Use of Whole Genome Sequencing Data for a First in Silico Specificity Evaluation of the RT-qPCR Assays Used for SARS-CoV-2 Detection

International Journal of Molecular Sciences

Philippe Herman¹  Arnaud Capron²  Kevin Vanneste³  Mathieu Gand³  NancyH. C. Roosens³  SigridC. J. De Keersmaecker³  Steven Van Gucht⁴  Isabelle Thomas⁴ 

[1] Expertise and Service Provision, Sciensano, J. Wytsmanstraat 14, B-1050 Brussels, Belgium;Quality of Laboratories, Sciensano, J. Wytsmanstraat 14, B-1050 Brussels, Belgium;Transversal activities in Applied Genomics, Sciensano, J. Wytsmanstraat 14, B-1050 Brussels, Belgium;Viral Diseases, Sciensano, J. Wytsmanstraat 14, B-1050 Brussels, Belgium;

关键词: SARS-CoV-2;    COVID-19;    detection;    diagnosis;    RT-qPCR;    in silico specificity evaluation;   

DOI  :  10.3390/ijms21155585

来源: DOAJ

【 摘 要 】

The current COronaVIrus Disease 2019 (COVID-19) pandemic started in December 2019. COVID-19 cases are confirmed by the detection of SARS-CoV-2 RNA in biological samples by RT-qPCR. However, limited numbers of SARS-CoV-2 genomes were available when the first RT-qPCR methods were developed in January 2020 for initial in silico specificity evaluation and to verify whether the targeted loci are highly conserved. Now that more whole genome data have become available, we used the bioinformatics tool SCREENED and a total of 4755 publicly available SARS-CoV-2 genomes, downloaded at two different time points, to evaluate the specificity of 12 RT-qPCR tests (consisting of a total of 30 primers and probe sets) used for SARS-CoV-2 detection and the impact of the virus’ genetic evolution on four of them. The exclusivity of these methods was also assessed using the human reference genome and 2624 closely related other respiratory viral genomes. The specificity of the assays was generally good and stable over time. An exception is the first method developed by the China Center for Disease Control and prevention. NIID: National Institute for Infectious Diseases (CDC), which exhibits three primer mismatches present in 358 SARS-CoV-2 genomes sequenced mainly in Europe from February 2020 onwards. The best results were obtained for the assay of Chan et al. (2020) targeting the gene coding for the spiking protein (S). This demonstrates that our user-friendly strategy can be used for a first in silico specificity evaluation of future RT-qPCR tests, as well as verifying that the former methods are still capable of detecting circulating SARS-CoV-2 variants.

【 授权许可】

Unknown