title: Specific detection of Clostridium difficile toxin A gene sequences in clinical isolates.
abstract: The polymerase chain reaction (PCR) was used to specifically detect toxin A gene sequences of Clostridium difficile in DNA isolated from human faeces. A set of oligonucleotide primers derived from the non-repetitive region of the toxin A gene was developed to amplify a 634-bp DNA fragment. All 28 cytotoxic strains of C. difficile, previously characterized by a toxin B-PCR assay, were positive for the presence of toxin A gene sequences. No amplification products were obtained from DNAs extracted from non-toxigenic strains, strains of C. sordellii, or C. bifermentans. In addition, amplification of DNA extracted from C. difficile 8864, a strain which does not produce toxin A, resulted in multiple bands which probed negative for toxin A gene sequences. DNAs extracted from nine stool specimens which were positive for toxin B by the cytotoxicity assay and by the toxin B-PCR assay were also positive in this assay. Toxin A gene sequences were detected in DNAs obtained from 4/11 stool specimens which were negative by the toxin B cytotoxicity assay. These four specimens were from patients who had a history of relapses due to C. difficile-associated colitis, and whose stools had previously been found to be positive by the toxin B-PCR test despite no detectable toxin B in the specimens. These data indicate a comparable degree of clinical sensitivity between these two toxin-gene PCR-based assays. This rapid, sensitive and specific assay may be useful not only in the diagnosis of C. difficile infections, but also in molecular studies of the toxin A gene in C. difficile strains.
authors.affiliation: Department of Internal Medicine, University of California, Davis School of Medicine.
authors.forename: Y J
authors.initials: YJ
authors.lastname: Tang
authors.forename: P H
authors.initials: PH
authors.lastname: Gumerlock
authors.forename: J B
authors.initials: JB
authors.lastname: Weiss
authors.forename: J
authors.initials: J
authors.lastname: Silva
mesh_terms: D001426:Bacterial Proteins; D001427:Bacterial Toxins / Q000235:genetics; D001483:Base Sequence; D016360:Clostridioides difficile / Q000737:chemistry* / Q000235:genetics; D017931:DNA Primers; D004269:DNA, Bacterial / Q000235:genetics* / Q000302:isolation & purification; D004768:Enterotoxins / Q000235:genetics*; D005243:Feces / Q000382:microbiology; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D017931:DNA Primers; D004269:DNA, Bacterial; D004768:Enterotoxins; C041348:tcdA protein, Clostridium difficile; C031815:toxB protein, Clostridium difficile