食品科学 ›› 2019, Vol. 40 ›› Issue (2): 281-286.doi: 10.7506/spkx1002-6630-20180110-124

• 安全检测 • 上一篇    下一篇

牛奶源金黄色葡萄球菌血清型、毒力基因及PFGE分型

刘保光1,蔡田1,李小申1,刘营营1,贺丹丹1,匡秀华2,高延玲3,胡功政1,*   

  1. (1.河南农业大学牧医工程学院,河南?郑州 450002;2.河南牧业经济学院制药工程学院,河南?郑州 450046;3.河南省兽药监察所,河南?郑州 450008)
  • 出版日期:2019-01-25 发布日期:2019-01-22
  • 基金资助:
    “十三五”国家重点研发计划重点专项(2016YFD0501304);国家自然科学基金面上项目(31772799)

Serotyping, Virulence Genes and Pulsed Field Gel Electrophoresis (PFGE) Typing of Staphylococcus aureus Isolated from Fresh Milk

LIU Baoguang1, CAI Tian1, LI Xiaoshen1, LIU Yingying1, HE Dandan1, KUANG Xiuhua2, GAO Yanling3, HU Gongzheng1,*   

  1. (1. College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, China; 2. College of Pharmaceutical Engineering, Henan University of Animal Husbandry and Economy, Zhengzhou 450046, China; 3. Henan Supervision Institute of Veterinary Drug and Feed, Zhengzhou 450008, China)
  • Online:2019-01-25 Published:2019-01-22

摘要: 目的:研究生鲜牛奶中金黄色葡萄球菌分离、荚膜多糖血清型分布、毒力基因携带及脉冲场凝胶电泳(pulsed field gel electrophoresis,PFGE)分型情况。方法:从河南省4?个地区奶牛养殖场采集生鲜牛奶样品按照国标法进行分离,扩增耐热核酸酶基因nuc鉴定金黄色葡萄球菌,利用聚合酶链式反应方法测定荚膜多糖血清型和毒力基因携带情况,采用PFGE分析菌株间的相关性和遗传关系。结果:从350?份生鲜牛奶样品中分离鉴定到80?株金黄色葡萄球菌,分离率为22.86%。荚膜多糖血清型测定发现,cap5(60%)是流行血清型。从这些阳性菌株中,发现有62?株(77.5%)携带有毒力基因,毒力基因set、hlb、hld、lukED、ebp、clfA和clfB,检出率分别为40.00%、51.25%、57.50%、60.00%、58.75%、57.50%和58.75%。此外,47?株(58.75%)菌携带不少于6?个毒力基因,流行的毒力基因谱型为set-hla-hlb-hld-lukED-cna-ebp-clfA-clfB。PFGE结果显示,获得72?株菌的PFGE图谱,按90%的相似性可分为12?个簇和46?种PFGE型。D簇(3?种PFGE型)、G簇(3?种PFGE型)和J簇(5?种PFGE型)菌株中均检出一定基因类型的毒力基因,表明河南地区生鲜牛奶中金黄色葡萄球菌毒力基因广泛存在于多种PFGE型别中。结论:生鲜牛奶均有一定程度的金黄色葡萄球菌污染,多数菌株携带毒力基因,且毒力基因的类型较为复杂,这对消费这些牛奶的人群构成潜在的健康威胁。PFGE分型菌株主要以克隆形式进行传播,且克隆型具有多样性和差异性,故临床应加强生奶及乳品血清型、毒力基因检测及分子分型研究。

关键词: 牛奶, 金黄色葡萄球菌, 血清型, 毒力基因, 脉冲场凝胶电泳

Abstract: Objective: To investigate the serotyping, virulence genes and pulsed field gel electrophoresis (PFGE) typing of Staphylococcus aureus in fresh milk for a better understanding of the epidemiological characteristics of S. aureus. Methods: S. aureus strains were isolated from fresh milk samples collected from dairy farms in four regions of Henan province and they were identified by PCR amplification of the thermostable nuclease gene (nuc). The capsular polysaccharide (Cap) serotypes and virulence genes were determined by PCR. The correlation and genetic relatedness among these S. aureu isolates were analyzed by PFGE. Results: A total of 80 S. aureus strains were isolated from 350 fresh milk samples, with a separation rate of 22.86%. Cap5 (60%) was found to be a popular serotype. Of the positive isolates, 62 (77.5%) carried virulence genes, and the prevalence of the virulence genes set, hlb, hld, lukED, ebp, clfA and clfB were 40.00%, 51.25%, 57.50% , 60.00%, 58.75%, 57.50% and 58.75%, respectively. In addition, 47 (58.75%) strains carried at least 6 virulence genes, set-hla-hlb-hld-lukED-cna-ebp-clfA-clfB were found to be the prevalent virulence genotypes. PFGE profiles of 72 strains were obtained. Based on a 90% similarity, they were divided into 12 clusters and 46 PFGE types. The strains in clusters D, G and J were detected to carry some virulence genotypes, suggesting that virulence genes widely existed in a variety of PFGE types of S. aureus isolated from fresh milk in Henan. Conclusions: Fresh milk samples were contaminated by S. aureu to a certain degree. Most of the isolates carried virulence genes, and the virulence genotypes were complicated, posing a potential threat to the health of consumers. The PFGE subtypes were mainly transmitted in the form of clones, and the clone types were diversified and differentiated. Therefore, further clinical studies are needed for serotyping, virulence gene detection and molecular typing of S. aureus in raw milk and dairy products.

Key words: milk, Staphylococcus aureus, serotype, virulence genes, PFGE

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