Emergency SARS-CoV-2 Variants of Concern: Novel Multiplex Real-Time RT-PCR Assay for Rapid Detection and Surveillance.
Emergency SARS-CoV-2 Variants of Concern: Novel Multiplex Real-Time RT-PCR Assay for Rapid Detection and Surveillance.
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DOI：
10.1128/spectrum.02513-21
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发表时间：
2022-02-23
影响因子：
3.7
通讯作者：
Shang HS
中科院分区：
文献类型：
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作者：
Chung HY;Jian MJ;Chang CK;Lin JC;Yeh KM;Chen CW;Hsieh SS;Hung KS;Tang SH;Perng CL;Chang FY;Wang CH;Shang HS
由严重急性呼吸道综合征冠状病毒2型（SARS-CoV-2）引起的2019冠状病毒病（COVID-19）已在全球范围内蔓延。已经报道了SARS-CoV-2的许多变体，其中一些变体增加了传播性和/或降低了对疫苗的敏感性。有一个迫切需要的变异表型流行病学监测循环谱系。全基因组测序是确定SARS-CoV-2变异体的黄金标准，这是发展中国家的一个主要瓶颈。简化方法可提高流行病学监测的可行性和效率。我们设计了一种新的多重实时荧光RT-PCR方法，用于检测SARS-CoV-2 S基因突变的变异株。建立了多重PCR分型方法，使用针对SARS-CoV-2变异体刺突蛋白受体结合域的特异性引物和探针（ΔHV 69/70、K417 T、K417 N、L452 R、E484 K、E484 Q、N501 Y、P681 H和P681 R）检测9种突变。计算机模拟分析显示该测定具有高度特异性。发现关注变体（VOC）分型结果对我们的预期目标具有高度特异性，与其他上呼吸道病毒没有观察到交叉反应性。使用全基因组测序和商业试剂盒进一步验证了基于PCR的分型方法，该试剂盒应用于250名来自台湾的COVID-19患者的临床样本。通过对这些样本的筛查，可以按时间间隔确定流行趋势，包括台湾第三波疫情中的B.1.617.2。这种PCR分型策略允许检测五种主要的关注变体，并且还提供了一种开源PCR测定法，该测定法可以迅速部署在世界各地的实验室中，以加强对B.1.1.7、B.1.351、P.1和B.1.617.2变体以及刺突蛋白上的四种Omicron突变的局部出现和传播的监测（ΔHV 69/70，K417 N，N501 Y，P681 H）。重要性COVID-19已在全球蔓延。关注的SARS-CoV-2变体（VOC）正在引领COVID-19大流行的下一波浪潮。此前有研究指出，这些VOCs可能增加了传染性，降低了疫苗敏感性，改变了治疗方案，增加了防疫政策的难度。了解SARS-CoV-2变种仍然是所有地方政府当局关注的问题，对于制定和实施有效的公共卫生措施至关重要。本研究建立了一种基于多重实时荧光RT-PCR的SARS-CoV-2变异株鉴定方法。使用该方法同时鉴定了5种SARS-CoV-2变体（Alpha、Beta、Gamma、Delta和Omicron）。PCR分型可以提供快速的检测结果，成本较低，可行性较高，这是任何诊断实验室都有能力做到的。表征这些变异体及其突变对于跟踪SAR-CoV-2的进化非常重要，并且有助于公共感染控制和政策制定策略。
Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread worldwide. Many variants of SARS-CoV-2 have been reported, some of which have increased transmissibility and/or reduced susceptibility to vaccines. There is an urgent need for variant phenotyping for epidemiological surveillance of circulating lineages. Whole-genome sequencing is the gold standard for identifying SARS-CoV-2 variants, which constitutes a major bottleneck in developing countries. Methodological simplification could increase epidemiological surveillance feasibility and efficiency. We designed a novel multiplex real-time reverse transcriptase PCR (RT-PCR) to detect SARS-CoV-2 variants with S gene mutations. This multiplex PCR typing method was established to detect 9 mutations with specific primers and probes (ΔHV 69/70, K417T, K417N, L452R, E484K, E484Q, N501Y, P681H, and P681R) against the receptor-binding domain of the spike protein of SARS-CoV-2 variants. In silico analyses showed high specificity of the assays. Variants of concern (VOC) typing results were found to be highly specific for our intended targets, with no cross-reactivity observed with other upper respiratory viruses. The PCR-based typing methods were further validated using whole-genome sequencing and a commercial kit that was applied to clinical samples of 250 COVID-19 patients from Taiwan. The screening of these samples allowed the identification of epidemic trends by time intervals, including B.1.617.2 in the third Taiwan wave outbreak. This PCR typing strategy allowed the detection of five major variants of concern and also provided an open-source PCR assay which could rapidly be deployed in laboratories around the world to enhance surveillance for the local emergence and spread of B.1.1.7, B.1.351, P.1, and B.1.617.2 variants and of four Omicron mutations on the spike protein (ΔHV 69/70, K417N, N501Y, P681H). IMPORTANCE COVID-19 has spread globally. SARS-CoV-2 variants of concern (VOCs) are leading the next waves of the COVID-19 pandemic. Previous studies have pointed out that these VOCs may have increased infectivity, have reduced vaccine susceptibility, change treatment regimens, and increase the difficulty of epidemic prevention policy. Understanding SARS-CoV-2 variants remains an issue of concern for all local government authorities and is critical for establishing and implementing effective public health measures. A novel SARS-CoV-2 variant identification method based on a multiplex real-time RT-PCR was developed in this study. Five SARS-CoV-2 variants (Alpha, Beta, Gamma, Delta, and Omicron) were identified simultaneously using this method. PCR typing can provide rapid testing results with lower cost and higher feasibility, which is well within the capacity for any diagnostic laboratory. Characterizing these variants and their mutations is important for tracking SAR-CoV-2 evolution and is conducive to public infection control and policy formulation strategies.
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影响因子：
3.9
作者：
Jian MJ;Chung HY;Chang CK;Lin JC;Yeh KM;Chen CW;Li SY;Hsieh SS;Liu MT;Yang JR;Tang SH;Perng CL;Chang FY;Shang HS
通讯作者：
Shang HS
SARS-CoV-2 在中国航空乘客中传播的风险。
影响因子：
11.8
作者：
Hu, Maogui;Wang, Jinfeng;Lai, Shengjie
通讯作者：
Lai, Shengjie
影响因子：
5.8
作者：
Ioannou, Petros;Karakonstantis, Stamatis;Kofteridis, Diamantis P.
通讯作者：
Kofteridis, Diamantis P.
影响因子：
11.8
作者：
Gharpure R;Sami S;Vostok J;Johnson H;Hall N;Foreman A;Sabo RT;Schubert PL;Shephard H;Brown VR;Brumfield B;Ricaldi JN;Conley AB;Zielinski L;Malec L;Newman AP;Chang M;Finn LE;Stainken C;Mangla AT;Eteme P;Wieck M;Green A;Edmundson A;Reichbind D;Brown V Jr;Quiñones L;Longenberger A;Hess E;Gumke M;Manion A;Thomas H;Barrios CA;Koczwara A;Williams TW;Pearlowitz M;Assoumou M;Senisse Pajares AF;Dishman H;Schardin C;Wang X;Stephens K;Moss NS;Singh G;Feaster C;Webb LM;Krueger A;Dickerson K;Dewart C;Barbeau B;Salmanson A;Madoff LC;Villanueva JM;Brown CM;Laney AS
通讯作者：
Laney AS
影响因子：
64.8
作者：
通讯作者：
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