猪弓形虫病特异PCR诊断方法的建立

Development of PCR assay for diagnosing swine toxoplasmosis

以核糖体DNA第一内转录间隔区(ITS1)序列作为弓形虫种特异的遗传标记,建立了猪弓形虫病特异PCR诊断方法。应用人工感染的方法建立了猪弓形虫病动物模型,摸索出猪弓形虫病PCR检测的最佳条件。敏感性和特异性试验结果显示,该PCR方法最低能检测到10 个弓形虫速殖子的DNA,且与相关的8种原虫和线虫均无交叉反应,证明该PCR方法具有高度的敏感性和特异性。动物感染试验结果证明,在试验猪出现明显临床症状时(感染后第5 d),对取材的8 个组织样品用该PCR方法检测,有6个组织样品为阳性,其中检出率最高的组织为肺门淋巴结。

The first internal transcribed spacer (ITS1) of nuclear ribosomal DNA was amplified from the genomic DNA of Toxoplasma gondii by PCR using the species-specific primers. The assay was highly sensitive and was able to detect 10 tachyzoites of T. gondii. On day 5 post-infection, the specific PCR products of T. gondii could be amplified from six of the eight examined tissues of pig after experimental infection, and the bronchopulmonary lymph node could be selected as tissue for the PCR assay.