绵羊肺炎支原体和精氨酸支原体双重PCR检测方法的建立及应用  

Development and Application of a Duplex PCR Assay for Detecting Mycoplasma ovipneumiae and Mycoplasma arginini

作　　者：郑佳琪[1] 黄海碧[1] 王晓晖[1] 李真亚 邢蒙恩 郝永清[1] 

机构地区：[1]内蒙古农业大学兽医学院,微生物学与免疫学实验室,呼和浩特010018

出　　处：《中国畜牧兽医》2015年第9期2240-2245,共6页China Animal Husbandry & Veterinary Medicine

基　　金：国家科技支撑计划(2012BAD13B00)“重点牧区“生产生态生活”保障技术集成与示范”;国家科技支撑计划(2011BAD18B01)“草原肉牛肉羊绿色养殖关键技术集成研究与产业化”

摘　　要：为建立绵羊肺炎支原体和精氨酸支原体的双重PCR检测方法,本试验分别设计了绵羊肺炎支原体和精氨酸支原体的特异性引物,优化反应条件后对其特异性和敏感性进行评价,并对40份鼻拭子进行了检测。结果显示,该方法能同时扩增出绵羊肺炎支原体545bp和精氨酸支原体806bp的特异性片段,而对其他病原的DNA扩增均为阴性。该双重PCR方法对绵羊肺炎支原体和精氨酸支原体的最低检测限分别为100和10pg/μL。40份鼻拭子检测结果显示,双重PCR检测方法与分离培养法符合率高达92.5%,均能鉴定出绵羊肺炎支原体和精氨酸支原体。结果表明,本研究建立的双重PCR方法可用于绵羊肺炎支原体和精氨酸支原体的临床快速诊断。In order to establish a duplex PCR method for simultaneous detection of Mycoplasma ovipneumiae and Mycoplasma arginini,specific primers of Mycoplasma ovipneumiae and Mycoplasma arginini were designed,and evaluated its sensitivity and specificity after optimizing the reaction conditions of PCR.Then,a total of 40 nasal swabs were tested by duplex PCR.The assay could specifically amplify PCR fragments of 545 and 806bp from Mycoplasma ovipneumiae and Mycoplasma arginini,respectively.While no PCR products were detected for other pathogens.The detection limits of the assay were determined to be 100pg/μL for Mycoplasma ovipneumiae and 10pg/μL for Mycoplasma arginini.The duplex PCR could detect Mycoplasma ovipneumiae and Mycoplasma arginini,and the coincidence rate could reach as high as 92.5% with enrichment culture about the 40 nasal swabs.The results suggested that the duplex PCR could be useful for clinical detection of Mycoplasma ovipneumiae and Mycoplasma arginini.

关 键 词：绵羊肺炎支原体 精氨酸支原体 双重PCR 

分 类 号：S858.26[农业科学—临床兽医学]