Improved multiplex polymerase chain reaction (PCR) detection of Bacillus cereus group and its toxic strains in food and environmental samples

A multiplex PCR assay for the rapid detection of Bacillus cereus group, enterotoxic and emetic strains was developed. A panel of emetic and enterotoxic reference strains, B. cereus group members and non-target strains were used for the evaluation of the assay. Verification of PCR results on pure culture and inoculated foods successfully confirmed the specificity of approach for detection of target genes for B. cereus group (groEL), diarrheal (cytK, nheA, hblC, entFM) and emetic strains (CER). The sensitivity of approach was satisfying in pure culture as 20 pg of DNA per reaction tube. Artificial contamination of seven different food matrices with distinct bacterial counts revealed a minimum detection limit of 103 cfu/g in food samples. The detection limits were improved to approximately 101 cfu/g after 7 h enrichment. Natural contamination of rice and kimbab as well as environmental samples (soil, cow feces) was studied. The incidence of B. cereus was 63.88 and 38.88% in rice and kimbab, and 84.61 and 69.23% in soil and feces, respectively. To the best of our knowledge, this is the first time that an assay for simultaneous detection of B. cereus group, emetic and enterotoxic strains with such a wide range of detection target genes in food and environmental samples has been described.