从湖羊肺脏中分离绵羊肺炎支原体的鉴定  

Identification of a strain of Mycoplasma ovipneumonia from the lung of a neumonia sheep

作　　者：李媛[1] 陶岳 阿依吐拉.肉孜 高玉龙[1] 尹训南[1] 李新萍 张孝恩 王砚范[1] 辛九庆[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/国家牛传染性胸膜肺炎参考实验室,黑龙江哈尔滨150001 [2]新疆石河子畜牧兽医站,新疆石河子832000 [3]新疆维吾尔自治区动物防疫监督总站,新疆乌鲁木齐830063

出　　处：《中国预防兽医学报》2006年第4期375-379,共5页Chinese Journal of Preventive Veterinary Medicine

摘　　要：从新疆湖羊肺脏病料中分离出一株支原体XJ-3f,用其培养物人工感染80日龄健康绵羊,28d后剖杀,剖检可见肺表面肉粉色实变,显微病理变化为大灶性融合性肺炎。参考国际已知支原体16SrRNA序列,设计一对扩增700bp片段的通用引物,直接提取肺脏组织DNA进行PCR扩增并克隆、测序。将该序列与GenBank中33种支原体序列比较,结果证明该序列与绵羊肺炎支原体(M.ovipneumonia)标准株Y-98同源性为99.9%,而与山羊支原体山羊亚种(M.capricolumsubsp.capricolum,Mcc)、丝状支原体山羊亚种(M.mycoidessubsp.Capri,Mmc)、丝状支原体丝状亚种LC型(M.mycoidessubsp.mycoidesLC,M.mmLC)等同源率为81%。以感染羊血清和Y-98与从发病羊肺脏中分离出的三株支原体菌体蛋白进行Westernblot,证明均与感染羊血清有特异性反应,且与Y-98相比无明显差异,故确定分离株为绵羊肺炎支原体(M.ovipneumonia)。One Mycoplasma isolate XJ-3f was isolated from the lung of diseased sheep with the respiratory symptoms. The strain was cultured in modified Hayrick Medium and the culture were infected intratracheal to 2 sheep of 80 days. Lobular catarrhal bronchopneumonia was established by postmortem examinations, 28 d from infection, and Mycoplasma was re-isolated from the lungs. We designed a pair of primers to amplify a 700 bp fragment of 16S rRNA gene of all Mycoplasma. The genomic DNA of lung were extracted, amplified by the primers and cloned into the pMD18-T vector. 33 reference strains were loaded down from GenBank, the result of sequence analysis indicated that the isolated strain, XJ-3f, exhibited 99.9 % homology with reference strain Y-98, 81% with reference strains Mccp, MmmLC, Mcc ,Mmc. The result of Westem blot showed that they are all recognized by the serum as seem as Y-98. The results indicated that the isolated strain XJ-3f is belong to the Mycoplasma ovipneumonia.

关 键 词：绵羊肺炎支原体 16 S RRNA 鉴定 

分 类 号：S852.65[农业科学—基础兽医学]