Development and evaluation of tailored specific real-time RT-PCR assays for detection of foot-and-mouth disease virus serotypes circulating in East Africa
Development and evaluation of tailored specific real-time RT-PCR assays for detection of foot-and-mouth disease virus serotypes circulating in East Africa
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DOI：
10.1016/j.jviromet.2016.08.002
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发表时间：
2016-11-01
影响因子：
3.1
通讯作者：
King, Donald P.
中科院分区：
文献类型：
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作者：
Bachanek-Bankowska, Katarzyna;Mero, Herieth R.;King, Donald P.
快速、可靠和准确的诊断方法为监测和控制口蹄疫的规划提供了必不可少的支持。虽然在口蹄疫流行国家和无口蹄疫国家已经建立并广泛使用了检测口蹄疫病毒（FMDV）的泛特异性分子检测方法，但目前的血清分型方法主要依赖抗原检测elisa或核苷酸测序方法。本报告描述了一组血清型特异性实时RT-PCR检测方法（rRT-PCR）的发展，该方法专门用于检测目前在东非流行的口蹄疫病毒谱系。这些检测的目标序列位于vp1编码区域内，具有较高的谱系内同一性，但与该区域传播的其他血清型口蹄疫病毒不发生交叉反应。这些血清型特异性检测与泛诊断测试具有相同的热剖面，因此可以并行运行它们以产生与检测3d编码区域的泛诊断测试相当的C-T值。这些检测方法与已建立的泛特异性分子测试一起进行了评估，该测试使用了从坦桑尼亚、肯尼亚和埃塞俄比亚收集的现场样本和病毒分离物，这些分离物以前已通过核苷酸测序进行了表征。这些样本（n= 71）分别代表血清型A（拓扑型AFRICA， G-I）、血清型O（拓扑型EA-2和EA-4）、血清型SAT 1（拓扑型I (NWZ)）和血清型SAT2（拓扑型IV）。此外，来自不含感染性病毒的样品的FMDV RNA仍然可以使用这些检测方法进行血清分型。这些血清型特异性实时RT-PCR检测可以检测目前在东非流行的口蹄疫病毒并确定其特征，从而改善该地区的疾病控制。(C) 2016年作者。Elsevier B.V.出版
Rapid, reliable and accurate diagnostic methods provide essential support to programmes that monitor and control foot-and-mouth disease (FMD). While pan-specific molecular tests for FMD virus (FMDV) detection are well established and widely used in endemic and FMD-free countries, current serotyping methods mainly rely either on antigen detection ELISAs or nucleotide sequencing approaches. This report describes the development of a panel of serotype-specific real-time RT-PCR assays (rRT-PCR) tailored to detect FMDV lineages currently circulating in East Africa. These assays target sequences within the VP1-coding region that share high intra-lineage identity, but do not cross-react with FMD viruses from other serotypes that circulate in the region. These serotype-specific assays operate with the same thermal profile as the pan-diagnostic tests making it possible to run them in parallel to produce C-T values comparable to the pan-diagnostic test detecting the 3D-coding region. These assays were evaluated alongside the established pan-specific molecular test using field samples and virus isolates collected from Tanzania, Kenya and Ethiopia that had been previously characterised by nucleotide sequencing. Samples (n= 71) representing serotype A (topotype AFRICA, lineage G-I), serotype O (topotypes EA-2 and EA-4), serotype SAT 1 (topotype I (NWZ)) and serotype SAT2 (topotype IV) were correctly identified with these rRT-PCR assays. Furthermore, FMDV RNA from samples that did not contain infectious virus could still be serotyped using these assays. These serotype-specific real-time RT-PCR assays can detect and characterise FMDVs currently circulating in East Africa and hence improve disease control in this region. (C) 2016 The Authors. Published by Elsevier B.V.