牛支原体套式PCR检测方法的建立

Development of a nested PCR assay for detection of Mycoplasma Bovis

为建立牛支原体检测方法,本研究采用牛支原体的oppD/F基因的两对特异性引物,建立了牛支原体的套式PCR检测方法。该套式PCR可以从100ccu/mL(color change unit颜色变化单位)的牛支原体培养物中检出目的片段;同时还可以从病牛肺脏、病牛鼻拭子中扩增出目的片段,而且结果与病原分离结果一致。特异性试验结果表明,该方法与其它支原体无交叉反应,是一种特异性强、敏感性高的牛支原体检测方法。

Mycoplasma boris is a pathogen associated with pneumonia, arthritis, mastitis and keratoconjunctivitis in cattle. In this work, a nest PCR method to detect M. boris was established using two pairs of primers based on opp D/F genes. This method had a sensitivity of 100 ccu （color change unit） equivalents per milliliter, and was highly specific in detecting M. bovis. From other phylogenetically related organisms. The PCR procedure was validated by comparison with isolation of M. bovis from the M. boris culture, nose swabs and lungs of diseased calves. The results confirmed that the nested PCR was a fast, sensitive, specific and reliable method for detecting M. boris infection in cattle.