The Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF, UK.
Viruses. 2019 Jul 31;11(8):699. doi: 10.3390/v11080699.
Peste des petits ruminants (PPR) is a disease of small ruminants caused by peste des petits ruminants virus (PPRV), and is endemic in Asia, the Middle East and Africa. Effective control combines the application of early warning systems, accurate laboratory diagnosis and reporting, animal movement restrictions, suitable vaccination and surveillance programs, and the coordination of all these measures by efficient veterinary services. Molecular assays, including conventional reverse transcription-polymerase chain reaction (RT-PCR) and real-time RT-PCR (RT-qPCR) have improved the sensitivity and rapidity of diagnosing PPR. However, currently these assays are only performed within laboratory settings; therefore, the development of field diagnostics for PPR would improve the fast implementation of control policies, particularly when PPR has been targeted to be eradicated by 2030. Loop-mediated isothermal amplification (LAMP) assays are simple to use, rapid, and have sensitivity and specificity within the range of RT-qPCR; and can be performed in the field using disposable consumables and portable equipment. This study describes the development of a novel RT-LAMP assay for the detection of PPRV nucleic acid by targeting the N-protein gene. The RT-LAMP assay was evaluated using cell culture propagated PPRVs, field samples from clinically infected animals and samples from experimentally infected animals encompassing all four lineages (I-IV) of PPRV. The test displayed 100% concordance with RT-qPCR when considering an RT-qPCR cut-off value of C >40. Further, the RT-LAMP assay was evaluated using experimental and outbreak samples without prior RNA extraction making it more time and cost-effective. This assay provides a solution for a pen-side, rapid and inexpensive PPR diagnostic for use in the field in nascent PPR eradication programme.
小反刍兽疫（PPR）是一种由小反刍兽疫病毒（PPRV）引起的小反刍动物疾病，在亚洲、中东和非洲流行。有效的控制措施结合了早期预警系统的应用、准确的实验室诊断和报告、动物流动限制、适当的疫苗接种和监测计划，以及通过高效的兽医服务协调所有这些措施。分子检测方法，包括常规逆转录聚合酶链反应（RT-PCR）和实时 RT-PCR（RT-qPCR），提高了 PPR 的诊断灵敏度和速度。然而，目前这些检测方法仅在实验室环境中进行；因此，开发用于 PPR 的现场诊断方法将改善控制政策的快速实施，特别是当 PPR 被定为 2030 年消除的目标时。环介导等温扩增（LAMP）检测方法简单易用、快速，其灵敏度和特异性与 RT-qPCR 相当；并且可以使用一次性耗材和便携式设备在现场进行。本研究描述了一种针对 N 蛋白基因的新型 RT-LAMP 检测方法，用于检测 PPRV 核酸。使用细胞培养繁殖的 PPRVs、来自临床感染动物的现场样本和涵盖 PPRV 所有四个谱系（I-IV）的实验感染动物样本对 RT-LAMP 检测方法进行了评估。当考虑 RT-qPCR 截断值 C>40 时，该检测方法与 RT-qPCR 具有 100%的一致性。此外，该 RT-LAMP 检测方法无需预先进行 RNA 提取即可用于实验和暴发样本的评估，使其更加省时和节省成本。该检测方法为新兴的 PPR 根除计划提供了一种在现场进行快速、廉价的 PPR 诊断方法。
