Rapid and visual detection of 2019 novel coronavirus (SARS-CoV-2) by a reverse transcription loop-mediated isothermal amplification assay

Rapid and visual detection of 2019 novel coronavirus (SARS-CoV-2) by a reverse transcription loop-mediated isothermal amplification assay

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. We designed primers specific to the orf1ab and S genes of SARS-CoV-2. Total viral RNA was extracted us...

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Bibliographic Details
Published in:Clinical microbiology and infection 2020-06, Vol.26 (6), p.773-779
Main Authors: Yan, C., Cui, J., Huang, L., Du, B., Chen, L., Xue, G., Li, S., Zhang, W., Zhao, L., Sun, Y., Yao, H., Li, N., Zhao, H., Feng, Y., Liu, S., Zhang, Q., Liu, D., Yuan, J.
Format: Article
Language:eng
Subjects:
COVID-19
Detection
RT-LAMP
SARS-CoV-2
Visual
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Summary: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. 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. 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 × 101 copies and 2 × 102 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. 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.
ISSN:1198-743X
1469-0691