CRISPR-Cas3-based diagnostics for SARS-CoV-2 and influenza virus

Kazuto Yoshimi; Kohei Takeshita; Seiya Yamayoshi; Satomi Shibumura; Yuko Yamauchi; Masaki Yamamoto; Hiroshi Yotsuyanagi; Yoshihiro Kawaoka; Tomoji Mashimo

doi:10.1016/j.isci.2022.103830

CRISPR-Cas3-based diagnostics for SARS-CoV-2 and influenza virus

iScience. 2022 Feb 18;25(2):103830. doi: 10.1016/j.isci.2022.103830. Epub 2022 Jan 30.

Authors

Kazuto Yoshimi^{1

2}, Kohei Takeshita³, Seiya Yamayoshi⁴, Satomi Shibumura⁵, Yuko Yamauchi¹, Masaki Yamamoto³, Hiroshi Yotsuyanagi⁶, Yoshihiro Kawaoka^{4

7}, Tomoji Mashimo^{1

2}

Affiliations

¹ Division of Animal Genetics, Laboratory Animal Research Center, Institute of Medical Science, The University of Tokyo, Tokyo 108-8639, Japan.
² Division of Genome Engineering, Center for Experimental Medicine and Systems Biology, Institute of Medical Science, The University of Tokyo, Tokyo 108-8639, Japan.
³ Advanced Photon Technology Division, RIKEN SPring-8 Center, Hyogo 679-5148, Japan.
⁴ Division of Virology, Department of Microbiology and Immunology, Institute of Medical Science, The University of Tokyo, Tokyo 108-8639, Japan.
⁵ C4U Corporation, Osaka 565-0871, Japan.
⁶ Division of Infectious Diseases and Applied Immunology, Institute of Medical Science, The University of Tokyo, Tokyo 108-8639, Japan.
⁷ Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, Madison, WI 53711, USA.

Abstract

CRISPR-based diagnostics (CRISPR-dx), including the Cas12-based DETECTR and Cas13-based SHERLOCK Class 2 CRISPRs, have been used to detect the presence of DNA or RNA from pathogens, such as the 2009 pandemic influenza virus A (IAV) and the 2019 novel coronavirus SARS-CoV-2. Here, we describe the collateral single-stranded DNA cleavage with Class 1 type I CRISPR-Cas3 and highlight its potential for development as a Cas3-mediated rapid (within 40 min), low-cost, instrument-free detection method for SARS-CoV-2. This assay, which we call Cas3-Operated Nucleic Acid detectioN (CONAN), not only detects SARS-CoV-2 in clinical samples, but also offers specific detection of single-base-pair mutations in IAV variants. This tool allows rapid and accurate point-of-care testing for patients with suspected SARS-CoV-2 or drug-resistant IAV infections in hospitals.

Keywords: Diagnostics; Virology.

Grants and funding

HHSN272201400008C/AI/NIAID NIH HHS/United States