Rapid and Accurate Detection of SARS Coronavirus 2 by Nanopore Amplicon Sequencing.

Autor:	Li XX; Department of Pharmacy, Department of Intensive Care Unit, and Department of Medical Affairs, Peking University Third Hospital, Beijing, China., Li C; Department of Pharmacy, Department of Intensive Care Unit, and Department of Medical Affairs, Peking University Third Hospital, Beijing, China., Du PC; Beijing YuanShengKangTai (ProtoDNA) Genetech Co. Ltd., Beijing, China., Li SY; Department of Pharmacy, Department of Intensive Care Unit, and Department of Medical Affairs, Peking University Third Hospital, Beijing, China., Yu L; Beijing YuanShengKangTai (ProtoDNA) Genetech Co. Ltd., Beijing, China., Zhao ZQ; Beijing YuanShengKangTai (ProtoDNA) Genetech Co. Ltd., Beijing, China., Liu TT; Beijing YuanShengKangTai (ProtoDNA) Genetech Co. Ltd., Beijing, China., Zhang CK; Beijing YuanShengKangTai (ProtoDNA) Genetech Co. Ltd., Beijing, China., Zhang SC; Beijing YuanShengKangTai (ProtoDNA) Genetech Co. Ltd., Beijing, China., Zhuang Y; Department of Pharmacy, Department of Intensive Care Unit, and Department of Medical Affairs, Peking University Third Hospital, Beijing, China., Dong CR; Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China., Ge QG; Department of Pharmacy, Department of Intensive Care Unit, and Department of Medical Affairs, Peking University Third Hospital, Beijing, China.
Jazyk:	angličtina
Zdroj:	Frontiers in microbiology [Front Microbiol] 2022 Apr 06; Vol. 13, pp. 735363. Date of Electronic Publication: 2022 Apr 06 (Print Publication: 2022).
DOI:	10.3389/fmicb.2022.735363
Abstrakt:	Objective: We aimed to evaluate the performance of nanopore amplicon sequencing detection for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in clinical samples. Method: We carried out a single-center, prospective cohort study in a Wuhan hospital and collected a total of 86 clinical samples, including 54 pharyngeal swabs, 31 sputum samples, and 1 fecal sample, from 86 patients with coronavirus disease 2019 (COVID-19) from Feb 20 to May 15, 2020. We performed parallel detection with nanopore-based genome amplification and sequencing (NAS) on the Oxford Nanopore Technologies (ONT) minION platform and routine reverse transcription quantitative polymerase chain reaction (RT-qPCR). In addition, 27 negative control samples were detected using the two methods. The sensitivity and specificity of NAS were evaluated and compared with those of RT-qPCR. Results: The viral read number and reference genome coverage were both significantly different between the two groups of samples, and the latter was a better indicator for SARS-CoV-2 detection. Based on the reference genome coverage, NAS revealed both high sensitivity (96.5%) and specificity (100%) compared with RT-qPCR (80.2 and 96.3%, respectively), although the samples had been stored for half a year before the detection. The total time cost was less than 15 h, which was acceptable compared with that of RT-qPCR (∼2.5 h). In addition, the reference genome coverage of the viral reads was in line with the cycle threshold value of RT-qPCR, indicating that this number could also be used as an indicator of the viral load in a sample. The viral load in sputum might be related to the severity of the infection, particularly in patients within 4 weeks after onset of clinical manifestations, which could be used to evaluate the infection. Conclusion: Our results showed the high sensitivity and specificity of the NAS method for SARS-CoV-2 detection compared with RT-qPCR. The sequencing results were also used as an indicator of the viral load to display the viral dynamics during infection. This study proved the wide application prospect of nanopore sequencing detection for SARS-CoV-2 and may more knowledge about the clinical characteristics of COVID-19. Competing Interests: P-CD, LY, Z-QZ, T-TL, C-KZ, and S-CZ were employed by Beijing YuanShengKangTai (ProtoDNA) Genetech Co. Ltd., Beijing, China. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. (Copyright © 2022 Li, Li, Du, Li, Yu, Zhao, Liu, Zhang, Zhang, Zhuang, Dong and Ge.)
Databáze:	MEDLINE
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