TY - JOUR TI - FnCas9-based CRISPR diagnostic for rapid and accurate detection of major SARS-CoV-2 variants on a paper strip AU - Kumar, Manoj AU - Gulati, Sneha AU - Ansari, Asgar H AU - Phutela, Rhythm AU - Acharya, Sundaram AU - Azhar, Mohd AU - Murthy, Jayaram AU - Kathpalia, Poorti AU - Kanakan, Akshay AU - Maurya, Ranjeet AU - Vasudevan, Janani Srinivasa AU - S, Aparna AU - Pandey, Rajesh AU - Maiti, Souvik AU - Chakraborty, Debojyoti A2 - Krishnan, Yamuna A2 - VijayRaghavan, K A2 - Chandrasekaran, Arun Richard VL - 10 PY - 2021 DA - 2021/06/09 SP - e67130 C1 - eLife 2021;10:e67130 DO - 10.7554/eLife.67130 UR - https://doi.org/10.7554/eLife.67130 AB - The COVID-19 pandemic originating in the Wuhan province of China in late 2019 has impacted global health, causing increased mortality among elderly patients and individuals with comorbid conditions. During the passage of the virus through affected populations, it has undergone mutations, some of which have recently been linked with increased viral load and prognostic complexities. Several of these variants are point mutations that are difficult to diagnose using the gold standard quantitative real-time PCR (qRT-PCR) method and necessitates widespread sequencing which is expensive, has long turn-around times, and requires high viral load for calling mutations accurately. Here, we repurpose the high specificity of Francisella novicida Cas9 (FnCas9) to identify mismatches in the target for developing a lateral flow assay that can be successfully adapted for the simultaneous detection of SARS-CoV-2 infection as well as for detecting point mutations in the sequence of the virus obtained from patient samples. We report the detection of the S gene mutation N501Y (present across multiple variant lineages of SARS-CoV-2) within an hour using lateral flow paper strip chemistry. The results were corroborated using deep sequencing on multiple wild-type (n = 37) and mutant (n = 22) virus infected patient samples with a sensitivity of 87% and specificity of 97%. The design principle can be rapidly adapted for other mutations (as shown also for E484K and T716I) highlighting the advantages of quick optimization and roll-out of CRISPR diagnostics (CRISPRDx) for disease surveillance even beyond COVID-19. This study was funded by Council for Scientific and Industrial Research, India. KW - CRISPRDx KW - FnCas9 KW - COVID-19 KW - variants KW - FELUDA KW - N501Y JF - eLife SN - 2050-084X PB - eLife Sciences Publications, Ltd ER -