Unité des Arbovirus et virus de fièvres hémorragiques, Institut Pasteur Dakar, 36, Avenue Pasteur, BP 220 Dakar, Senegal.
Virol J. 2013 Oct 22;10:311. doi: 10.1186/1743-422X-10-311.
Zika virus (ZIKV), a mosquito borne flavivirus is a pathogen affecting humans in Asia and Africa. ZIKV infection diagnosis relies on serology-which is challenging due to cross-reactions with other flaviviruses and/or absence or low titer of IgM and IgG antibodies at early phase of infection- virus isolation, which is labor intensive, time consuming and requires appropriate containment. Therefore, real-time RT-PCR (rRT-PCR) is an appealing option as a rapid, sensitive and specific method for detection of ZIKV in the early stage of infection. So far, only one rRT-PCR assay has been described in the context of the outbreak in Micronesia in 2007. In this study, we described a one step rRT-PCR for ZIKV which can detect a wider genetic diversity of ZIKV isolates from Asia and Africa.
The NS5 protein coding regions of African ZIKV isolates were sequenced and aligned with representative flaviviruses sequences from GenBank to design primers and probe from conserved regions. The analytical sensitivity of the assay was evaluated to be 32 genome-equivalents and 0.05 plaque forming unit (pfu). The assay was shown to detect 37 ZIKV isolates covering a wide geographic in Africa and Asia over 36 years but none of the 31 other flaviviruses tested showing high analytical specificity. The rRT-PCR could be performed in less than 3 hours. This method was used successfully to detect ZIKV strains from field-caught mosquitoes.
We have developed a rapid, sensitive and specific rRT-PCR for detection of ZIKV. This assay is a useful tool for detection of ZIKV infection in regions where a number of other clinically indistinguishable arboviruses like dengue or chikungunya co-circulate. Further studies are needed to validate this assay in clinical positive samples collected during acute ZIKV infection.
寨卡病毒（ZIKV）是一种通过蚊子传播的黄病毒，是影响亚洲和非洲人类的病原体。ZIKV 感染的诊断依赖于血清学检测——由于与其他黄病毒的交叉反应以及感染早期 IgM 和 IgG 抗体的缺失或低滴度，这一检测方法具有挑战性——病毒分离，这一方法既费力又耗时，且需要适当的隔离。因此，实时 RT-PCR（rRT-PCR）是一种有吸引力的选择，作为一种快速、敏感和特异的方法，可用于在感染的早期阶段检测 ZIKV。到目前为止，只有一种 rRT-PCR 检测方法在 2007 年密克罗尼西亚爆发时被描述过。在这项研究中，我们描述了一种一步法 rRT-PCR，可检测来自亚洲和非洲的更广泛遗传多样性的 ZIKV 分离株。
对非洲 ZIKV 分离株的 NS5 蛋白编码区进行测序，并与 GenBank 中代表性黄病毒序列进行比对，以设计来自保守区的引物和探针。评估该检测方法的分析灵敏度为 32 个基因组当量和 0.05 噬菌斑形成单位（pfu）。该检测方法显示可检测来自非洲和亚洲的 37 株 ZIKV 分离株，覆盖了 36 年的广泛地理范围，但在 31 种其他测试的黄病毒中，没有一种表现出很高的分析特异性。该 rRT-PCR 可在不到 3 小时内完成。该方法成功地用于检测野外捕获的蚊子中的 ZIKV 株。
我们开发了一种快速、敏感和特异的 rRT-PCR 检测 ZIKV 的方法。该检测方法是一种有用的工具，可用于检测在与其他临床上难以区分的虫媒病毒（如登革热或基孔肯雅热）共同传播的地区的 ZIKV 感染。需要进一步的研究来验证该检测方法在急性 ZIKV 感染期间采集的临床阳性样本中的应用。
