为研究牛源金黄色葡萄球菌耐药性以及甲氧西林敏感(MSSA)和耐甲氧西林(MRSA)菌携带的耐药基因、MRSA的SCCmec基因型,揭示牛源MSSA与MRSA之间的演化相关性和MRSA起源和扩散途径,对2009年以来中国五省区不同牛场分离的54株金黄色葡萄球菌进行了药敏试验.对确定的12株MSSA和MRSA进行了耐药基因检测、SCCmec基因型分型和多位点测序分型(MLST)研究.54株菌的药敏结果显示,对红霉素、克林霉素、青霉素、复方新诺明、多西环素、四环素、氯霉素、环丙沙星、庆大霉素的耐药菌株分别为88.8％、81.5％、88.9％、90.7％、92.6％、94.4％、79.6％、63.0％和70.4％.其中,对所测10种抗生素完全耐药的占5.6％,对5种以上耐药的占85.2％,6株(11.1％)对头孢西丁耐药.12株MSSA和MRSA菌株的药敏和耐药基因检测结果显示,3株MRSA对10种抗生素耐药；6株MSSA菌中,新疆分离株对2～4种抗生素耐药,其它省区分离株对7种以上抗菌素耐药.12株MSSA和MRSA菌均检出ermC和tetK基因；6株MRSA中均检出aac(6′)/aph(2")基因,5株检出tetM基因,4株检出aph(3′)-Ⅲ基因;6株MSSA均未检出aac(6′)/aph(2")和aph(3′)-Ⅲ,4株检出tetM基因.用多重PCR对MRSA进行SCCmec基因分型显示,5株为SCCmec Ⅳ,1株未能分型；12株菌的MLST分型发现,所有菌株分为ST50、ST965、ST6、ST97和ST9序列型.经eBURST v3分析,MRSA分布在CC7、CC4、CC21和CC9四个克隆复合群(CCs)中,MSSA分布在CC7和CC32克隆复合群中.以上研究表明,我国牛源金黄色葡萄球菌不仅耐药严重,且表现多重耐药；MRSA基因型以SCCmec Ⅳ为主.对牛源SCCmec Ⅳ型MRSA与同型人源MRSA的耐药谱分析发现有较大差异,推测位于质粒或转座子上的耐药基因可能插入到金黄色葡萄球菌基因组中,导致耐药基因在不同菌株间传播.根据ST97中有MSSA和MRSA以及牛源MRSA与人源MRSA的MLST比较,确认我国牛源MRSA是在抗生素的选择压力下由来源于不同克隆复合群的MSSA获得SCCmec Ⅳ而产生,推测同一克隆株MRSA的扩散并不是MRSA大范围出现的主要原因.%The aim of this study was to investigate the current status of the antimicrobial resistance of bovine Staphylococcus aureus isolates, so as to do research on carried drug-resistant genes and molecular epidemiology profiles of methicillin-susceptibility Staphylococcus aureus (MSSA) and methicillin-resistant Staphylococcus aureus(MRSA) which isolated from bovine, further, to reveal the evolutionary relationship between MSSA and MRSA, and origin and diffusion of MRSA. Total 54 isolates which isolated from Xinjiang, Zhejiang, Shandong, Neimenggu and Shanghai in 2009 were investigated antibiotics susceptibility by disc diffusion method, and 12 isolates of MSSA and MRSA which confirmed by Cefoxitin sensitive test by disc diffusion method and mecA PCR were used to detected the antimicrobial resistance genes and typed by multilocus sequence typing (MLST). Drug sensitive test results showed that 88. 8% isolates resisted to erythromy-cin, 81. 5% to clindamycin, 88. 9% to penicillin, 90. 7% to ulfamethoxazole compound, 92. 6% to doxycycline, 94. 4% to tetracycline, 79. 6% to chloramphenicol, 63. 0% to ciprofloxacin, 70. 4% to gentamicin. Besides that, 5. 6% isolates resisted to all 10 antibiotics, 85. 2% isolates resisted to more than 5 antibiotics, 6 isolates (11. 1%) resisted to cefoxitin. Furthermore, 3 MRSA isolates resisted to 10 antibiotics, Xinjiang MSSA isolates resisted to 2-4 antibiotics, other provinces MSSA isolates resisted to 7 antibiotics, and analyzed result of the antimicrobial resistance gene of MSSA and MRSA, found all isolates carried ermC and tetK genes, but no ermA gene, 6 MRSA isolates carried aac(6')/aph(2"), 5 and 4 MRSA isolates carried tetM and aph (3')- Ⅲ respectively, but only 4 MSSA isolates carried tetM gene, no MSSA isolates carried aac (6')/aph(2") and aph(3')-Ⅲ. Staphylococcal chromosomal cassette (SCC) mec of MRSA was typed by multi-PCR, the results showed that 5 isolates were SCCmec Ⅳ and 1 isolate can not identificated type. The MLST results of MSSA and MRSA proved that the isolates belonged to ST50, ST965,ST6,ST97 and ST9, further analysis found that 6 MRSA isolates distributed in CC7,CC4,CC21 and CC9 and MSSA isolates distributed in CC32 and CC7. The results suggest that drug resistance of Chinese bovine Staphylococcus aureus isolates are serious and most isolates represent multidrug resistance, MSSA and MRSA are multidrug-resistant strains and carries more than 2 antimicrobial resistance genes, epidemical bovine MRSA is MRSA-SCCmec Ⅳ. By analyzing the results of MLST and comparing with MLST results of human MRSA, found that, in China, bovine MRSA was different with human's and speculated that bovine MRSA emerged when MSSA which belonged to different CCs gained SCCmec Ⅳ by antibiotic selection, in addition, MRSA stains isolated from different areas were not the same ancestor, further analysis, presume those genes similar to plasmid or transposon can free communication in different isolates than depend on SCCmec.
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