SPECIFIC DETECTION OF SARS-COV-2 VARIANTS B.1.1.7 (ALPHA) AND B.1.617.2 (DELTA) USING A ONE-STEP QUANTITATIVE PCR ASSAY

Erster, O.; Mendelson, E.; Kabat, A.; Levy, V.; Mannasse, B.; Asraf, H.; Azar, R.; Ali, Y.; Bucris, E.; Bar-Ilan, D.; Mor, O.; Elul, M.; Mandelboim, M.; Sofer, D.; Fleishon, S.; Zuckerman, N. S.; Bar-Or, I.

In this report, we describe the development of an RT-qPCR assay, termed Alpha Delta assay, which can detect SARS-COV-2 (SC-2) and distinguish between the Alpha (B.1.1.7) and Delta (B.1.617.2) variants. The Alpha- and Delta-specific reactions in the assay target mutations that are strongly linked to the target variant. The Alpha reaction targets the D3L substitution in N gene, and the Delta reaction targets the spike gene 156-158 mutations. Additionally, we describe a second Delta-specific assay that we use as a confirmatory test for the Alpha Delta assay that targets the 119-120 deletion in the Orf8 gene. Both reactions have similar sensitivities of 15-25 copies per reaction, similar to the sensitivity of commercial SC-2 detection tests. The Alpha Delta assay and the Orf8-119del assay were successfully used to classify clinical samples that were subsequently analyzed by whole genome sequencing. Lastly, the capability of the Alpha Delta assay and Orf8-119del assay to identify correctly the presence of Delta RNA in wastewater samples was demonstrated. This study provides a rapid, sensitive and cost-effective tool for detecting and classifying two worldwide dominant SC-2 variants. It also highlights the importance of a timely diagnostic response to the emergence of new SC-2 variants with significant consequences on global health.