A Portable Reverse Transcription Recombinase Polymerase Amplification Assay for Rapid Detection of Foot-and-Mouth Disease Virus
A Portable Reverse Transcription Recombinase Polymerase Amplification Assay for Rapid Detection of Foot-and-Mouth Disease Virus
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DOI：
10.1371/currents.outbreaks.62df1c7c75ffc96cd59034531e2e8364
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发表时间：
2013-08-20
期刊：
影响因子：
3.7
通讯作者：
Weidmann, Manfred
中科院分区：
文献类型：
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作者：
Abd El Wahed, Ahmed;El-Deeb, Ayman;Weidmann, Manfred
口蹄疫（Foot-and-mouth disease，FMD）是一种跨国界的家畜病毒性传染病，给世界范围内的畜牧业造成巨大的经济损失和严重的传染性威胁。口蹄疫的早期诊断有助于通过适当的暴发管理减少其影响。在这项研究中，我们描述了一个实时逆转录重组酶聚合酶扩增（RT-RPA）检测口蹄疫病毒（FMDV）的发展。FMDV RT-RPA设计靶向FMDV的3D基因，并且使用260 nt分子RNA标准品进行测定验证。RT-RPA测定是快速的（4-10分钟），并且通过概率单位回归分析在检测到1436个RNA分子时确定分析灵敏度。FMDV RT-RPA检测法检测了所有7种FMDV血清型制备的RNA，但未检测到猪瘟病毒或猪水泡病病毒。在埃及最近的口蹄疫爆发期间，在现场使用了FMDV RT-RPA检测。在临床样本中，逆转录聚合酶链反应（RT-PCR）和RT-RPA的诊断灵敏度分别为100%和98%。总之，FMDV RT-RPA比实时RT-PCR更快，更容易在现场处理。因此，RT-RPA可以很容易地在检疫站或农场进行诊断，以快速检测感染点。
Foot-and-mouth disease (FMD) is a trans-boundary viral disease of livestock, which causes huge economic losses and constitutes a serious infectious threat for livestock farming worldwide. Early diagnosis of FMD helps to diminish its impact by adequate outbreak management. In this study, we describe the development of a real-time reverse transcription recombinase polymerase amplification (RT-RPA) assay for the detection of FMD virus (FMDV). The FMDV RT-RPA design targeted the 3D gene of FMDV and a 260 nt molecular RNA standard was used for assay validation. The RT-RPA assay was fast (4-10 minutes) and the analytical sensitivity was determined at 1436 RNA molecules detected by probit regression analysis. The FMDV RT-RPA assay detected RNA prepared from all seven FMDV serotypes but did not detect classical swine fever virus or swine vesicular disease virus. The FMDV RT-RPA assay was used in the field during the recent FMD outbreak in Egypt. In clinical samples, reverse transcription polymerase chain reaction (RT-PCR) and RT-RPA showed a diagnostic sensitivity of 100% and 98%, respectively. In conclusion, FMDV RT-RPA was quicker and much easier to handle in the field than real-time RT-PCR. Thus RT-RPA could be easily implemented to perform diagnostics at quarantine stations or farms for rapid spot-of-infection detection.