青藏高原牦牛携带屎肠球菌及其毒力基因和耐药性检测  

Virulence genes and antibiotic resistance of Enterococcus faecium isolated from yaks in Qinghai-Tibet Plateau,China

作　　者：周娟[1] 金东[1] 卢珊[1] 濮吉[1] 白向宁[1] 杨晶[1] 胡守奎[1] 徐建国[1] 

机构地区：[1]中国疾病预防控制中心传染病预防控制所,传染病预防控制国家重点实验室,北京102206

出　　处：《中国人兽共患病学报》2015年第4期298-302,共5页Chinese Journal of Zoonoses

基　　金：国家自然科学基金重大课题(No.81290340,No.81290345)~~

摘　　要：目的了解青藏高原牦牛携带屎肠球菌(Enterococcus faecium)情况,以及耐药性和毒力基因。方法使用链球菌培养基分离细菌;使用生化反应和16srDNA序列分析方法鉴定;采用PCR方法检测从牦牛粪便中分离的屎肠球菌携带细胞溶血素(cytolysin,cylA)、明胶酶E(gelatinase,gelE)、表面蛋白(enterococcal surface protein,esp)、胶质蛋白粘附素collagen-binding-adhesin of Enterococcal faecium,acm)、聚集物质(aggregation substance,asa1)及透明质酸酶(hyalronidase,hyl)6种毒力基因的情况;应用K-B纸片法对屎肠球菌分离株进行16种抗生素的敏感性分析;参照PulseNet脉冲场凝胶电泳(Pulsed-field gel electrophoresis,PFGE)实验方法对屎肠球菌分离株进行PFGE分型分析。结果我们从320份牦牛粪便分离到33株屎肠球菌。这些菌株中毒力基因asa1的阳性率最高,为6.1%,acm和hyl次之,均为3.0%,其他毒力基因皆未检出。33株牦牛屎肠球菌中,有19株菌株具有抗生素耐药性,包括4株多重耐药菌株和15株单耐菌株。牦牛屎肠球菌对利福平耐药率最高,为48.5%;对其他抗生素的耐药率从高到低依次为青霉素G15.2%、四环素12.1%、强力霉素12.1%、红霉素9.1%、环丙沙星6.1%、氨苄西林6.1%、高浓度庆大霉素6.1%、磷霉素6.1%、左氧氟沙星6.1%。所有菌株对万古霉素、替拉考宁和氯霉素均敏感。细菌染色体DNA酶切片段的PFGE分析发现,33株屎肠球菌分离株共产生30种PFGE带型,可分为A-H 8个聚类群,耐药菌株分布在6个聚类群中。结论青藏高原牦牛携带屎肠球菌,呈现高度遗传多态性,部分菌株携带毒力基因,对多种抗生素耐药。研究提示,屎肠球菌可能会通过牦牛相关食品传播。To isolate Enterococcus faeciurn from yaks in Qinghai-Tibet Plateau, China and detect its virulence genes and an- tibiotic resistance, the strains were isolated by Streptococcal medium and were identified by biochemical and 16s rDNA meth- ods. The strains were then analyzed by pulsed-field gel-electrophoresis of digested chromosomal DNA fragments. The virulence genes were detected by PCR and confirmed by sequencing. Antibiotic resistance was performed by the K-B disk method with 16 antibiotics that were widely used in E. faecium. Results showed that 33 E. faeciurn strains were isolated from 320 fecal sam- ples of yaks. A total of six virulence genes were detected by PCR method, including cytolysin （cylA）, gelatinase （gelE）, en- terococcal surface protein （esp）, collagen-binding-adhesin （acre）, aggregation substance （asal） and hyalronidase （hyl）. The gene asal was found in 6.1% strains tested, followed by acm and hyl （3.0 %）. None of isolates was tested positive for the rest three virulence genes. Nineteen of 33 isolates were resistant to antibiotics, including 4 isolates having multidrug-resistance and 15 isolates being resistant to only one antibiotic. Of the 16 antibiotics tested, rifampicin resistance was found to be the most prevalent, accounting for 48.5 % of isolates, followed by penicillin G （15.2 ~）, tetracycline （12.1%）, doxycycline （12.1%）, erythromycin （9.1%）, ampieillin （6.1%）, high-level gentamicin （6.1%）, ciprofloxacin （6.1%）, fosfomycin （6.1G）, and levofloxacin （6.1%）. All of the isolates were 100% susceptible to vancomycin, teicoplanin and chloromycetin. Thirty-three E. faecium strains displayed 30 PFGE patterns and then were divided into 8 clusters from A to H, 6 of which included antibiotic-resistant strains. The yaks in the Qinghai-Tibetan Plateau are releasing E. faecium through feces. Those isolates car ried some virulence genes and were resistant to various anti biotics, indicating potential hazard to human health.

关 键 词：屎肠球菌 毒力基因 耐药性 脉冲场凝胶电泳 牦牛 

分 类 号：R378.1[医药卫生—病原生物学]