Multiplex, variant-tolerant, real-time RT-LAMP for SARS-CoV-2 detection using HFman probe

Dong, Y. et al. (2022) Multiplex, variant-tolerant, real-time RT-LAMP for SARS-CoV-2 detection using HFman probe. ACS Sensors, 7(3), pp. 730-739. (doi: 10.1021/acssensors.1c02079) (PMID:35192340) (PMCID:PMC8887655)

Abstract

Viral evolution impacts diagnostic test performance through the emergence of variants with sequences affecting the efficiency of primer binding. Such variants that evade detection by nucleic acid-based tests are subject to selective pressure, enabling them to spread more efficiently. Here, we report a variant-tolerant diagnostic test for SARS-CoV-2 using a loop-mediated isothermal nucleic acid-based amplification (LAMP) assay containing high-fidelity DNA polymerase and a high-fidelity DNA polymerase-medicated probe (HFman probe). In addition to demonstrating a high tolerance to variable SARS-CoV-2 viral sequences, the mechanism also overcomes frequently observed limitations of LAMP assays arising from non-specific amplification within multiplexed reactions performed in a single “pot”. Results showed excellent clinical performance (sensitivity 94.5%, specificity 100%, n = 190) when compared directly to a commercial gold standard reverse transcription quantitative polymerase chain reaction assay for the extracted RNA from nasopharyngeal samples and the capability of detecting a wide range of sequences containing at least alpha and delta variants. To further validate the test with no sample processing, directly from nasopharyngeal swabs, we also detected SARS-CoV-2 in positive clinical samples (n = 49), opening up the possibility for the assay’s use in decentralized testing.

• Item Type: Articles
• Keywords: Multiplex LAMP, HFman probe, high-fidelity DNA polymerase, non-specific amplification, COVID-19/SARS- CoV-2, point-of-care testing (POCT).
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Cooper, Professor Jonathan and Reboud, Dr Julien
• Authors: Dong, Y., Zhao, Y., Li, S., Wan, Z., Lu, R., Yang, X., Yu, G., Reboud, J., Cooper, J. M., Tian, Z., and Zhang, C.
• College/School: College of Science and Engineering > School of Engineering > Biomedical Engineering
• Journal Name: ACS Sensors
• Publisher: American Chemical Society
• ISSN: 2379-3694
• ISSN (Online): 2379-3694
• Published Online: 22 February 2022
• Copyright Holders: Copyright © 2022 American Chemical Society
• First Published: First published in ACS Sensors 7(3): 730-739
• Publisher Policy: Reproduced under a Creative Commons licence

Project Code	Award No	Project Name	Principal Investigator	Funder's Name	Funder Ref	Lead Dept
300573		Novel low cost diagnostic tools and their impact in Africa	Jonathan Cooper	Engineering and Physical Sciences Research Council (EPSRC)	EP/R01437X/1	ENG - Biomedical Engineering
311995		COVID	Jonathan Cooper	Medical Research Council (MRC)	MR/V035401/1	ENG - Biomedical Engineering