title: Improvement in the detection rate of diarrhoeagenic bacteria in human stool specimens by a rapid real-time PCR assay.
abstract: A rapid laboratory system has been developed and evaluated that can simultaneously identify major diarrhoeagenic bacteria, including Salmonella enterica, Vibrio parahaemolyticus, Campylobacter jejuni and Shiga toxin-producing Escherichia coli, in stool specimens by real-time PCR. Specific identification was achieved by using selective TaqMan probes, detecting two targets in each pathogen. A positive result was scored only when both targets of a pathogen were amplified and the difference between threshold cycles for detection was less than five. Diagnosis of enteric bacterial infections using this highly sensitive method, including DNA extraction and real-time PCR, requires only 3 h. Forty stool specimens related to suspected food poisoning outbreaks were analysed: 16 (40%) of these samples were found to be positive for diarrhoeagenic bacteria using a conventional culture method; 28 (70%) were positive using the real-time PCR assay. Of the 12 PCR-positive but culture-negative cases, 11 patients had consumed pathogen-contaminated or high-risk food. Analysis of faecal samples from 105 outpatients who complained of diarrhoea and/or abdominal pain identified 19 (18%) patients as being positive for diarrhoeagenic bacteria using the culture method. An additional six (6%) patients were found to be positive by PCR analysis.
authors.affiliation: Department of Microbiology, Kobe Institute of Health, Minatojima-nakamachi, Chuo-ku, Kobe 650-0046, Japan 2Department of Microbiology, Tenri Hospital, Mishima-cho, Tenri 632-8552, Japan.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Iijima
authors.affiliation: Department of Microbiology, Kobe Institute of Health, Minatojima-nakamachi, Chuo-ku, Kobe 650-0046, Japan 2Department of Microbiology, Tenri Hospital, Mishima-cho, Tenri 632-8552, Japan.
authors.forename: Nahoko T
authors.initials: NT
authors.lastname: Asako
authors.affiliation: Department of Microbiology, Kobe Institute of Health, Minatojima-nakamachi, Chuo-ku, Kobe 650-0046, Japan 2Department of Microbiology, Tenri Hospital, Mishima-cho, Tenri 632-8552, Japan.
authors.forename: Masanori
authors.initials: M
authors.lastname: Aihara
authors.affiliation: Department of Microbiology, Kobe Institute of Health, Minatojima-nakamachi, Chuo-ku, Kobe 650-0046, Japan 2Department of Microbiology, Tenri Hospital, Mishima-cho, Tenri 632-8552, Japan.
authors.forename: Kozaburo
authors.initials: K
authors.lastname: Hayashi
mesh_terms: D015746:Abdominal Pain / Q000382:microbiology; D001419:Bacteria / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D001424:Bacterial Infections / Q000382:microbiology*; D016123:Campylobacter jejuni / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000032:analysis / Q000302:isolation & purification; D003967:Diarrhea / Q000382:microbiology*; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification; D005243:Feces / Q000382:microbiology*; D005517:Foodborne Diseases / Q000382:microbiology; D005759:Gastroenteritis / Q000382:microbiology; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D019779:Salmonella enterica / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D014736:Vibrio parahaemolyticus / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification
chemical_list: D004269:DNA, Bacterial