Diagnostic real-time PCR assays for the detection of emetic Bacillus cereus strains in foods and recent food-borne outbreaks.
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.
Fricker
Martina
M
Abt. Mikrobiologie/ZIEL, TUM, Weihenstephaner Berg 3, 85354 Freising, Germany. monika.ehling-schulz@wzw.tum.de
Messelhäusser
Ute
U
Busch
Ulrich
U
Scherer
Siegfried
S
Ehling-Schulz
Monika
M
D001409:Bacillus cereus / Q000235:genetics / Q000302:isolation & purification*; D052160:Benzothiazoles; D017931:DNA Primers / Q000235:genetics; D003937:Diagnosis, Differential; D003959:Diamines; D004196:Disease Outbreaks; D005516:Food Microbiology; D005517:Foodborne Diseases / Q000175:diagnosis* / Q000453:epidemiology / Q000382:microbiology*; D006801:Humans; D009930:Organic Chemicals / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D011804:Quinolines; D012015:Reference Standards; D012680:Sensitivity and Specificity; D013194:Staining and Labeling / Q000379:methods
D052160:Benzothiazoles; D017931:DNA Primers; D003959:Diamines; D009930:Organic Chemicals; D011804:Quinolines; C098022:SYBR Green I