Tracking SARS-CoV-2 Omicron Lineages Using Real-Time Reverse Transcriptase PCR Assays and Prospective Comparison with Genome Sequencing

Omicron has become the dominant SARS-CoV-2 variant globally since December 2021, with distinct waves being associated with separate Omicron sublineages. Rapid detection of BA.1, BA.2, BA.4, and BA.5 was accomplished in the province of Alberta, Canada, through the design and implementation of real-time reverse transcriptase PCR assays targeting S:N501Y, S:ins214EPE, S:H69/V70, ORF7b:L11F, and M:D3N. Using the combination of results for each of these markers, samples could be designated as belonging to sublineages within BA.1, BA.2, BA.4, or BA.5. The analytical sensitivity of these markers ranged from 132 to 2229 copies/mL and in-laboratory accuracy was 98.9-100%. A 97.3% agreement using 12,592 specimens was demonstrated for the assays compared to genome sequencing. The use of these assays, combined with genome sequencing, facilitated the surveillance of SARS-CoV-2 lineages throughout a BA.5-dominated period.