A two-step process for in silico screening to assess the performance of qRTPCR kits against variant strains of SARS-CoV-2.

ABSTRACT

BACKGROUND: Since inception of the COVID-19 pandemic, early detection and isolation of positive cases is one of the key strategies to restrict disease transmission. Real time reverse transcription polymerase chain reaction (qRTPCR) has been the mainstay of diagnosis. Most of the qRTPCR kits were designed against the target genes of original strain of SARS-CoV-2. However, with the emergence of variant strains of SARS-CoV-2, sensitivity of the qRTPCR assays has reportedly reduced. In view of this, it is critical to continuously monitor the performance of the qRTPCR kits in the backdrop of variant strains of SARS-CoV-2. Real world monitoring of assay performance is challenging. Therefore, we developed a two-step in-silico screening process for evaluating the performance of various qRTPCR kits used in India. RESULTS: We analysed 73 qRT-PCR kits marketed in India, against the two SARS-CoV-2 VoCs. Sequences of both Delta (B.1.617.2) and Omicron (B.1.1.529) VoCs submitted to GISAID within a specific timeframe were downloaded, clustered to identify unique sequences and aligned with primer and probe sequences. Results were analysed following a two-step screening process. Out of 73 kits analysed, seven were unsatisfactory for detection of both Delta and Omicron VoCs, 10 were unsatisfactory for Delta VoC whereas 2 were unsatisfactory for only Omicron VoC. CONCLUSION: Overall, we have developed a useful screening process for evaluating the performance of qRTPCR assays against Delta and Omicron VoCs of SARS-CoV-2 which can be used for detecting SARS-CoV-2 VoCs that may emerge in future and can also be redeployed for other evolving pathogens of public health importance.