奶牛乳房炎源金黄色葡萄球菌的分离鉴定及耐药性分析

Isolation,idetification and drug resistance analysis of cow mastitis-derived Staphylococcus aureus

为了了解新疆石河子地区奶牛乳房炎源金黄色葡萄球菌的耐药表型及耐药基因携带情况,试验对采自石河子地区2个规模化奶牛场患乳房炎奶牛的乳样进行细菌分离,使用种特异性引物对分离株进行鉴定,并采用纸片琼脂扩散法、最小抑菌浓度(MIC)测定法和PCR方法对分离得到的金黄色葡萄球菌的耐药表型和耐药基因进行检测。结果表明:从患乳房炎奶牛的138份乳样中共分离出74株金黄色葡萄球菌,镜检为阳性球菌,经PCR扩增确定为金黄色葡萄球菌,分离率为53.62%(74/138)。所有分离株均有不同程度的耐药性,对万古霉素、甲氧西林、磺胺甲口恶唑、林可霉素、青霉素、恩诺沙星的耐药率分别为97.30%、82.43%、79.73%、71.62%、67.57%、67.57%,其他9种药物的耐药率为16.22%~59.46%;对庆大霉素和多西环素两种药物的敏感性较高,分别为52.70%、74.32%。耐3种及以上抗生素的菌株占90.54%,耐9种及以上抗生素的菌株占56.75%,其中1个分离株对所有检测的抗生素均耐受。74株分离株中共检测到blaZ、linA、ermB、aacA-aphD、aac6-aph2、aph(3′)-a和tetM 7种耐药基因,其中氨基糖苷类耐药基因aacA-aphD检出率最高,为8.10%,其余耐药基因的检出率在2.70%~4.05%之间。说明新疆石河子地区奶牛乳房炎源金黄色葡萄球菌普遍耐药,且多重耐药现象较为严重,建议临床上根据药敏试验结果科学治疗奶牛乳房炎。

In order to understand the drug resistance phenotype and the carrying of drug resistance genes of cow mastitis-derived Staphylococcus aureus in Shihezi area,in this experiment,bacteria isolation was conducted from milk samples of dairy cows with mastitis in two large-scale dairy farms in Shihezi area.Species-specific primers were used to identify the isolates;the detection of the drug-resistant phenotypes and the drug-resistant genes of the obtained Staphylococcus aureus were carried out by K-B method,MIC method and PCR method.The results showed that 74 isolates of Staphylococcus aureus were isolated from 138 milk samples of dairy cows with mastitis,which were positive cocci by microscopic examination,and identified as Staphylococcus aureus by PCR amplification,with an isolation rate of 53.62%(74/138).All isolates had drug resistance in varyous degrees.The drug resistance rates of vancomycin,methicillin,sulfamethoxazole,lincomycin,penicillin and enrofloxacin were 97.30%,82.43%,79.73%,71.62%,67.57% and 67.57%,respectively.The drug resistance rates of the other nine drugs were 16.22%-59.46%.The sensitivity to the two drugs gentamicin and doxycycline was 52.70% and 74.32% respectively.90.54% of the isolates were resistant to three or more antibiotics,and 56.75% of the isolates were resistant to nine or more antibiotics;one isolate was resistant to all test antibiotics.Seven resistance genes,including blaZ,linA,ermB,aacA-aphD,aac6-aph2,Aph(3′)-a and tetM,were detected in the 74 isolates.Among them,the detection rate of aminoglycoside resistance gene aacA-aphD was the highest of 8.10%,and the detection rates of other drug resistance genes were 2.70%-4.05%.The results suggested that the drug resistance of cow mastitis-derived Staphylococcus aureus existed widespread in Shihezi area,Xinjiang Uygur Autonomous Region,and the phenomenon of multiple drug resistance was serious.A recommendation was given that the scientific clinical treatment of dairy cow mastitis should be based on the results of drug sensitivity test.