猪嗜血支原体PCR及荧光定量PCR检测方法的建立和比较

摘要

To compare PCR and TaqMan real time PCR (FQ-PCR) for the detection of Mycoplasma haemosuis infection in pigs, the PCR method and FQ-PCR were set up with the primers targeting the 16S rRNA gene of M.haemosuis.Plasmid containing 16S rRNA gene was used as template to optimize the conditions of conventional PCR and FQ-PCR reactions.The results showed that these 2 assays have good sensitivity and specificity.The sensitivity of FQ-PCR was 1,000 times higher than conventional PCR.Twenty clinical samples were detected by these 2 assays, positive rate was 12/20 by conventional PCR, while 15/20 by FQ-PCR.In conclusion, these 2 assays can effectively detect the clinical infection of M.haemosuis and may provide useful tools in establishment of animal infection model.%为了解猪嗜血支原体(Mycoplasma haemosuis)对猪群的感染情况并建立该病的检测方法,本研究根据GenBank登录的M.haemosuis 16S rRNA基因序列(FJ263944)设计合成PCR引物以及荧光定量PCR(FQ-PCR)引物和探针.以含16S rRNA基因的重组质粒为模板,通过对PCR反应条件的优化,建立检测M.haemosuis的PCR和FQ-PCR检测方法.结果表明,这两种检测方法均具有较好的敏感性和特异性,与常规方法相比,FQ-PCR方法的敏感性高1000倍;用这两种PCR方法检测20份临床样品,常规PCR方法的阳性率为60%(12/20),而FQ-PCR方法的阳性率为75%(15/20).这两种检测方法的建立为确定M.haemosuis在我国猪群的感染情况和建立该病动物模型提供有效的检测手段.