严重急性呼吸综合征冠状病毒2型(SARS-CoV-2)
计算生物学
2019年冠状病毒病(COVID-19)
2019-20冠状病毒爆发
生物
计算机科学
地理
遗传学
病毒学
医学
疾病
病理
爆发
传染病(医学专业)
作者
Ting Yan,Xu Ye,Rongrong Zheng,Xiaohong Zeng,Zehui Chen,Su Lin,Zihan Xia,Yiqun Liao,Yongyou Zhang,Qingge Li
标识
DOI:10.1128/spectrum.03222-22
摘要
Rapid identification and continuous surveillance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are critical for guiding the response to the COVID-19 pandemic. Whole-genome sequencing (WGS) is a preferred tool for this aim, but many laboratories suffer from a lack of resources to support population-level sequencing. Here, we describe two PCR strategies targeting spike protein mutations to identify the Alpha, Delta, and Omicron variants. Signature mutations were selected using a dedicated bioinformatic program. The selected mutations in Alpha and Delta variants were detected using multicolor melting curve analysis (MMCA). Thirty-two mutations of the Omicron variant were targeted using the MeltArray approach in one reaction, which was able to detect the Omicron subvariants BA.1, BA.2, BA.3, and BA.4/5. The limits of detection varied from five to 50 copies of RNA templates/reactions. No cross-reactivity was observed with nine other respiratory viruses, including other coronaviruses. We validated the MMCA and MeltArray assays using 309 SARS-CoV-2 positive samples collected at different time points. These assays exhibited 98.3% to 100% sensitivity and 100% specificity compared with WGS. Multiplexed real-time PCR strategies represent an alternative tool capable of identifying current SARS-CoV-2 VOCs, adaptable for emerging variants and accessible for laboratories using existing equipment and personnel. IMPORTANCE Rapid detection and mutation surveillance of SARS-CoV-2 VOCs is crucial for COVID-19 control, management, and prevention. We developed two rapid molecular assays based on the real-time PCR platform to identify important variants of concern, including the Omicron variant with a large number of mutations. Signature mutations were selected by an R program. Then, MMCA assays were established for Alpha and Delta variants, and a MeltArray assay targeting 32 mutations was developed for Omicron variant. These multiplexed PCR assays could be performed in a 96-well real-time PCR instrument within 2.5 h, offering a high-throughput choice for dynamic monitoring of SARS-CoV-2 VOCs in a standard microbiology laboratory.
科研通智能强力驱动
Strongly Powered by AbleSci AI