification of strains of the same O serogroup is essential in outbreak investigations and surveillance. In a previous study, we analyzed the O-antigen biosynthesis gene cluster in all known E. coli O serogroups (A. Iguchi et al., DNA Res, 22:101–107, 2015, http://dx.doi.org/10.1093/dnares/dsu043 ). Based on those results, we have arranged 162 PCR primer pairs for the identification or classification of O serogroups. Of these, 147 pairs were used to identify 147 individual O serogroups with unique O-antigen biosynthesis genes, and the other 15 pairs were used to identify 15 groups of strains (Gp1 to Gp15). Each of these groups consisted of strains with identical or very similar O-antigen biosynthesis genes, and the groups represented a total of 35 individual O serogroups. We then used the 162 primer pairs to create 20 multiplex PCR sets. Each set contained six to nine primer pairs that amplify products of markedly different sizes. This genetic methodology ( E. coli O-genotyping PCR) allowed for comprehensive, rapid, and low-cost typing. Validation of the PCR system using O-serogroup references and wild strains showed that the correct O serogroups were specifically and accurately identified for 100% (182/182) and 90.8% (522/575) of references and wild strains, respectively. The PCR-based system reported here might be a promising tool for the subtyping of E. coli strains for epidemiological studies as well as for the surveillance of pathogenic E. coli during outbreaks.
AbstractList The O serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In particular, the identification of strains of the same O serogroup is essential in outbreak investigations and surveillance. In a previous study, we analyzed the O-antigen biosynthesis gene cluster in all known E. coli O serogroups (A. Iguchi et al., DNA Res, 22:101-107, 2015, http://dx.doi.org/10.1093/dnares/dsu043). Based on those results, we have arranged 162 PCR primer pairs for the identification or classification of O serogroups. Of these, 147 pairs were used to identify 147 individual O serogroups with unique O-antigen biosynthesis genes, and the other 15 pairs were used to identify 15 groups of strains (Gp1 to Gp15). Each of these groups consisted of strains with identical or very similar O-antigen biosynthesis genes, and the groups represented a total of 35 individual O serogroups. We then used the 162 primer pairs to create 20 multiplex PCR sets. Each set contained six to nine primer pairs that amplify products of markedly different sizes. This genetic methodology (E. coli O-genotyping PCR) allowed for comprehensive, rapid, and low-cost typing. Validation of the PCR system using O-serogroup references and wild strains showed that the correct O serogroups were specifically and accurately identified for 100% (182/182) and 90.8% (522/575) of references and wild strains, respectively. The PCR-based system reported here might be a promising tool for the subtyping of E. coli strains for epidemiological studies as well as for the surveillance of pathogenic E. coli during outbreaks.The O serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In particular, the identification of strains of the same O serogroup is essential in outbreak investigations and surveillance. In a previous study, we analyzed the O-antigen biosynthesis gene cluster in all known E. coli O serogroups (A. Iguchi et al., DNA Res, 22:101-107, 2015, http://dx.doi.org/10.1093/dnares/dsu043). Based on those results, we have arranged 162 PCR primer pairs for the identification or classification of O serogroups. Of these, 147 pairs were used to identify 147 individual O serogroups with unique O-antigen biosynthesis genes, and the other 15 pairs were used to identify 15 groups of strains (Gp1 to Gp15). Each of these groups consisted of strains with identical or very similar O-antigen biosynthesis genes, and the groups represented a total of 35 individual O serogroups. We then used the 162 primer pairs to create 20 multiplex PCR sets. Each set contained six to nine primer pairs that amplify products of markedly different sizes. This genetic methodology (E. coli O-genotyping PCR) allowed for comprehensive, rapid, and low-cost typing. Validation of the PCR system using O-serogroup references and wild strains showed that the correct O serogroups were specifically and accurately identified for 100% (182/182) and 90.8% (522/575) of references and wild strains, respectively. The PCR-based system reported here might be a promising tool for the subtyping of E. coli strains for epidemiological studies as well as for the surveillance of pathogenic E. coli during outbreaks.
The O serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In particular, the identification of strains of the same O serogroup is essential in outbreak investigations and surveillance. In a previous study, we analyzed the O-antigen biosynthesis gene cluster in all known E. coli O serogroups (A. Iguchi et al., DNA Res, 22:101–107, 2015, http://dx.doi.org/10.1093/dnares/dsu043 ). Based on those results, we have arranged 162 PCR primer pairs for the identification or classification of O serogroups. Of these, 147 pairs were used to identify 147 individual O serogroups with unique O-antigen biosynthesis genes, and the other 15 pairs were used to identify 15 groups of strains (Gp1 to Gp15). Each of these groups consisted of strains with identical or very similar O-antigen biosynthesis genes, and the groups represented a total of 35 individual O serogroups. We then used the 162 primer pairs to create 20 multiplex PCR sets. Each set contained six to nine primer pairs that amplify products of markedly different sizes. This genetic methodology ( E. coli O-genotyping PCR) allowed for comprehensive, rapid, and low-cost typing. Validation of the PCR system using O-serogroup references and wild strains showed that the correct O serogroups were specifically and accurately identified for 100% (182/182) and 90.8% (522/575) of references and wild strains, respectively. The PCR-based system reported here might be a promising tool for the subtyping of E. coli strains for epidemiological studies as well as for the surveillance of pathogenic E. coli during outbreaks.
The O serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In particular, the identification of strains of the same O serogroup is essential in outbreak investigations and surveillance. In a previous study, we analyzed the O-antigen biosynthesis gene cluster in all known E. coli O serogroups (A. Iguchi et al., DNA Res, 22:101-107, 2015, http://dx.doi.org/10.1093/dnares/dsu043). Based on those results, we have arranged 162 PCR primer pairs for the identification or classification of O serogroups. Of these, 147 pairs were used to identify 147 individual O serogroups with unique O-antigen biosynthesis genes, and the other 15 pairs were used to identify 15 groups of strains (Gp1 to Gp15). Each of these groups consisted of strains with identical or very similar O-antigen biosynthesis genes, and the groups represented a total of 35 individual O serogroups. We then used the 162 primer pairs to create 20 multiplex PCR sets. Each set contained six to nine primer pairs that amplify products of markedly different sizes. This genetic methodology (E. coli O-genotyping PCR) allowed for comprehensive, rapid, and low-cost typing. Validation of the PCR system using O-serogroup references and wild strains showed that the correct O serogroups were specifically and accurately identified for 100% (182/182) and 90.8% (522/575) of references and wild strains, respectively. The PCR-based system reported here might be a promising tool for the subtyping of E. coli strains for epidemiological studies as well as for the surveillance of pathogenic E. coli during outbreaks.
Author Iyoda, Sunao
Ohnishi, Makoto
Iguchi, Atsushi
Seto, Kazuko
Morita-Ishihara, Tomoko
Scheutz, Flemming
Author_xml – sequence: 1
  givenname: Atsushi
  surname: Iguchi
  fullname: Iguchi, Atsushi
  organization: Department of Animal and Grassland Sciences, Faculty of Agriculture, University of Miyazaki, Miyazaki, Japan
– sequence: 2
  givenname: Sunao
  surname: Iyoda
  fullname: Iyoda, Sunao
  organization: Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan
– sequence: 3
  givenname: Kazuko
  surname: Seto
  fullname: Seto, Kazuko
  organization: Division of Bacteriology, Osaka Prefectural Institute of Public Health, Osaka, Japan
– sequence: 4
  givenname: Tomoko
  surname: Morita-Ishihara
  fullname: Morita-Ishihara, Tomoko
  organization: Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan
– sequence: 5
  givenname: Flemming
  surname: Scheutz
  fullname: Scheutz, Flemming
  organization: Department of Microbiology Infection Control, Statens Serum Institut, Copenhagen, Denmark, WHO Collaborating Centre for Reference and Research on Escherichia and Klebsiella, Statens Serum Institut, Copenhagen, Denmark
– sequence: 6
  givenname: Makoto
  surname: Ohnishi
  fullname: Ohnishi, Makoto
  organization: Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan
BackLink https://www.ncbi.nlm.nih.gov/pubmed/25926488$$D View this record in MEDLINE/PubMed
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ContentType Journal Article
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Nakajima, Hiroshi
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Ishikawa, Kazuhiko
Saeki, Yuji
Okayama, Akihiko
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Isobe, Junko
Takeda, Yoshihiro
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  givenname: Masumi
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– sequence: 72
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Copyright Copyright © 2015, American Society for Microbiology. All Rights Reserved.
Copyright © 2015, American Society for Microbiology. All Rights Reserved. 2015 American Society for Microbiology
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Citation Iguchi A, Iyoda S, Seto K, Morita-Ishihara T, Scheutz F, Ohnishi M, Pathogenic E. coli Working Group in Japan. 2015. Escherichia coli O-genotyping PCR: a comprehensive and practical platform for molecular O serogrouping. J Clin Microbiol 53:2427–2432. doi:10.1128/JCM.00321-15.
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Snippet The O serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In...
The O serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In...
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SubjectTerms Animals
Bacteriology
DNA Primers - genetics
Escherichia coli
Escherichia coli - classification
Escherichia coli - genetics
Escherichia coli - isolation & purification
Escherichia coli Infections - microbiology
Escherichia coli Infections - veterinary
Genotyping Techniques - methods
Humans
Multiplex Polymerase Chain Reaction - methods
O Antigens - genetics
Sensitivity and Specificity
Serogroup
Serotyping - methods
Title Escherichia coli O-Genotyping PCR: a Comprehensive and Practical Platform for Molecular O Serogrouping
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