Laboratoire MIVEGEC, UMR 224 IRD/CNRS/UM1, Montpellier, France.
J Virol Methods. 2013 Oct;193(1):55-61. doi: 10.1016/j.jviromet.2013.04.016. Epub 2013 May 10.
Mosquitoes-borne viruses are a major threat for human populations. Among them, chikungunya virus (CHIKV) and dengue virus (DENV) cause thousands of cases worldwide. The recent propagation of mosquito vectors competent to transmit these viruses to temperate areas increases their potential impact on susceptible human populations. The development of sensitive methods allowing the detection and isolation of infectious viruses is of crucial interest for determination of virus contamination in humans and in competent mosquito vectors. However, simple and rapid method allowing the capture of infectious CHIKV and DENV from samples with low viral titers useful for further genetic and functional characterization of circulating strains is lacking. The present study reports a fast and sensitive isolation technique based on viral particles adsorption on magnetic beads coated with anionic polymer, poly(methyl vinyl ether-maleic anhydrate) and suitable for isolation of infectious CHIKV and DENV from the four serotypes. Starting from quite reduced biological material, this method was accurate to combine with conventional detection techniques, including qRT-PCR and immunoblotting and allowed isolation of infectious particles without resorting to a step of cultivation. The use of polymer-coated magnetic beads is therefore of high interest for rapid detection and isolation of CHIKV and DENV from samples with reduced viral loads and represents an accurate approach for the surveillance of mosquito vector in area at risk for arbovirus outbreaks.
蚊媒病毒是人类的主要威胁之一。其中,基孔肯雅病毒(CHIKV)和登革热病毒(DENV)在全球范围内导致数千例病例。蚊子传播媒介在温带地区的传播增加,增加了它们对易感人群的潜在影响。开发灵敏的方法来检测和分离有感染力的病毒,对于确定人类和有能力的蚊子媒介中的病毒污染至关重要。然而,简单快速的方法可以从低病毒滴度的样本中捕获有感染力的 CHIKV 和 DENV,这对于循环株的进一步遗传和功能特征分析非常有用,但目前尚缺乏这种方法。本研究报告了一种基于病毒颗粒吸附在带负电荷的聚合物(聚甲基乙烯基醚-马来酸酐)涂覆的磁性珠上的快速灵敏的分离技术,适用于从四个血清型中分离有感染力的 CHIKV 和 DENV。从相当少的生物材料开始,这种方法与常规检测技术(包括 qRT-PCR 和免疫印迹)结合使用非常准确,可以在不进行培养的情况下分离出有感染力的颗粒。因此,聚合物涂覆的磁性珠的使用对于从病毒载量降低的样本中快速检测和分离 CHIKV 和 DENV 非常有意义,并且是对有暴发 arbovirus 风险地区的蚊子媒介进行监测的准确方法。