Objective To evaluate a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and compare it with RT-PCR. Methods We designed primers specific to the orf1ab and S genes of SARS-CoV-2. Total viral RNA was extracted using the QIAamp Viral RNA Mini Kit. We optimized the RT-LAMP assay, and evaluated it for its sensitivity and specificity of detection using real-time turbidity monitoring and visual observation. Results The primer sets orf1ab-4 and S-123 amplified the genes in the shortest times, the mean (±SD) times were 18 ± 1.32 min and 20 ± 1.80 min, respectively, and 63°C was the optimum reaction temperature. The sensitivities were 2 × 10 1 copies and 2 × 10 2 copies per reaction with primer sets orf1ab-4 and S-123, respectively. This assay showed no cross-reactivity with 60 other respiratory pathogens. To describe the availability of this method in clinical diagnosis, we collected 130 specimens from patients with clinically suspected SARS-CoV-2 infection. Among them, 58 were confirmed to be positive and 72 were negative by RT-LAMP. The sensitivity was 100% (95% CI 92.3%–100%), specificity 100% (95% CI 93.7%–100%). This assay detected SARS-CoV-2 in a mean (±SD) time of 26.28 ± 4.48 min and the results can be identified with visual observation. Conclusion These results demonstrate that we developed a rapid, simple, specific and sensitive RT-LAMP assay for SARS-CoV-2 detection among clinical samples. It will be a powerful tool for SARS-CoV-2 identification, and for monitoring suspected patients, close contacts and high-risk groups.
【저자키워드】 COVID-19, SARS-CoV-2, detection, RT-LAMP, Visual, 【초록키워드】 coronavirus, SARS-COV-2 infection, RT-PCR, RNA, cross-reactivity, clinical samples, SARS-CoV-2 detection, sensitivity, specificity, Sensitivity and specificity, Patient, Respiratory pathogens, temperature, reverse transcription, isothermal amplification, Viral RNA, Clinical diagnosis, S gene, close contact, observation, acute respiratory syndrome, 95% CI, specimen, powerful tool, primer, positive, suspected patients, MINI, objective, Result, amplified, collected, evaluate, primer set, evaluated, clinically suspected, groups, the mean, Total, 【제목키워드】 2019 novel coronavirus, Rapid, reverse transcription, isothermal amplification,