A real-time reverse transcription loop-mediated isothermal amplification assay for the rapid detection of yellow fever virus
A real-time reverse transcription loop-mediated isothermal amplification assay for the rapid detection of yellow fever virus
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DOI：
10.1016/j.jviromet.2013.05.004
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发表时间：
2013-10-01
影响因子：
3.1
通讯作者：
Mwau, Matilu
中科院分区：
文献类型：
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作者：
Kwallah, Allan Ole;Inoue, Shingo;Mwau, Matilu
黄热病是一种蚊媒疾病，是非洲和南美洲的一种重要病毒性出血热，在那里流行。由于缺乏高度特异性的检测方法，非洲黄热病病毒（YFV）的检测仍然是一个挑战。本研究旨在建立并优化一种快速检测YFV的逆转录环介导等温扩增（RT-LAMP）方法。RT-LAMP在62 ° C下等温进行，使用允许在1小时内检测的实时浊度计。RT-LAMP的特异性使用来自黄病毒和其他相关病毒的RNA确定，其中仅检测到YFV RNA：西尼罗河病毒、登革热病毒、日本脑炎病毒、裂谷热病毒和基孔肯雅病毒。此外，当使用YFV加标的人血清样品比较RT-LAMP和实时RT-PCR时，也观察到相同的灵敏度，检测限为0.29 PFU/ml。在本研究中，两种肯尼亚YFV野生株的检出限与疫苗株17 D相同。与RT-PCR相比，RT-LAMP将反应时间从3 h缩短到1 h，灵敏度提高了10倍。因此，当肯尼亚和其他非洲国家爆发急性出血热时，这种检测方法为黄热病提供了一种简单、快速和可靠的诊断工具。(c)2013爱思唯尔有限公司版权所有。
Yellow fever, a mosquito-borne disease, is an important viral hemorrhagic fever in Africa and South America where it is endemic. Detection of yellow fever virus (YFV) in Africa remains a challenge due to a lack of highly specific tests. The aim of this study was to develop and optimize a rapid detection reverse transcription loop-mediated isothermal amplification (RT-LAMP) for YFV. The RT-LAMP was done isothermally at 62 degrees C using a real-time turbidimeter that allowed detection within 1 h. Specificity of the RT-LAMP was determined using RNA from flaviviruses and other related viruses where only YFV RNA was detected: West Nile virus, dengue viruses, Japanese encephalitis virus, Rift Valley fever virus, and chikungunya virus. In addition, equal sensitivity was also observed when the RT-LAMP and the real-time RT-PCR were compared using YFV-spiked human serum samples with a detection limit of 0.29 PFU/ml. Two Kenyan YFV wild strains showed an equal detection limit as the vaccine strain 17D in this study. The RT-LAMP reduced the time of reaction from 3 h to 1 h and increased sensitivity tenfold compared to RT-PCR. Therefore, this test offers a simple, rapid and reliable diagnostic tool for yellow fever when there are outbreaks of acute hemorrhagic fever in Kenya and other African countries. (c) 2013 Elsevier B.V. All rights reserved.