A RAPID POINT-OF-CARE DIPSTICK ASSAY FOR DIFFERENTIATION OF SARS-COV-2 VARIANTS IN COVID-19 PATIENTS

Abstract

Intro: Delta and Omicron variants of SARS-CoV-2 are highly contagious, currently dominating the globe and recognized as variant of concern (VOC). The transmissibility efficiency of viruses, disease symptoms, and severity of COVID- 19 is highly heterogeneous. Therefore, testing at the community level is essential to identify the infected people at an early stage-carrying VOC to reduce the spread of virus and combat the pandemic. Methods: In this study, we analysed thousands of genome sequences representing 30 different SARS-CoV-2 variants and identified Delta and Omicron variants specific nucleic acid signatures in the spike gene. Based on the variant specific nucleic acid sequences we synthesized different oligos and optimized a mPCR assay that can specifically differentiate the Delta and Omicron variants. We further translated our work into a dipstick assay (Tohoku Bio-array, Japan) by adding tag linker sequence to 5’ end of the forward primer and adding biotin in 3’ end of the oligos. Findings: A total of 250 samples were subjected to WGS using MiSeq platform and these confirmed samples were processed for validation of our specific designed probes using PCR assay and the readout was found to be 100% specific to Delta, BA.1 & BA.2 of SARS-CoV-2 variants which were further confirmed by Sanger sequencing. The dipstick was used to screen these samples, and specific signals were observed. WGS and Sanger sequencing were used to validate our PCR and dipstick assay results, and the readout was found to be 100% specific. The results can be visualised by the naked eye and interpreted easily. Conclusion: This study developed a rapid point-of-care test of SARS-CoV-2 patients, which can differentiate Delta, BA.1 and BA.2 variants at the same time of confirmation of the infection in patient. The current nucleic-acid chromatography-based dipstick assay is highly specific and can work even in the case of low viral load as well.