Development of a Real-Time PCR Assay for Detection and Quantification of Anaplasma ovis Infection
Development of a Real-Time PCR Assay for Detection and Quantification of Anaplasma ovis Infection
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DOI：
10.1111/tbed.12139
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发表时间：
2013-11-01
影响因子：
4.3
通讯作者：
Yin, H.
中科院分区：
文献类型：
--
作者：
Chi, Q.;Liu, Z.;Yin, H.
绵羊无形体是一种蜱传小反刍动物红细胞内立克次体病原体。实时PCR具有快速、准确、可靠、自动化和易于标准化的优点，但据我们所知，尚未用于绵羊曲霉的检测。在本研究中，开发了一种实时 PCR 检测方法来检测和定量羊曲霉。基于gltA基因设计物种特异性引物和TaqMan探针。与边缘无形体、牛无形体、嗜吞噬细胞无形体、伯氏疏螺旋体未观察到交叉反应。 l.、鹦鹉热衣原体、丝状支原体、鲁文舒尼泰勒虫和巴贝虫属。新疆隔离。分析灵敏度结果表明，实时 PCR 可以检测到少至 10 个拷贝的 gltA 基因。通过检测 254 份山羊血液样本并与传统 PCR 的结果进行比较来评估实时 PCR 的性能。这表明实时 PCR 检测比传统 PCR 检测更加灵敏。我们的结果表明，实时 PCR 是检测绵羊绵羊感染的有效方法，并且有潜力作为绵羊无形体病生态和流行病学监测的替代工具。
Anaplasma ovis is a tick-borne intra-erythrocytic rickettsial pathogen of small ruminants. Real-time PCR possesses merits of rapidity, accuracy, reliability, automation and ease of standardization, but has not been used for detection of A.ovis, to the best of our knowledge. In this study, a real-time PCR assay was developed for detection and quantification of A.ovis. Species-specific primers and TaqMan probe were designed based on the gltA gene. No cross-reactions were observed with Anaplasma marginale, Anaplasma bovis, Anaplasma phagocytophilum, Borrelia burgdorferi s. l., Chlamydia psittaci, Mycoplasma mycoides, Theileria luwenshuni and Babesia sp. Xinjiang isolate. Analytic sensitivity results revealed that real-time PCR could detect as few as 10 copies of the gltA gene. The performance of real-time PCR was assessed by testing 254 blood samples from goats and comparing with the results from conventional PCR. This demonstrated that the real-time PCR assay was significantly more sensitive than conventional PCR. Our results indicated that real-time PCR is a useful approach for detecting A.ovis infections and has potential as an alternative tool for ecological and epidemiological surveillance of ovine anaplasmosis.