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A fast extraction-free isothermal LAMP assay for detection of SARS-CoV-2 with potential use in resource-limited settings

Gartner, Kathleen; Meleke, Harry; Kamdolozi, Mercy; Chaima, David; Samikwa, Lyson; Paynter, Mary; Nyang'Wa, Maggie Nyirenda; ... Alber, Dagmar G; + view all (2022) A fast extraction-free isothermal LAMP assay for detection of SARS-CoV-2 with potential use in resource-limited settings. Virology Journal , 19 (1) , Article 77. 10.1186/s12985-022-01800-7. Green open access

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Abstract

BACKGROUND: To retain the spread of SARS-CoV-2, fast, sensitive and cost-effective testing is essential, particularly in resource limited settings (RLS). Current standard nucleic acid-based RT-PCR assays, although highly sensitive and specific, require transportation of samples to specialised laboratories, trained staff and expensive reagents. The latter are often not readily available in low- and middle-income countries and this may significantly impact on the successful disease management in these settings. Various studies have suggested a SARS-CoV-2 loop mediated isothermal amplification (LAMP) assay as an alternative method to RT-PCR. METHODS: Four previously published primer pairs were used for detection of SARS-CoV-2 in the LAMP assay. To determine optimal conditions, different temperatures, sample input and incubation times were tested. Ninety-three extracted RNA samples from St. George's Hospital, London, 10 non-extracted nasopharyngeal swab samples from Great Ormond Street Hospital for Children, London, and 92 non-extracted samples from Queen Elisabeth Central Hospital (QECH), Malawi, which have previously been tested for SARS-Cov-2 by quantitative reverse-transcription RealTime PCR (qRT-PCR), were analysed in the LAMP assay. RESULTS: In this study we report the optimisation of an extraction-free colourimetric SARS-CoV-2 LAMP assay and demonstrated that a lower limit of detection (LOD) between 10 and 100 copies/µL of SARS-CoV-2 could be readily detected by a colour change of the reaction within as little as 30 min. We further show that this assay could be quickly established in Malawi, as no expensive equipment is necessary. We tested 92 clinical samples from QECH and showed the sensitivity and specificity of the assay to be 86.7% and 98.4%, respectively. Some viral transport media, used routinely to stabilise RNA in clinical samples during transportation, caused a non-specific colour-change in the LAMP reaction and therefore we suggest collecting samples in phosphate buffered saline (which did not affect the colour) as the assay allows immediate sample analysis on-site. CONCLUSION: SARS-CoV-2 LAMP is a cheap and reliable assay that can be readily employed in RLS to improve disease monitoring and management.

Type: Article
Title: A fast extraction-free isothermal LAMP assay for detection of SARS-CoV-2 with potential use in resource-limited settings
Location: England
Open access status: An open access version is available from UCL Discovery
DOI: 10.1186/s12985-022-01800-7
Publisher version: https://doi.org/10.1186/s12985-022-01800-7
Language: English
Additional information: Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visithttp://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
Keywords: Science & Technology, Life Sciences & Biomedicine, Virology, SARS-CoV-2, qRT-PCR, Extraction-free LAMP, Resource-limited settings, AMPLIFICATION
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health > Infection, Immunity and Inflammation Dept
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
URI: https://discovery.ucl.ac.uk/id/eprint/10148593
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