Direct detection of SARS-CoV-2 using CRISPR-Cas13a and a mobile phone

Fozouni, P.; Son, S.; Diaz de Leon Derby, M.; Knott, G. J.; Gray, C. N.; D'Ambrosio, M. V.; Zhao, C.; Switz, N. A.; Kumar, G. R.; Stephens, S. I.; Boehm, D.; Tsou, C.-L.; Shu, J.; Bhuiya, A.; Armstrong, M.; Harris, A.; Osterloh, J. M.; Meyer-Franke, A.; Langelier, C.; Pollard, K. S.; Crawford, E. D.; Puschnik, A. S.; Phelps, M.; Kistler, A.; DeRisi, J. L.; Doudna, J. A.; Fletcher, D. A.; Ott, M.

The December 2019 outbreak of a novel respiratory virus, SARS-CoV-2, has become an ongoing global pandemic due in part to the challenge of identifying symptomatic, asymptomatic and pre- symptomatic carriers of the virus. CRISPR-based diagnostics that utilize RNA and DNA-targeting enzymes can augment gold-standard PCR-based testing if they can be made rapid, portable and accurate. Here we report the development of an amplification-free CRISPR-Cas13a-based mobile phone assay for direct detection of SARS-CoV-2 from nasal swab RNA extracts. The assay achieved ~100 copies/L sensitivity in under 30 minutes and accurately detected a set of positive clinical samples in under 5 minutes. We combined crRNAs targeting SARS-CoV-2 RNA to improve sensitivity and specificity, and we directly quantified viral load using enzyme kinetics. Combined with mobile phone-based quantification, this assay can provide rapid, low-cost, point-of-care screening to aid in the control of SARS-CoV-2.