宠物医院犬猫源肠球菌的耐药性检测及其耐药基因optrA的流行特征分析

Investigation for antibiotics resistance of pet Enterococcus in Jilin province and the epidemiological characteristics of optrA gene

为了解吉林地区宠物源肠球菌的耐药情况和噁唑烷酮类耐药基因optrA在宠物源肠球菌中的流行分布特征,本研究对来自长春市和吉林市4家宠物医院的200份宠物肛门拭子样品,利用肠球菌琼脂培养基进行细菌的分离,利用16S rRNA基因的PCR扩增及测序进行菌种的鉴定,采用琼脂稀释法对11类13种抗菌药物进行药敏试验,利用PCR检测酰胺醇类耐药基因fexA、fexB和噁唑烷酮类耐药基因cfr、optrA,并对携带optrA基因的菌株进行全基因组测序。结果显示,本研究分离到96株肠球菌;药敏试验结果显示,宠物源肠球菌对克林霉素、四环素、沃尼妙林、庆大霉素和红霉素耐药严重,耐药菌株分别占87.5%、76.0%、72.9%、65.6%和55.2%,其次对环丙沙星、青霉素、氨苄西林和利福平耐药的菌株分别占33.3%、24.0%、20.8%和12.5%,对氟苯尼考耐药的菌株相对较少,占5.2%,检测到1株对万古霉素耐药的屎肠球菌,未检测到对利奈唑胺和替加环素耐药的菌株。PCR检测到3株同时携带optrA及fexA耐药基因的粪肠球菌,且其均呈多重耐药性。对携带optrA基因肠球菌的全基因组测序结果显示,3株粪肠球菌分别为ST376型(n=2)和ST16型(n=1),且不同ST型的粪肠球菌携带的耐药基因存在显著差异,optrA基因常与fexA、tet、isaA等耐药基因协同传播。研究表明optrA基因在宠物源肠球菌中已经存在一定程度的流行,且人与宠物接触密切,易造成耐药菌在两者之间的传播,应加强对宠物源肠球菌耐药性的监测。

In order to understand the antibiotics resistance of pet-derived Enterococcus and the prevalence and distribution characteristics of the oxazolidinones-resistant gene optrA,two hunderd pet feces samples provided by 4 pet clinics in Changchun and Jilin were collected.Bacteria isolation was performed by using Enterococcus agar medium and the 16S rRNA was amplified and identified by sequencing.We tested 11 classes of 13 antibacterial drugs susceptibility by agar dilution method.The amphenicols-resistant genes of fexA,fexB,cfr and optrA were detected by PCR.The positive strains carrying optrA gene were subjected to whole-genome sequencing.In this study,we obtained 96 Enterococcus.Most of them were highly resistant to Clindamycin,Tetracycline,Vornimulin,Gentamicin,and Erythromycin,and the resistance rates were 87.5%,76.0%,72.9%,65.6%and 55.2%,respectively.The resistance rates of Ciprofloxacin,Penicillin,Ampicillin and Rifampicin were 33.3%,24.0%,20.8%and 12.5%,respectively.The proportion of strains resistant to Florfenicol was 5.2%.One strain was detected to be resistant to Vancomycin.Linezolid-and tigecycline-resistant Enterococcus strains were not detected.Three Enterococcus faecalis strains were detected by PCR for carrying both optrA and fexA genes showing multi-drug resistance.The whole-genome sequencing analysis of Enterococcus carrying the optrA gene showed that the three optrA-positive E.faecalis strains were ST376(n=2)and ST16(n=1),respectively.The resistance genes carried by different ST-type E.faecalis were significantly different.The optrA gene usually spreads cooperatively with other drug resistance genes such as fexA,tet,and isaA.Studies have shown that the optrA gene has a certain degree of prevalence in pet-derived Enterococcu.Moreover,the close daily contact between people and pets can easily lead to the spread of antibiotics-resistant bacteria between them.The monitoring of the drug resistance of pet-derived Enterococcus should be strengthened.