To establish a specific and sensitive TaqMan-MGB quantitative real-time PCR assay for the rapid detection of Cronobacter spp..MethodsBased on the conservative sequence of partial macromolecular synthesis operon gene of Cronobacter spp. published on GenBank, specific primers and TaqMan Minor groove binder (TaqMan-MGB) probes were designed, and the rapid real-time PCR assay was estabilished and optimized. The specificity was evaluated with 25strains of other Enterobacteriaceae and some common pathogens. The quantitative standard curve was established with the recombinant plasmids and the sensitivity for the assay was evaluated for recombinant plasmids, pure cultures and contaminated food samples. Comparing with the TaqMan real-time PCR recommended by U.S. FDA, paired-samples t-test for the variables of cycle threshold (Ct) and relative fluorescence intensity (△Rn) was done between the two methods. ResultsThe TaqMan-MGB quantitative real-time PCR assay could be finished detection in 40minutes. It was specific enough to discriminate Cronobacter spp. from all other Enterobacter and non-Enterobacter strains tested. The relative coefficient of the quantitative standard curve was 0.999, and the amplification efficiency of the quantitative standard curve was 99.972%. The sensitivity for the assay was 10copies per reaction for recombinant, 3.8cfu/ml for pure culture, and 38cfu/ml for contaminated food samples,respectively. There were statistical differences between two real-time PCR methods by paired-samples t-test (Ct:t=-14.406, P<0.01and △Rn:t=14.230, P<0.01). The TaqMan-MGB real-time PCR was better than the TaqMan real-time PCR recommended by U.S. FDA in sensitivity and resolution.ConclusionThe TaqMan-MGB quantitative real-time PCR assay targeted the partial macromolecular synthesis operon gene of Cronobacter spp. is rapid, specific and sensitive. It would had a good value in the screening and identification of Cronobacter spp. from infant milk powder for food safety and risk monitor.