应用多重PCR快速鉴定自然发酵肉制品中6种葡萄球菌  

Rapid Identification of Six Staphylococcus Species in Fermented Meat Products Using Multiplex

作　　者：许随根 陈曦[1] 李家鹏[1] 李金春[1] 杨君娜[1] 熊苏玥 戚彪[1] 米瑞芳 黄鑫[1] 乔晓玲[1] 王守伟[1] XU Suigen;CHEN Xi;LI Jiapeng;LI Jinchun;YANG Junna;XIONG Suyue;QI Biao;MI Ruifang;HUANG Xin;QIAO Xiaoling;WANG Shouwei(Beijing Key Laboratory of Meat Processing Technology, China Meat Processing and Engineering Center,Beijing Academy of Food Sciences, China Meat Research Center, Beijing 100068, China)

机构地区：[1]中国肉类食品综合研究中心北京食品科学研究院国家肉类加工工程技术研究中心肉类加工技术北京市重点实验室,北京100068

出　　处：《食品科学》2018年第24期303-310,共8页Food Science

基　　金："十三五"国家重点研发计划重点专项(2018YFD0401200);北京市优秀人才项目(2017754154700G098)

摘　　要：肉葡萄球菌(Staphylococcuscarnosus)、木糖葡萄球菌(S.xylosus)、腐生葡萄球菌(S. saprophyticus)、表皮葡萄球菌(S. epidermidis)、马胃葡萄球菌(S. equorum)和松鼠葡萄球菌(S. sciuri)是发酵肉制品中常见的葡萄球菌,为准确、快速地检测和鉴定这些菌种,建立6种葡萄球菌的多重聚合酶链式反应(polymerase chain reaction,PCR)方法并进行验证。以上述菌种的gap、rpoB、SodA和SNc为靶基因,设计和筛选菌种特异性引物6对,优化PCR体系,建立6种葡萄球菌的多重PCR鉴定方法。实验结果显示多重PCR的基因组DNA检测灵敏度可达到4 pg,活菌检测灵敏度可达到2×10~2 CFU;采用建立的多重PCR方法对传统肉制品中分离的8株葡萄球菌进行鉴定,结果与16S rDNA序列分析、生理生化鉴定结果一致。建立的多重PCR方法具有较高的种间特异性,可快速、准确地用于发酵肉制品中肉葡萄球菌、木糖葡萄球菌、表皮葡萄球菌、马胃葡萄球菌、松鼠葡萄球菌和腐生葡萄球菌的检测和鉴定,具有较好的应用前景。A multiplex polymerase chain reaction(mPCR)assay was developed and validated for simultaneous detection of Staphylococcus carnosus,S.xylosus,S.saprophyticus,S.epidermidis,S.equorum and S.sciuri,which are commonly found in fermented meat products.In this study,six pairs of species-specific primers were designed targeting the gap,rpoB,SodA and SNc genes,and the PCR reaction system was optimized.The results showed that The sensitivity of the mPCR system was4pg for bacterial genomic DNA and2×10^2CFU for living cultures.Meanwhile,eight Staphylococcus strains from traditional fermented meat products were consistently identified by this mPCR assay and16S rDNA sequence and biochemical analysis.This mPCR method was species-specific and effective,and could be used in the rapid and accurate identification of S.carnosus,S.xylosus,S.epidermidis,S.equorum,S.sciuri and S.saprophyticus.Thus,it has great application prospects.

关 键 词：多重PCR 葡萄球菌 发酵肉制品 快速鉴定 

分 类 号：TS201.3[轻工技术与工程—食品科学]