SARS-CoV-2 Variant Surveillance Using Tandem Targeted RT-PCR-based Genotyping Assays and Whole Genome Sequencing

Genomic surveillance is critical for tracking SARS-CoV-2 Variants of Concern (VOC) and for rapid detection of emerging variants. Whole genome sequencing (WGS) is the predominant method for genomic surveillance; but it is a laborious process for large-scale testing. The aim of this study was to assess the performance of a PCR-based mutation panel for the discrimination of 5 known VOC; Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Delta (B.1.617.2) and Omicron (B.1.1.529). Genotyping analysis was performed on 128 SARS-CoV-2 positive samples collected at the Life Science Testing Center at Northeastern University from April-December 2021. RNA extraction was performed using MagMax™ Viral/Pathogen II Nucleic Acid Isolation Kit. SARS-CoV-2 detection was confirmed using the TaqPath™ COVID-19 Combo Kit. Variant determination was conducted using a panel of TaqMan™ SARS-CoV-2 single nucleotide polymorphism (SNP) assays. On November 25, 2021, the emerging VOC (Omicron) was reported by South Africa and the panel was quickly modified to detect Omicron by substituting P681H and K417N assays. Based on the SNP panel analysis, variant identification in 128 samples were as follows: Alpha (N=34), Beta (N=1), Gamma (N=7), Delta (N=41) and Omicron (N=21). The genotyping panel accurately assigned lineages to 104 samples, confirmed by Ion Torrent GeneStudio S5 WGS. VOC discrimination using RT-PCR genotyping is a rapid, versatile method for detecting known and emerging SARS-CoV-2 variants. The versatility of SNP panels allows monitoring of emerging strains by simple layout adaptations. RT-PCR genotyping assays can expedite variant identification, enable high-throughput variant surveillance, and support WGS prioritization for detection of new variants. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This study was funded by Northeastern University's Life Sciences Testing Center ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: All samples were deidentified for study purposes in accordance with IRB approval by the Northeastern 223 University Office of Human Subject Research Protection (HSRP). I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes All data produced in the present study are available upon reasonable request to the authors [https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0024768\_TaqManSARS-CoV-2\_MutationPanel_UG.pdf][1] [https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0019746\_MagMAX\_MVP\_II\_NA\_IsolationKit\_CE-IVD_IFU.pdf][2] [https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0019277\_Ion\_AmpliSeq\_SARS-CoV-2\_Research\_Panel\_GeneStudio_QR.pdf][3] [1]: https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0024768_TaqManSARS-CoV-2_MutationPanel_UG.pdf [2]: https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0019746_MagMAX_MVP_II_NA_IsolationKit_CE-IVD_IFU.pdf [3]: https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0019277_Ion_AmpliSeq_SARS-CoV-2_Research_Panel_GeneStudio_QR.pdf