tetano
Editor, Senior Moderator
Clin Microbiol Infect. 2020 Apr 7. pii: S1198-743X(20)30186-5. doi: 10.1016/j.cmi.2020.04.001. [Epub ahead of print]
Rapid and visual detection of 2019 novel coronavirus (SARS-CoV-2) by a reverse transcription loop-mediated isothermal amplification assay.
Yan C[SUP]1[/SUP], Cui J[SUP]1[/SUP], Huang L[SUP]2[/SUP], Du B[SUP]1[/SUP], Chen L[SUP]3[/SUP], Xue G[SUP]1[/SUP], Li S[SUP]1[/SUP], Zhang W[SUP]1[/SUP], Zhao L[SUP]1[/SUP], Sun Y[SUP]1[/SUP], Yao H[SUP]1[/SUP], Li N[SUP]1[/SUP], Zhao H[SUP]1[/SUP], Feng Y[SUP]1[/SUP], Liu S[SUP]1[/SUP], Zhang Q[SUP]1[/SUP], Liu D[SUP]4[/SUP], Yuan J[SUP]5[/SUP].
Author information
Abstract
OBJECTIVE:
To evaluate a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for detection of SARS-CoV-2, and compare it with RT polymerase chain reaction (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, this assay was evaluated 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) time was 18 ? 1.32 min and 20 ? 1.80 min, respectively, and 63?C was the optimum reaction temperature. The sensitivity was 2?10[SUP]1[/SUP] copies and 2?10[SUP]2[/SUP] copies per reaction with primer sets orf1ab-4 and S-123, respectively. This assay showed no cross-reactivity with other 60 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 sensiticity was 100% (95% CI 92.3% - 100%), specificity 100% (95% CI 93.7% - 100%). This assay detected SARS-CoV-2 in the 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.
Copyright ? 2020. Published by Elsevier Ltd.
PMID:32276116PMCID:PMC7144850DOI:10.1016/j.cmi.2020.04.001
Rapid and visual detection of 2019 novel coronavirus (SARS-CoV-2) by a reverse transcription loop-mediated isothermal amplification assay.
Yan C[SUP]1[/SUP], Cui J[SUP]1[/SUP], Huang L[SUP]2[/SUP], Du B[SUP]1[/SUP], Chen L[SUP]3[/SUP], Xue G[SUP]1[/SUP], Li S[SUP]1[/SUP], Zhang W[SUP]1[/SUP], Zhao L[SUP]1[/SUP], Sun Y[SUP]1[/SUP], Yao H[SUP]1[/SUP], Li N[SUP]1[/SUP], Zhao H[SUP]1[/SUP], Feng Y[SUP]1[/SUP], Liu S[SUP]1[/SUP], Zhang Q[SUP]1[/SUP], Liu D[SUP]4[/SUP], Yuan J[SUP]5[/SUP].
Author information
Abstract
OBJECTIVE:
To evaluate a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for detection of SARS-CoV-2, and compare it with RT polymerase chain reaction (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, this assay was evaluated 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) time was 18 ? 1.32 min and 20 ? 1.80 min, respectively, and 63?C was the optimum reaction temperature. The sensitivity was 2?10[SUP]1[/SUP] copies and 2?10[SUP]2[/SUP] copies per reaction with primer sets orf1ab-4 and S-123, respectively. This assay showed no cross-reactivity with other 60 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 sensiticity was 100% (95% CI 92.3% - 100%), specificity 100% (95% CI 93.7% - 100%). This assay detected SARS-CoV-2 in the 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.
Copyright ? 2020. Published by Elsevier Ltd.
PMID:32276116PMCID:PMC7144850DOI:10.1016/j.cmi.2020.04.001