Establishment of a novel diagnostic test for Bovine leukaemia virus infection using direct filter PCR
Establishment of a novel diagnostic test for Bovine leukaemia virus infection using direct filter PCR
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使用直接过滤 PCR 建立牛白血病病毒感染的新型诊断测试
DOI：
10.1111/tbed.13506
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发表时间：
2020
影响因子：
4.3
通讯作者：
Sekiguchi Satoshi
中科院分区：
文献类型：
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作者：
El Daous Hala;Mitoma Shuya;Elhanafy Eslam;Thi Nguyen Huyen;Thi Mai Ngan;Hara Akihiro;Duangtathip Karn;Takezaki Yuka;Kaneko Chiho;Norimine Junzo;Sekiguchi Satoshi
地方性牛白血病（EBL）是由牛白血病病毒（BLV）引起的牛的肿瘤性疾病。 EBL造成巨大的经济损失，因此快速可靠的诊断方法对于了解BLV的状况至关重要。这将使我们能够有效控制 BLV 感染并减轻经济损失。在这项研究中，我们使用干燥的血斑滤纸建立了 BLV 的直接诊断测试，无需对样品进行预处理。该研究基于从日本九州的一个农场采集的 159 份 EDTA 临床血液样本。血点滤纸用作直接过滤 PCR 的模板。当使用 ELISA 作为诊断金标准时，直接过滤 PCR 的灵敏度和特异性分别为 90.1% 和 97.5%。直接过滤 PCR 和实时 PCR 方法的 kappa 值为 0.97。点在滤纸上的干燥血液样本在室温下可稳定至少 10 天，即使样本来自 BLV 前病毒载量较低的牛。直接过滤 PCR 是一种快速、简单、可靠且经济高效的诊断测试，可直接检测临床血液样本中的 BLV 原病毒基因组，无需提取 DNA。而且简化了临床血液标本的采集、运输和储存程序。
Enzootic bovine leucosis (EBL) is a neoplastic disease of cattle caused byBovine leukaemia virus(BLV). EBL causes great economic losses, so a fast and reliable diagnostic method is critical for understanding the status of BLV. This will allow us to control BLV infections efficiently and mitigate economic losses. In this study, we established a direct diagnostic test for BLV using dried blood‐spotted filter papers without sample pre‐treatment. The study was based on 159 clinical blood specimens collected in EDTA from one farm in Kyushu, Japan. The blood‐spotted filter papers were used as the template for direct filter PCR. When an ELISA was used as the diagnostic gold standard, the sensitivity and specificity of the direct filter PCR were 90.1% and 97.5%, respectively. The kappa value for the direct filter PCR and real‐time PCR methods was 0.97. The dried blood samples spotted onto filter papers were stable for at least 10 days at room temperature, even when the samples were from cattle with a low BLV proviral load. Direct filter PCR is a rapid, easy, reliable and cost‐effective diagnostic test that directly detects the BLV proviral genome in clinical blood specimens without DNA extraction. Moreover, it simplifies the collection, transportation and storage procedures for clinical blood specimens.