Multiplex PCR Targeting<i>tpi</i>(Triose Phosphate Isomerase),<i>tcdA</i>(Toxin A), and<i>tcdB</i>(Toxin B) Genes for Toxigenic Culture of<i>Clostridium difficile</i>

DOI Web Site 被引用文献2件
  • Ludovic Lemee
    Groupe de Recherche sur les Antimicrobiens et les Microorganismes (G.R.A.M. EA 2656, I.F.R. 23), Université de Rouen, U.F.R. Médecine-Pharmacie, and Service de Bactériologie, Centre Hospitalier Universitaire, Rouen
  • Anne Dhalluin
    Groupe de Recherche sur les Antimicrobiens et les Microorganismes (G.R.A.M. EA 2656, I.F.R. 23), Université de Rouen, U.F.R. Médecine-Pharmacie, and Service de Bactériologie, Centre Hospitalier Universitaire, Rouen
  • Sabrina Testelin
    Groupe de Recherche sur les Antimicrobiens et les Microorganismes (G.R.A.M. EA 2656, I.F.R. 23), Université de Rouen, U.F.R. Médecine-Pharmacie, and Service de Bactériologie, Centre Hospitalier Universitaire, Rouen
  • Marie-Andre Mattrat
    Groupe de Recherche sur les Antimicrobiens et les Microorganismes (G.R.A.M. EA 2656, I.F.R. 23), Université de Rouen, U.F.R. Médecine-Pharmacie, and Service de Bactériologie, Centre Hospitalier Universitaire, Rouen
  • Karine Maillard
    Laboratoire Départemental Frank Duncombe, Département Santé Animale, Caen, France
  • Jean-François Lemeland
    Groupe de Recherche sur les Antimicrobiens et les Microorganismes (G.R.A.M. EA 2656, I.F.R. 23), Université de Rouen, U.F.R. Médecine-Pharmacie, and Service de Bactériologie, Centre Hospitalier Universitaire, Rouen
  • Jean-Louis Pons
    Groupe de Recherche sur les Antimicrobiens et les Microorganismes (G.R.A.M. EA 2656, I.F.R. 23), Université de Rouen, U.F.R. Médecine-Pharmacie, and Service de Bactériologie, Centre Hospitalier Universitaire, Rouen

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<jats:title>ABSTRACT</jats:title><jats:p>A multiplex PCR toxigenic culture approach was designed for simultaneous identification and toxigenic type characterization of<jats:italic>Clostridium difficile</jats:italic>isolates. Three pairs of primers were designed for the amplification of (i) a species-specific internal fragment of the<jats:italic>tpi</jats:italic>(triose phosphate isomerase) gene, (ii) an internal fragment of the<jats:italic>tcdB</jats:italic>(toxin B) gene, and (iii) an internal fragment of the<jats:italic>tcdA</jats:italic>(toxin A) gene allowing distinction between toxin A-positive, toxin B-positive (A+B+) strains and toxin A-negative, toxin B-positive (A−B+) variant strains. The reliability of the multiplex PCR was established by using a panel of 72<jats:italic>C. difficile</jats:italic>strains including A+B+, A−B−, and A−B+ toxigenic types and 11 other<jats:italic>Clostridium</jats:italic>species type strains. The multiplex PCR assay was then included in a toxigenic culture approach for the detection, identification, and toxigenic type characterization of<jats:italic>C. difficile</jats:italic>in 1,343 consecutive human and animal stool samples. Overall, 111 (15.4%) of 721 human samples were positive for<jats:italic>C. difficile</jats:italic>; 67 (60.4%) of these samples contained A+B+ toxigenic isolates, and none of them contained A−B+ variant strains. Fifty (8%) of 622 animal samples contained<jats:italic>C. difficile</jats:italic>strains, which were toxigenic in 27 (54%) cases, including 1 A−B+ variant isolate. Eighty of the 721 human stool samples (37 positive and 43 negative for<jats:italic>C. difficile</jats:italic>culture) were comparatively tested by Premier Toxins A&B (Meridian Bioscience) and Triage<jats:italic>C. difficile</jats:italic>Panel (Biosite) immunoassays, the results of which were found concordant with toxigenic culture for 82.5 and 92.5% of the samples, respectively. The multiplex PCR toxigenic culture scheme described here allows combined diagnosis and toxigenic type characterization for human and animal<jats:italic>C. difficile</jats:italic>intestinal infections.</jats:p>

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