ABSTRACT
SARS-CoV-2, influenza, and respiratory syncytial viruses (RSVs) cause acute respiratory infections with similar symptoms. Since the treatments and outcomes of these infections are different, the early detection and accurate differentiation of the viruses are clinically important for the prevention and treatment of the diseases. We previously demonstrated that clustered regularly interspaced short palindromic repeats (CRISPR) could rapidly and precisely detect SARS-CoV-2. The objective of this study was to develop CRISPR as a test for simultaneously detecting and accurately distinguishing the viruses. The CRISPR assay with an RNA guide against each virus was performed in the reference standards of SARS-CoV-2, influenza A and B, and RSV. The CRISPR assay had a limit of detection of 1-100 copies/µL for specifically detecting SARS-CoV-2, influenza A and B, and RSV without cross-reaction with other respiratory viruses. The validation of the test in nasopharyngeal specimens showed that it had a 90-100% sensitivity and 100% specificity for the detection of SARS-CoV-2, influenza A and B, and RSV. The CRISPR assay could potentially be used for sensitive detection and specific differentiation of the respiratory viruses.