OmniSARS2: a highly sensitive and specific RT-qPCR-based COVID-19 diagnostic method designed to withstand SARS-CoV-2 lineage evolution

• Autor(a) principal: Carvalho-Correia, Eduarda
• Data de Publicação: 2021
• Outros Autores: Calçada, Carla Sofia Martins, Branca, Fernando, Estévez-Gómez, Nuria, De Chiara, Loretta, Varela, Nair, Gallego-García, Pilar, Posada, David, Sousa, Hugo, Sousa, João Carlos, Veiga, Maria Isabel, Osório, Nuno S.
• Tipo de documento: Artigo
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/1822/75604
• topic: SARS-CoV-2; COVID-19; RT-qPCR; B.1.1.7; Science & Technology

Extensive transmission of SARS-CoV-2 during the COVID-19 pandemic allowed the generation of thousands of mutations within its genome. While several of these become rare, others largely increase in prevalence, potentially jeopardizing the sensitivity of PCR-based diagnostics. Taking advantage of SARS-CoV-2 genomic knowledge, we designed a one-step probe-based multiplex RT-qPCR (OmniSARS2) to simultaneously detect short fragments of the SARS-CoV-2 genome in ORF1ab, E gene and S gene. Comparative genomics of the most common SARS-CoV-2 lineages, other human betacoronavirus and alphacoronavirus, was the basis for this design, targeting both highly conserved regions across SARS-CoV-2 lineages and variable or absent in other <i>Coronaviridae</i> viruses. The highest analytical sensitivity of this method for SARS-CoV-2 detection was 94.2 copies/mL at 95% detection probability (~1 copy per total reaction volume) for the S gene assay, matching the most sensitive available methods. In vitro specificity tests, performed using reference strains, showed no cross-reactivity with other human coronavirus or common pathogens. The method was compared with commercially available methods and detected the virus in clinical samples encompassing different SARS-CoV-2 lineages, including B.1, B.1.1, B.1.177 or B.1.1.7 and rarer lineages. OmniSARS2 revealed a sensitive and specific viral detection method that is less likely to be affected by lineage evolution oligonucleotide–sample mismatch, of relevance to ensure the accuracy of COVID-19 molecular diagnostic methods.