title: Rapid diagnosis of toxinogenic Clostridium difficile in faecal samples with internally controlled real-time PCR.
abstract: A real-time PCR assay for Clostridium difficile was developed, based on the tcdB gene, which detected all known toxinogenic reference strains (n = 45), within 30 serogroups and 24 toxinotypes. The analytical sensitivity was 1 x 10(3) CFU/mL, and the detection limit in faeces was 1 x 10(5) CFU/g. The optimal protocol for DNA extraction from faecal samples involved use of the MagnaPure system with a Stool Transport and Recovery (STAR) buffer pre-treatment. In a 1-month prospective study of 85 patients with diarrhoea, the sensitivity, specificity and positive and negative predictive values of the assay were 100%, 94%, 55% and 100%, respectively, compared with the standard cell cytotoxicity assay.
authors.affiliation: Department of Medical Microbiology, Center of Infectious Diseases, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: R J
authors.initials: RJ
authors.lastname: van den Berg
authors.forename: E J
authors.initials: EJ
authors.lastname: Kuijper
authors.forename: L E S Bruijnesteijn
authors.initials: LE
authors.lastname: van Coppenraet
authors.forename: E C J
authors.initials: EC
authors.lastname: Claas
mesh_terms: D000328:Adult; D001426:Bacterial Proteins / Q000235:genetics*; D001427:Bacterial Toxins / Q000235:genetics*; D016360:Clostridioides difficile / Q000235:genetics / Q000302:isolation & purification*; D004269:DNA, Bacterial / Q000032:analysis*; D003967:Diarrhea / Q000382:microbiology; D004761:Enterocolitis, Pseudomembranous / Q000175:diagnosis* / Q000382:microbiology; D005243:Feces / Q000382:microbiology*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011237:Predictive Value of Tests; D011786:Quality Control; D012015:Reference Standards; D012680:Sensitivity and Specificity
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004269:DNA, Bacterial; C031815:toxB protein, Clostridium difficile