耐甲氧西林葡萄球菌牛乳分离株耐药特征和分型研究  

Study on drug resistance characteristics and typing of methicillin-resistant Staphylococcus milk-derived isolates

作　　者：胡嘉宁 钟普鹏 杨培川 赵瑞利 秦顺义 孙英峰 马吉飞 HU Jianing;ZHONG Pupeng;YANG Peichuan;ZHAO Ruili;QIN Shunyi;SUN Yingfeng;MA Jifei(Changchun Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Changchun 130122,China)

机构地区：[1]天津农学院动物科学与动物医学学院天津市农业动物繁育与健康养殖重点实验室,天津300384

出　　处：《黑龙江畜牧兽医》2022年第1期82-91,136,137,共12页Heilongjiang Animal Science And veterinary Medicine

基　　金：河北省重点研发计划项目(19227523D);天津市生猪产业技术体系创新团队项目(ITTPRS2018004)。

摘　　要：为了研究天津地区牛源耐甲氧西林葡萄球菌(methicillin-resistant Staphylococci,MRS)的耐药特征和分型情况,试验通过选择培养、溶源检测、革兰氏染色、PCR检测、测序分型分析、耐药基因和毒力基因检测、药敏试验对从天津市郊4个区5所规模牛场126头泌乳牛采集的499份乳样进行研究。结果表明:临床型乳房炎患牛17头,仅隐性乳房炎患牛55头,完全健康牛54头,χ;检验显示患不同程度乳房炎牛的数量与牛龄存在相关性;经BP固体培养基培养,可见黑色菌落,革兰氏染色阳性;共分离得到370株葡萄球菌属菌株,鉴定出9种MRS,共58株(占比15.7%),含耐甲氧西林金黄色葡萄球菌(methicillin-resistant Staphylococcus aureus,MRSA)7株(占比1.9%)、耐万古霉素葡萄球菌(vancomycin-resistant Staphylococci,VRS)5株(占比1.4%);鉴定出6种SCCmec分型,Ⅰ型占1.7%、Ⅱ型占13.8%、Ⅲ型占12.1%、Ⅳ型占24.1%、Ⅴ型占46.6%、Ⅻ型占1.7%;金黄色葡萄球菌共计67株,溶源菌占金黄色葡萄球菌分离株的74.6%、coa基因阳性分离株占71.6%、pvl基因阳性分离株占28.4%;MRSA的有效spa分型中t6172型居多(占42.8%),其余分型为t007、t233、t3523型;甲氧西林敏感金黄色葡萄球菌(methicillin-sensitive Staphylococcus aureus,MSSA)的有效spa分型中t034型居多(占31.7%),其余分型为t189、t164、t899、t377、t127、t159、t409、t745型;通过与NCBI数据库比较,有1株MRSA的spa分型为t3523型,SCCmec分型为Ⅻ型;spa分型MRSA分离株的系统发育树可分为4个不同群组,即3株t899型、3株t6172型、t007型和t355型、t437型和t3523型,t233型单独为一支;有7株MRSA对利奈唑胺完全敏感和对头孢西丁完全耐药,有5株MRSA对万古霉素敏感,各有1株MRSA对万古霉素中介和耐药。说明MRS牛乳分离株呈现遗传多样性,其中MRSA分离株具有多药耐药特征;MRSA-t3523-SCCmecⅫ牛乳分离株是一株spa分型和SCCmec分型的新组合。In order to study the prevalence of bovine-derived methicillin-resistant Staphylococcus(MRS) in Tianjin area, the experiment used selective culture, lysogen detection, Gram staining, PCR detection, sequencing typing analysis, antibiotic resistance gene and virulence gene detection, and drug sensitivity test to study 499 milk samples collected from 126 lactating cows from five large-scale cattle farms in four areas in the suburbs of Tianjin City. The results showed that there were 17 cows with clinical mastitis, 55 cows with subclinical mastitis, and 54 completely healthy cows;the χ;test showed a correlation between the number of cows with different levels of mastitis and their age. After culturing on BP solid medium, black colonies could be seen, and Gram staining was positive. A total of 370 non-repetitive Staphylococcus strains were isolated, of which nine species containing a total of 58 MRS strains were identified(accounting for 15.7%), including seven strains of methicillin-resistant Staphylococcus aureus(MRSA, accounting for 1.9%), and five strains of vancomycin-resistant Staphylococcus(VRS, accounting for 1.4%). SCCmec typing identified six types, of which type Ⅰ accounted for 1.7%, type Ⅱ accounted for 13.8%, type Ⅲ accounted for 12.1%, type Ⅳ accounted for 24.1%, type Ⅴ accounted for 46.6%, and type Ⅻ accounted for 1.7%. A total of 67 strains of Staphylococcus aureus were obtained, including lysogenic bacteria accounting for 74.6% of Staphylococcus aureus isolates, coa gene positive isolates accounting for 71.6%, pvl gene positive isolates accounting for 28.4%. Most of the effective spa typing of MRSA was t6172(42.8%), and the remaining types were t007, t233, and t3523;most of the effective spa typing of methicillin-sensitive Staphylococcus aureus(MSSA) was t034(31.7%), and the rest types were t189, t164, t899, t377, t127, t159, t409, t745;by comparison with the NCBI database, one strain of MRSA had a spa typing of t3523 and an SCCmec typing of Ⅻ;the spa gene phylogenetic tree of MRSA isol

关 键 词：奶牛乳房炎 耐甲氧西林葡萄球菌 耐药基因 毒力基因 葡萄球菌染色体盒 基因分型 

分 类 号：S852.611[农业科学—基础兽医学]