Clinical comparison, standardization and optimization of Zika virus molecular detection
Clinical comparison, standardization and optimization of Zika virus molecular detection
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寨卡病毒分子检测的临床比较、标准化和优化
DOI：
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发表时间：
2016
期刊：
影响因子：
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通讯作者：
J. Drexler
中科院分区：
文献类型：
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作者：
V. Corman;D. Cadar;S. Pas;J. Schinkel;R. Molenkamp;A. Eis;M. Koopmans;M. Grobusch;C. Drosten;J. Drexler
目的分子寨卡病毒(ZIKV)检测是当前寨卡病毒暴发疫情中患者诊断的关键。在这里，我们讨论了广泛使用的实时RT-PCR(QPCR)检测ZIKV的标准化和诊断性能。方法对目前已知ZIKV基因变异的两种新的qPCR方法和已发表的全部6种qPCR方法进行分析。使用新构建的通用对照RNA(UcRNA)对所有检测方法的性能进行比较，该通用对照RNA包含一条合成RNA链上所有比较检测的靶区。发现在已公布的qPCR方案中，存在多达10个与ZIKV暴发毒株的寡核苷酸错配。大多数检测方法的分析灵敏度约为每次反应5个拷贝，而三种检测方法的灵敏度降低了3-250倍。新的ucRNA实现了统一的ZIKV定量，而PCR阈值周期(CT值)的比较导致了不同方案之间高达20倍的错误定量。33份寨卡病毒暴发样本在发病后2周内的平均病毒载量分别为血10 4RNA拷贝数(10 2~4×10 5)和尿液5×10 3拷贝数(4×10 2~5 9×10 4)。结论几种ZIKV定量聚合酶链式反应方法对ZIKV暴发毒株的敏感性有限且不相容。ZIKV感染导致病毒浓度低，无论样本类型如何，都接近技术检测限值，这意味着可能有20%-80%的患者由于分子测试的有限敏感性而无法确诊。我们提供了最新的ZIKV检测方法，适用于所有ZIKV毒株。UcRNA将使跨地区和多中心临床内协调实施寨卡病毒分子诊断
Objective Molecular Zika virus (ZIKV) detection is key to patient diagnostics during the current outbreak. Here, we address standardization and diagnostic performance of widely used realtime RT-PCR (qPCR) protocols for ZIKV detection. Methods Two novel qPCR protocols covering the currently known ZIKV genetic variability were analyzed together with all six published qPCR protocols. The performance of all assays was compared using a newly constructed universal control RNA (ucRNA) that contains the target regions of all compared assays on one strand of synthetic RNA. Findings Up to 10 oligonucleotide mismatches with ZIKV outbreak strains existed in published qPCR protocols. The analytical sensitivity of most assays was around 5 copies per reaction, whereas three assays showed a 3-250-fold decreased sensitivity. The novel ucRNA enabled uniform ZIKV quantification, whereas comparisons of PCR threshold cycles (CT-values) resulted in up to 20-fold misquantification between protocols. Mean ZIKV loads in 33 outbreak samples were 10 4 RNA copies/mL of blood (range; 10 2 -4x10 5 ) and 5x10 3 RNA copies/ mL of urine (range; 4x10 2 -5.9x10 4 ) within two weeks after symptom onset. Conclusion Several ZIKV qPCR protocols show limited sensitivity and incompatibility with ZIKV outbreak strains. ZIKV infection results in low virus concentrations close to the technical limit of detection irrespective of sample type, implying that 20%-80% of patients may go undiagnosed due to limited sensitivity of molecular tests. We provide updated protocols for ZIKV detection that are suitable for all ZIKV strains. The ucRNA will enable coordinated implementation of ZIKV molecular diagnostics across regions and within multicentric clinical
影响因子：
6.4
作者：
Vaughn, DW;Green, S;Nisalak, A
通讯作者：
Nisalak, A