A comparative study of real-time RT-PCR-based SARS-CoV-2 detection methods and its application to human-derived and surface swabbed material

Tastanova, Aizhan; Stoffel, Corinne Isabelle; Dzung, Andreas; Cheng, Phil Fang; Bellini, Elisa; Johansen, PÃ¥l; Duda, Agathe; Nobbe, Stephan; Lienhard, Reto; Bosshard, Philipp Peter; Levesque, Mitchell Paul

Real-time reverse transcription polymerase chain reaction (RT-PCR) remains a gold standard in detection of various viral diseases. In the COVID-19 pandemic, multiple RT-PCR based tests were developed to screen for viral infection. As an emergency response to growing testing demand, we established a SARS-CoV-2 PCR diagnostics platform for which we compared different commercial and in-house RT-PCR protocols. Four commercial (CDC 2019-nCoV, Applied BiosystemsTM 2019-nCoV Assay Kit v1 TF-SinglePlex, 2019-nCoV Assay Kit v2 TF-MultiPlex, and EURORealTime SARS-CoV-2), one customized (Institute Pasteur), and one in-house RT-PCR protocols were evaluated with 92 SARS-CoV-2 positive and 92 SARS-CoV-2 negative samples. Furthermore, economical and practical characteristics of these protocols were compared. Additionally, a highly sensitive digital droplet PCR (ddPCR) method was developed and application of RT- and ddPCR methods on SARS-CoV-2 environmental samples was examined. Very low limits of detection (1 or 2 viral copies/μL), high sensitivities (93.6-97.8%) and specificities (98.7-100%) for the tested RT-PCR protocols were found. Furthermore, the feasibility of downscaling two of the commercial protocols, which could optimize testing capacity was demonstrated. Tested commercial and customized RT-PCR detection kits show very good and comparable sensitivity, and specificity, and the kits could be further optimized for use on SARS-CoV-2 viral samples derived from human and surface swabbed samples.