SARS-CoV-2 variants detection using TaqMan SARS-CoV-2 mutation panel molecular genotyping assays

Abstract

Purpose: For rapid detection and tracking of SARS-CoV-2, a simple screening method alternative to laborious and expensive sequencing is highly desirable. Here, we evaluated performance characteristics of TaqMan SARS-CoV-2 mutation panel genotyping molecular assay for detection of most common reported SARS-CoV-2 variants using specific RT-PCR assays targeting single nucleotide polymorphisms (SNP). Patients and Methods: A total of 150 SARS-CoV-2 positive samples from March to July were included for this study. In addition, five controls comprised of synthetic RNA B.1.1.7_601443, B.1.351_678597, P.1_792683, B.1.617.1_1662307 and MN908947.3-Wuhan-hu-1 from Twist bioscience and B.1.1.7 (England/204820464/2020) and B.1.351 (South Africa/KRISP-K005325/2020) from Zeptometrix, NY, USA were used for validation. Total RNA from specimens was extracted using Omega Bio-Tek Mag-Bind Viral RNA Xpress Extraction Kit and tested for known SARS-CoV2 variants using ThermoFisher TaqMan SARS-CoV-2 mutation panel molecular assay on the QuantStudio 12K Flex. Nine representative samples have been compared with sequencing. Data were analyzed by genotype calling using QuantStudio™ design and analysis software v 2.5 with the genotyping analysis module. Results: All validation controls were tested in triplicate and repeated in singlet on three different days and all reported variants were matched as expected. Out of 150 SARS-CoV-2 positive specimens, 69 (46%) were B.1.617.2, 49 (32.7%) were B.1.1.7, P.1 and P.2 were 4 (2.7%) each and B.1.351 and B.1.427/B.1429 were 2 (1.3%) each. Three (2%) were B.1.526, and 17 (11.3%) have a mutation in D614G. Genotyping results from the present study showing B.1.617.2, B.1.1.7, and B.1.526 variants and their mutation genes were concordant with sequencing results. Conclusion: Our study indicates that TaqMan SARS-CoV-2 mutation panel molecular genotyping assays detect and differentiate all published common variants B.1.617.2 (Delta), B.1.1.7 (Alpha), B.1.526 (Iota), B.1.351 (Beta), P.1 (Gamma), P.2 (Zeta), B.1.617.1 (Kappa) and B.1.427/B.1.429 (Epsilon) that can be used for surveillance and epidemic control and prevention.