奶牛附红细胞体和伊氏锥虫二重PCR诊断方法的建立

The Establishment of a Double PCR Assay for Diagnosis of E.wenyoni and T.evansi Infection

为建立奶牛附红细胞体和伊氏锥虫两种病原诊断方法并探索二者之间在奶牛感染中的关系,本研究针对两病原分别设计两对特异性引物,建立了奶牛附红细胞体和伊氏锥虫感染的二重PCR诊断方法,其扩增片段大小分别为415bp和237bp。敏感性试验和特异性试验表明,附红细胞体和伊氏锥虫的DNA的最低检测量为0.154pg和0.105pg;与猪肺炎支原体、大肠杆菌、葡萄球菌、鸡艾美耳球虫、牛双芽巴贝斯虫无交叉反应。35份临床血样检测结果为:奶牛附红细胞体阳性率22.89%,伊氏锥虫阳性率8.89%,其中共感染率为2.89%。临床试验表明,该方法可用于奶牛附红细胞体和伊氏锥虫的诊断,特别适用于早期诊断。

In order to study the relationship between E.wenyoni and T.evansi infection in bovine a double PCR assay was established for detecting E.wenyoni and T.evansi infection. 415bp and 237 bp segment was amplified ,and their lowest DNA quantity that could be detected was 0.154pg and 0.105pg. The specific test showed that there were no cross reactions with Mycoplasma hyopneumoniae,E.coli,Staphylococcus, Eimeria gallus, Babesia bigemina et al. Thirty-five clinical samples were detected. The results showed that the positive rate of the E.wenyoni and T.evansi was 22.89% and 8.89% respectively, and 2.89% infected the both diseases. The clinic sample detections have proved that the assay could be used in diagnoses of E.wenyoni and T.evansi infection, especially in early diagnosis.