Detection of Emetic Bacillus cereus by Real-Time PCR in Foods

Abstract

The simplex real-time PCR assays based on the TaqMan probe and SYBR green I which target cereulide synthetase genes ces genes were used for rapidly, reliably and sensitively identifying the emetic strains from among Bacillus cereus strains isolated from different sources. Only the emetic strains showed positive reactions to the real-time PCR assays, but all examined strains of diarrheal B. cereus, other Bacillus species and other gram positive and negative bacteria gave negative results. The final identification of emetic B. cereus was possible within 1 to 1.5 h in both simplex real-time PCR procedures. The detection limit of emetic strains in food was suggested to be 10 4 -10 5 cfu/g. Both simplex real-time PCR assays were found to be rapid, sensitive and reliable diagnostic tools that complement the different cellular, immunological and chemical detection methods for cereulide-producing B. cereus, even if a relatively expensive device is required for the assays.

Comparative analysis of the sensitivity of metagenomic sequencing and PCR to detect a biowarfare simulant (Bacillus atrophaeus) in soil samples

“…In S2 Table, for comparative purposes values from the literature [5–6, 8, 11, 12, 19, 46–48] are presented as absolute amounts ( i . e .…”

“…Amplification monitoring can be performed using the intercalating dye SYBR Green or fluorogenic probes (such as TaqMan probes), the later detection system being much more specific than the former. Real-time PCR assays, including assays compatible with handheld devices [8], are available for a number of biothreat agents such as ricin [9], Bacillus anthracis [10–11], Yersinia pestis [11], and emetic Bacillus cereus [12]. …”

Microbiological safety of industrially reared insects for food: Identification of bacterial endospores and targeted detection of foodborne viruses

“…qPCR reaction conditions for the panC detection followed the master mix supplier’s protocol: 2 min at 50 °C (UNG activation) and 2 min of initial denaturation at 95 °C, followed by 40 cycles of 15 s denaturation at 95 °C and combined annealing and elongation for 1 min at 60 °C. For the cesA detection, the same conditions were applied, except for the initial denaturation step which was shortened to 20 seconds 51 . Each qPCR was followed by a melt curve analysis by raising the temperature gradually from 60 to 95 °C (0.1 °C/s) while continuously monitoring fluorescence.…”

Rapid Testing of Food Matrices for Bacillus cereus Enterotoxins

“…; Ueda et al . ; Hariram and Labbe ). However, the most accurate quantitative method available for cereulide detection in contaminated food products is high performance liquid chromatography connected to mass spectrometry (LC/MS) (Biesta‐Peters et al .…”

“…Various methods using different cells lines and boar spermatozoa were evaluated, but results were subjective and sometimes inconclusive (Szabo et al 1991;Agata et al 1994;Hughes et al 1998;Jaaskelainen et al 2003). The genetic locus encoding cereulide biosynthetase (ces) in the emetic B. cereus strains has been identified and sequenced (Ehling-Schulz et al 2004;Horwood et al 2004;Dommel et al 2010) which has been a key factor for the development of molecular assays used for strain characterization and food testing (Ehling-Schulz et al 2006;Fricker et al 2007;Ueda et al 2013;Hariram and Labbe 2015). However, the most accurate quantitative method available for cereulide detection in contaminated food products is high performance liquid chromatography connected to mass spectrometry (LC/MS) (Biesta-Peters et al 2010;International Organization for Standaradization 2014).…”