应用微滴数字PCR技术快速检测食用菌中沙门氏菌

Rapid Detection of Salmonellla spp. in Edible Fungi by Droplet Digital PCR

定量风险评估是整个风险评估体系的发展趋势和终极体现形式,而快速、准确、简便的定量技术则是保障整个体系顺利、有效完成的核心支撑。作者根据沙门氏菌inv A毒力基因序列,在前人对引物研究结果基础上,合成特异性引物和探针。以沙门氏菌标准菌株为研究对象,建立沙门氏菌微滴数字PCR的快速定量检测方法,并对其特异性、灵敏度、样品实测等关键指标与传统培养法进行对照验证。结果表明,所建立的微滴数字PCR方法具有较好的特异性,检测灵敏度可达到10~2CFU/m L,样品实测与传统国标法对比,二者符合率100%。所建立的沙门氏菌微滴数字PCR定量检测方法,可以快速、准确、直接地分析沙门氏菌的含量,无需构建质粒和标准曲线,避免了因标准曲线量值偏差而影响样品定值的准确性,所建立的方法为食源性沙门氏菌污染的快速定量检测提供了技术支撑,为微生物风险评估体系的完善奠定了基础。

Quantitative Risk Assessment （QRA） is the future trend of the risk assessment system in which simple,rapid and accurate quantitative technologies made crucial contributions to the smooth and efficient accomplishment of QRA.In this study,a method for detecting the virulence-specific gene,invA in Salmonellla spp.was established by using droplet digital PCR technology.In contrast to the traditional method of national standard （GB 4789.4-2010） for Salmonellla detection,our developed method showed higher specificity and sensitivity as low as 102 CFU/mL.The coincidence rate was 100 % as compared with simulation specimens with culture strains.The establishment of this method based on droplet digital PCR for Salmonella detection provided a rapid and accurate approach for identification of food-borne pathogens without the use of standard curves or plasmids.These results demonstrated that this developed method offered a promising way for rapid quantitative detection of Salmonella and provided technical support for the future studies of the microbial risk assessment system.