Cortes Helder C E, Reis Yara, Gottstein Bruno, Hemphill Andrew, Leitão Alexandre, Müller Norbert
Laboratório de Parasitologia, Núcleo da Mitra, ICAM, Universidade de Evora, Apartado 94, 7000-554 Evora, Portugal.
Vet Parasitol. 2007 May 31;146(3-4):352-6. doi: 10.1016/j.vetpar.2007.03.003. Epub 2007 Apr 6.
Besnoitia besnoiti, an apicomplexan protozoan parasite, is the causative agent of bovine besnoitiosis. This infection may dramatically affect body condition, and, in males, lead to irreversible infertility. While identification of clinical cases and their histopathological confirmation is relatively simple to carry out, finding subclinical forms of infection is more difficult, thus a more sensitive test for the identification of the etiological agent may be an appropriate diagnostic tool. We have developed the ITS1 rDNA-sequence-based conventional and real-time PCR which are highly sensitive and specific for the detection of B. besnoiti infection in cattle. A recombinant internal positive control was introduced to assess possible sample-related inhibitory effects during the amplification reaction and, in order to prevent false-positive results, a pre-PCR treatment of potentially contaminating dU-containing PCR product with uracil-DNA-glycosylase (UDG) was followed.
贝氏贝诺孢子虫是一种顶复门原生动物寄生虫，是牛贝诺孢子虫病的病原体。这种感染可能会严重影响牛的身体状况，对于公牛而言，还会导致不可逆的不育。虽然临床病例的识别及其组织病理学确认相对容易进行，但发现亚临床感染形式则较为困难，因此，一种用于识别病原体的更灵敏检测方法可能是合适的诊断工具。我们开发了基于ITS1 rDNA序列的常规PCR和实时PCR，它们对检测牛的贝氏贝诺孢子虫感染具有高度的敏感性和特异性。引入了重组内部阳性对照以评估扩增反应过程中可能与样本相关的抑制作用，并且为了防止出现假阳性结果，随后对可能被污染的含dU PCR产物进行了尿嘧啶-DNA糖基化酶（UDG）的PCR前处理。
