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)

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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).
Glasgow Author(s) Enlighten ID:Cooper, Professor Jonathan and Reboud, Professor 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 (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

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
300573Novel low cost diagnostic tools and their impact in AfricaJonathan CooperEngineering and Physical Sciences Research Council (EPSRC)EP/R01437X/1ENG - Biomedical Engineering
311995COVIDJonathan CooperMedical Research Council (MRC)MR/V035401/1ENG - Biomedical Engineering