Foreign Animal Disease Diagnostic Laboratory, Animal Plant Health Inspection Service, Foreign Animal Disease Research Unit, Agricultural Research Service, US Department of Agriculture, Plum Island Animal Disease Center, Orient Pt, New York 11957, USA.
J Clin Microbiol. 2010 Nov;48(11):4094-101. doi: 10.1128/JCM.01081-10. Epub 2010 Sep 15.
A highly sensitive detection test for Rinderpest virus (RPV), based on a real-time reverse transcription-PCR (rRT-PCR)system, was developed. Five different RPV genomic targets were examined, and one was selected and optimized to detect viral RNA in infected tissue culture fluid with a level of detection ranging from 0.59 to 87.5 50% tissue culture infectious doses (TCID(50)) per reaction depending on the viral isolate. The strain sensitivity of the test was validated on 16 RPV strains belonging to all three phylogenetic branches described for RPV. No cross-reactivity was detected with closely related peste des petit ruminants or with symptomatically similar viruses, including all seven serotypes of foot-and-mouth disease virus, two serotypes of vesicular stomatitis virus, bluetongue virus, and bovine herpes virus type 2. In samples from experimentally infected cattle, our real-time RT-PCR test was significantly more sensitive than the gold standard test of virus isolation, allowing the detection of the disease 2 to 4 days prior to the appearance of clinical signs. The comparison of clinical samples with putative diagnostic value from live animals showed that conjunctival swabs and blood buffy coat were the samples of choice for epidemiological surveillance, while lymph nodes performed the best as postmortem specimens. This portable and rapid real-time RT-PCR has the capability of the preclinical detection of RPV and provides differential diagnosis from look-alike diseases of cattle. As RPV is declared globally eradicated, this test provides an important rapid virus detection tool that does not require the use of infectious virus and allows the processing of a large number of samples.
一种基于实时逆转录聚合酶链反应（rRT-PCR）系统的高敏感检测牛瘟病毒（RPV）的方法被开发出来。该方法检测了 5 个不同的 RPV 基因组靶标，然后选择并优化了一个靶标，用于检测感染的组织培养液中的病毒 RNA，检测下限为 0.59 至 87.5 50%组织培养半数感染剂量（TCID(50)）/反应，具体取决于病毒分离株。该检测方法的菌株敏感性在属于 RPV 描述的三个系统发育分支的 16 个 RPV 菌株上得到了验证。该检测方法与密切相关的绵羊肺腺瘤病或症状相似的病毒（包括口蹄疫病毒的所有 7 个血清型、2 种水疱性口炎病毒、蓝舌病病毒和牛疱疹病毒 2 型）均无交叉反应。在来自实验感染牛的样品中，我们的实时 RT-PCR 检测方法比病毒分离的金标准检测方法灵敏得多，允许在出现临床症状前 2 至 4 天检测到疾病。对具有潜在诊断价值的来自活畜的临床样本进行比较显示，结膜拭子和血白细胞层是用于流行病学监测的首选样本，而淋巴结作为尸检样本的效果最佳。这种便携式快速实时 RT-PCR 具有 RPV 的临床前检测能力，并提供了与牛的类似疾病的鉴别诊断。由于全球宣布消灭了 RPV，因此该检测方法提供了一种重要的快速病毒检测工具，不需要使用传染性病毒，并且允许处理大量的样品。
