生牛乳中金黄色葡萄球菌异质性耐药及机制研究  

Investigation of Heteroresistance and Its Mechanism of Staphylococcus aureus in Raw Milk

作　　者：和晓兰 赵艳坤 孟璐[1] 刘慧敏[1] 高姣姣 郑楠[1] HE Xiaolan;ZHAO Yankun;MENG Lu;LIU Huimin;GAO Jiaojiao;ZHENG Nan(Institute of Animal Sciences,Chinese Academy of Agricultural Sciences,Beijing 100193,China;Key Laboratory of Agro-Products Quality and Safety of Xinjiang,Ministry of Agriculture and Rural Affairs-Laboratory of Quality and Safety Risk Assessment for Agro-Products,Institute of Quality Standards&Testing Technology for Agro-Products,Xinjiang Academy of Agricultural Sciences,Urumqi 830091,China)

机构地区：[1]中国农业科学院北京畜牧兽医研究所,北京100193 [2]新疆农业科学院农业质量标准与检测技术研究所农业农村部农产品质量安全风险评估实验室(乌鲁木齐)新疆农产品质量安全重点实验室,乌鲁木齐830091

出　　处：《畜牧兽医学报》2025年第4期1934-1946,共13页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家重点研发计划(2022YFD1301000);国家现代农业产业技术体系(CARS36);农产品质量安全风险评估(GJFP20220304);中国农业科学院科技创新工程(ASTIP-IAS12);天山创新团队(2022D14016);新疆奶产业技术体系专项经费资助(XJARS-11)。

摘　　要：旨在了解生牛乳中分离的金黄色葡萄球菌耐药情况以及金黄色葡萄球菌的异质性耐药流行情况,并探究异质性耐药机制。从东北、华北、华东、华南、西北和西南六个地区采集179批次生牛乳,分离鉴定出金黄色葡萄球菌36株。采用肉汤微量稀释法鉴定金黄色葡萄球菌的敏感性和耐药性,联合纸片扩散法和E-test法从抗菌药物敏感表型的菌株中筛选疑似异质性耐药菌株,利用菌群谱型分析法(PAP)进行确证,传代测定异质性耐药稳定性,通过全基因组和比较基因组学分析异质性耐药机制。结果表明,36株金黄色葡萄球菌对β-内酰胺类抗生素耐药率达11.11%~61.11%,对四环素和氨基糖苷类抗生素表现高度敏感。初筛得到19种金黄色葡萄球菌和抗菌药物组合异质性耐药,确证3种组合为异质性耐药表型,其中仅金黄色葡萄球菌J23保持稳定的异质性耐药表型。本研究表明,生牛乳源的金黄色葡萄球菌对β-内酰胺类和喹诺酮类抗菌药物耐药性较高,对阿莫西林/克拉维酸的异质性耐药机制与blaZ和金属β-内酰胺酶的表达有关,对四环类药物的异质性耐药与四环素外排泵表达和耐药蛋白有关。This experiment was conducted to study the drug resistances of Staphylococcus aureus isolated from raw milk,the prevalence of heteroresistance of Staphylococcus aureus and explore the mechanism of heteroresistance.Thirty-six Staphylococcus aureus isolates were isolated from 179 batches of raw milk collected from Northeast,North China,East China,South China,Northwest and Southwest.The sensitivity and resistance of Staphylococcus aureus were identified using broth microdilution method.The combination of disk diffusion method and E-test method was used to screen suspected heteroresistant strains from strains with sensitive phenotypes,and then the suspected heteroresistant strains were confirmed by Population analysis profiling(PAP).The stability of the heteroresistant phenotype after serial passage was measured.Finally,whole-genome sequencing and whole genome re-sequencing analysis were used to determine the heteroresistant mechanisms.The results showed that 36 strains of Staphylococcus aureus were resistant toβ-lactam antibiotics,the resistant rate to ranges from 11.11%to 61.11%,and had high sensitivity to tetracycline and aminoglycoside antibiotics.19 combinations of Staphylococcus aureus and antibiotics with heteroresistant were identified through initial screening,and it was confirmed that the 3 combinations exhibited heteroresistant phenotypes,of which only amoxicillin-clavulanate-J23 heteroresistant stabled after serial passage.This study demonstrates that S.aureus of raw milk origin is high resistant toβ-lactam and quinolone antimicrobials.Heteroresistant mechanisms to amoxicillin-clavulanic acid are associated with blaZ and metalβ-lactamase expression.Heteroresistant mechanisms to tetracyclines is associated with tetracycline efflux pumps and resistance proteins.

关 键 词：生牛乳 金黄色葡萄球菌 异质性耐药 耐药亚群 全基因组学 

分 类 号：S852.611[农业科学—基础兽医学]