摘要
为了解榆林地区部分养牛场牛支原体的感染情况,采集临床表现为咳嗽、呼吸困难或患有关节炎的病牛肺脏组织3份、鼻腔分泌物9份,患乳房炎的泌乳奶牛乳样10份,对病料采用支原体分离纯化培养,Dienes染色,生化鉴定,支原体16S rRNA和特异性基因uvrC PCR扩增与测序,并检测分离株对抗生素药物的敏感性。结果显示,支原体固体培养基表面产生了针尖状、圆形、中间隆起、橙色的“煎蛋样”菌落,共获得8个分离株。Dienes染色可见分离株菌落颜色为深蓝色。分离株不水解精氨酸、明胶、七叶苷,不发酵葡萄糖、甘露醇、乳糖,能还原2,3,5-氯化三苯基四氮唑。利用支原体16S rRNA通用引物和牛支原体uvrC特异性引物进行PCR扩增,分别得到长度为1500 bp和238 bp的目的条带。分离株16S rRNA测序比对,与国内地方支原体流行株XBY01、Hbei-1、Ningxia-1、Tibet-10和NM2012同源相似性最高,均达99.9%,与无乳支原体PG2的同源相似性在99.3%~99.7%之间,与绵羊肺炎支原体Y98的相似性在81.5%~82.7%之间,分离株之间的同源性达到99.9%以上。分离株uvrC测序比对,与牛支原体同源性均在99.3%~99.8%之间,与无乳支原体的同源性在80.3%以下。测序结果表明,分离株均为牛支原体。分离株对诺氟沙星、洛美沙星、阿奇霉素、阿莫西林、红霉素表现为耐药,对卡那霉素、庆大霉素、多西环素、四环素、环丙沙星表现为敏感。
To investigate the infection of Mycoplasma bovis in some cattle farms in Yulin area,3 lung tissues,9 nasal secretions and 10 milk samples from lactating cows with clinical manifestations of cough,dyspnea and arthritis were collected.The experiment involved isolation,purification,and cultivation of Mycoplasma from diseased materials,Dienes staining,biochemical identification,PCR amplification and sequencing of Mycoplasma 16S rRNA and specific gene uvrC,and detection of antibiotic sensitivity for isolated strains.The results showed that there were needle-like,round,middle raised,orange"fried egg"colonies on the surface of Mycoplasma solid medium,and 8 isolates were obtained.The colony color of the isolates was dark blue by Dienes staining.The isolates did not hydrolyze arginine,gelatin and esculin,did not ferment glucose,mannitol and lactose,and could reduce 2,3,5-triphenyltetrazolium chloride.PCR amplification was performed using Mycoplasma 16S rRNA universal primers and mycoplasma bovis uvrC specific primers to obtain 1500 bp and 238 bp target bands,respectively.The 16S rRNA sequencing of isolates showed the highest homologous similarity(99.9%)with domestic endemic Mycoplasma strains XBY01,HBE-1,Ningxia-1,Tibet-10 and NM2012,and the homologous similarity with PG2 was 99.3%-99.7%.The similarity to Mycoplasma sheep pneumoniae Y98 was 81.5%-82.7%,and the homology of isolates reached more than 99.9%.By uvrC sequencing,the homology with Mycoplasma bovis was 99.3%-99.8%,and the homology with Mycoplasma agalactiae was less than 80.3%.Sequencing results showed that all isolates were Mycoplasma bovis.The isolates were resistant to norfloxacin,lomefloxacin,azithromycin,amoxicillin and erythromycin,and sensitive to kanamycin,gentamicin,doxycycline,tetracycline and ciprofloxacin.
作者
王峰
刘欢
白崇生
WANG Feng;LIU Huan;BAI Chongsheng(Yulin Animal Husbandry and Veterinary Service Center,Yulin 719000,China;Yulin Agricultural Publicity and Information Center,Yulin 719000,China)
出处
《中国草食动物科学》
CAS
2023年第5期61-65,共5页
China Herbivore Science
基金
陕西省榆林市科技计划项目(CXY-2020-077)。
关键词
榆林地区
牛支原体
分离鉴定
药敏试验
Yulin area
Mycoplasma bovis
isolation and identification
drug sensitivity experiment