Division of Immunobiology, Department of Medicine, Robert Larner, M.D. College of Medicine, University of Vermont, Burlington, Vermont, United States of America.
Virology Division, Department of Laboratory Medicine and Pathology, University of Washington, Seattle, Washington, United States of America.
PLoS Biol. 2020 Oct 2;18(10):e3000896. doi: 10.1371/journal.pbio.3000896. eCollection 2020 Oct.
The ongoing COVID-19 pandemic has created an unprecedented need for rapid diagnostic testing. The World Health Organization (WHO) recommends a standard assay that includes an RNA extraction step from a nasopharyngeal (NP) swab followed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to detect the purified SARS-CoV-2 RNA. The current global shortage of RNA extraction kits has caused a severe bottleneck to COVID-19 testing. The goal of this study was to determine whether SARS-CoV-2 RNA could be detected from NP samples via a direct RT-qPCR assay that omits the RNA extraction step altogether. The direct RT-qPCR approach correctly identified 92% of a reference set of blinded NP samples (n = 155) demonstrated to be positive for SARS-CoV-2 RNA by traditional clinical diagnostic RT-qPCR that included an RNA extraction. Importantly, the direct method had sufficient sensitivity to reliably detect those patients with viral loads that correlate with the presence of infectious virus. Thus, this strategy has the potential to ease supply choke points to substantially expand COVID-19 testing and screening capacity and should be applicable throughout the world.
持续的 COVID-19 大流行对快速诊断检测提出了前所未有的需求。世界卫生组织（WHO）推荐了一种标准检测方法，包括从鼻咽（NP）拭子中提取 RNA，然后进行逆转录定量聚合酶链反应（RT-qPCR），以检测纯化的 SARS-CoV-2 RNA。目前全球 RNA 提取试剂盒短缺，导致 COVID-19 检测严重受阻。本研究的目的是确定是否可以通过直接 RT-qPCR 检测方法从 NP 样本中检测到 SARS-CoV-2 RNA，该方法完全省略了 RNA 提取步骤。直接 RT-qPCR 方法正确识别了 92%的参考集（n = 155）的 NP 样本，这些样本通过包括 RNA 提取的传统临床诊断 RT-qPCR 被证明为 SARS-CoV-2 RNA 阳性。重要的是，直接方法具有足够的灵敏度，可以可靠地检测出那些病毒载量与传染性病毒存在相关的患者。因此，这种策略有可能缓解供应瓶颈，从而大幅扩大 COVID-19 检测和筛查能力，并且应该适用于世界各地。
