Diagnostic Real-Time PCR Assays for the Detection of Emetic Bacillus cereus Strains in Foods and Recent Food-Borne Outbreaks

• Published in: Applied and Environmental Microbiology Vol. 73; no. 6; pp. 1892 - 1898
• Main Authors: Fricker, Martina, Messelhäusser, Ute, Busch, Ulrich, Scherer, Siegfried, Ehling-Schulz, Monika
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.03.2007
• Subjects: Bacillus cereus; Bacillus cereus - genetics; Bacillus cereus - isolation & purification; Bacteria; Bacteriology; Benzothiazoles; Biological and medical sciences; Brassica; Deoxyribonucleic acid; Diagnosis, Differential; Diamines; Disease Outbreaks; DNA; DNA Primers - genetics; Food contamination & poisoning; Food Microbiology; Foodborne Diseases - diagnosis; Foodborne Diseases - epidemiology; Foodborne Diseases - microbiology; Fundamental and applied biological sciences. Psychology; Genetics; Humans; Microbiology; Organic Chemicals - metabolism; Oryza sativa; Pathology; Polymerase Chain Reaction - methods; Polymerase Chain Reaction - standards; Quinolines; Real time; Reference Standards; Sensitivity and Specificity; Staining and Labeling - methods; Staphylococcus aureus; Germany; Epidemic; Polymerase chain reaction; Bacillus cereus; Bacillales; Bacteria; Foodstuff; Diagnosis; Bacillaceae; Detection; Real time
• ISSN: 0099-2240; 1098-5336
• DOI: 10.1128/AEM.02219-06

Cereulide-producing Bacillus cereus can cause an emetic type of food-borne disease that mimics the symptoms provoked by Staphylococcus aureus. Based on the recently discovered genetic background for cereulide formation, a novel 5' nuclease (TaqMan) real-time PCR assay was developed to provide a rapid and sensitive method for the specific detection of emetic B. cereus in food. The TaqMan assay includes an internal amplification control and primers and a probe designed to target a highly specific part of the cereulide synthetase genes. Additionally, a specific SYBR green I assay was developed and extended to create a duplex SYBR green I assay for the one-step identification and discrimination of the two emesis-causing food pathogens B. cereus and S. aureus. The inclusivity and exclusivity of the assay were assessed using a panel of 100 strains, including 23 emetic B. cereus and 14 S. aureus strains. Different methods for DNA isolation from artificially contaminated foods were evaluated, and established real-time assays were used to analyze two recent emetic food poisonings in southern Germany. One of the food-borne outbreaks included 17 children visiting a day care center who vomited after consuming a reheated rice dish, collapsed, and were hospitalized; the other case concerned a single food-poisoning incident occurring after consumption of cauliflower. Within 2 h, the etiological agent of these food poisonings was identified as emetic B. cereus by using the real-time PCR assay.