Development of a simple genotyping method based on indel mutations to rapidly screen SARS-CoV-2 circulating variants: Delta, Omicron BA.1 and BA.2

Highlights

• Rapid monitoring of SARS-CoV-2 variants of concern (VOCs) is an ongoing challenge.

• Indel mutations in the spike gene are interesting targets for VOCs screening assays.

• Two qPCR duplex assays perfectly discriminated VOCs Delta, Omicron BA.1 and BA.2.

• The developed genotyping method is rapid and cost-effective for VOCs screening.

Abstract

The high need of rapid and flexible tools that facilitate the identification of circulating SARS-CoV-2 Variants of Concern (VOCs) remains crucial for public health system monitoring.

Here, we develop allele-specific (AS)-qPCR assays targeting three recurrent indel mutations, ΔEF156–157, Ins214EPE and ΔLPP24–26, in spike (S) gene to identify the Delta VOC and the Omicron sublineages BA.1 and BA.2, respectively.

After verification of the analytical specificity of each primer set, two duplex qPCR assays with melting curve analysis were performed to screen 129 COVID-19 cases confirmed between December 31, 2021 and February 01, 2022 in Sfax, Tunisia.

The first duplex assay targeting ΔEF156–157 and Ins214EPE mutations successfully detected the Delta VOC in 39 cases and Omicron BA.1 in 83 cases. All the remaining cases (n = 7) were identified as Omicron BA.2, by the second duplex assay targeting Ins214EPE and ΔLPP24–26 mutations. The results of the screening method were in perfect concordance with those of S gene partial sequencing. In conclusion, our findings provide a simple and flexible screening method for more rapid and reliable monitoring of circulating VOCs. We highly recommend its implementation to guide public health policies.