RUNCOV: A one-pot triplex real-time RT-LAMP as a point-of-care diagnostic tool for detecting SARS-CoV-2
Robène I., Jouen E., Maillot-Lebon M.V., Fenelon B., Hascoat J., Pecrix Y., Richard D., Jaffar-Bandjee M.C., Mavingui P., Chiroleu F., Becker N., Poubeau P., Ramiandrisoa M., Sin M., Costet L., Laurent A., Laurent P., Chabirand A., Moreau A., Reynaud B., Jeuffrault E., Cetre-Sossah C.. 2025. Biology Methods and Protocols, 10 (1) : 16 p..
Given the risk of zoonotic disease emergence, including new SARS-CoV-2 variants of COVID-19, rapid diagnostic tools are urgently needed to improve the control of the spread of infectious diseases. A one-pot triplex real-time RT-LAMP (reverse-transcription-loop-mediated isothermal amplification) assay, based on two regions of the genome SARS-CoV-2—specifically the Orf1ab and N genes—along with one internal control, the human RNase P gene, was developed. The multiplexing relies on the distinct melting peaks produced during an annealing step. This tool, named RUNCOV, was compared to the gold-standard reverse-transcription real-time quantitative PCR (RT-qPCR) assay. A simple sample preparation step was designed alongside the assay, making it ready for use on site, as a point-of-care diagnostic tool. RUNCOV is rapid (typically less than 40¿minutes), highly sensitive and specific. When tested on clinical samples with known SARS-CoV-2 status, its limit of detection (LOD) ranges between 5 and 20 copies per reaction and its diagnostic sensitivity (97.44%) and specificity (100%) values are high compared to the RT-qPCR gold standard. These results were supported with an extensive in silico analysis of over 14 million genomes, demonstrating this tool was capable of detecting all known SARS-CoV-2 variants, including the most recent ones KP.3.1.1 and BA2.86.1. This molecular assay is portable, as demonstrated when it was used successfully in La Réunion in different contexts outside the laboratory.
Mots-clés : coronavirus 2 du syndrome respiratoire aigu sévère; zoonose; covid-19; pcr; contrôle de maladies; génomique; maladie infectieuse; génome; france; la réunion
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Agents Cirad, auteurs de cette publication :
- Becker Nathalie — Bios / UMR PVBMT
- Cetre-Sossah Catherine — Bios / UMR ASTRE
- Chiroleu Frédéric — Bios / UMR PVBMT
- Costet Laurent — Bios / UMR PVBMT
- Hascoat Jérémy — Bios / UMR PVBMT
- Jeuffrault Eric — Dgdrd / Dgdrd - direction régionale Réunion-Mayotte
- Jouen Emmanuel — Bios / UMR PVBMT
- Maillot Marie-Véronique — Bios / UMR PVBMT
- Pecrix Yann — Bios / UMR PVBMT
- Reynaud Bernard — Bios / UMR PVBMT
- Robène Isabelle — Bios / UMR PVBMT