RT-LAMP: A Cheaper, Simpler and Faster Alternative for the Detection of SARS-CoV-2 in Wastewater.

Autor: Amoah ID; Institute for Water and Wastewater Technology, Durban University of Technology, P.O. Box 1334, Durban, 4000, South Africa., Mthethwa NP; Institute for Water and Wastewater Technology, Durban University of Technology, P.O. Box 1334, Durban, 4000, South Africa., Pillay L; Institute for Water and Wastewater Technology, Durban University of Technology, P.O. Box 1334, Durban, 4000, South Africa., Deepnarain N; Institute for Water and Wastewater Technology, Durban University of Technology, P.O. Box 1334, Durban, 4000, South Africa., Pillay K; Institute for Water and Wastewater Technology, Durban University of Technology, P.O. Box 1334, Durban, 4000, South Africa., Awolusi OO; Institute for Water and Wastewater Technology, Durban University of Technology, P.O. Box 1334, Durban, 4000, South Africa., Kumari S; Institute for Water and Wastewater Technology, Durban University of Technology, P.O. Box 1334, Durban, 4000, South Africa., Bux F; Institute for Water and Wastewater Technology, Durban University of Technology, P.O. Box 1334, Durban, 4000, South Africa. faizalb@dut.ac.za.
Jazyk: angličtina
Zdroj: Food and environmental virology [Food Environ Virol] 2021 Dec; Vol. 13 (4), pp. 447-456. Date of Electronic Publication: 2021 Jul 26.
DOI: 10.1007/s12560-021-09489-7
Abstrakt: Reverse transcription loop-mediated isothermal amplification (RT-LAMP) has the potential to become a cheaper and faster option for monitoring COVID-19 infections through wastewater-based epidemiology. However, its application in COVID-19 surveillance has been limited to clinical testing only. We present in this paper two optimized RT-LAMP protocols based on colour change and fluorescence detection and application of these protocols for wastewater monitoring from four wastewater treatment plants over 4 weeks. The optimized RT-LAMP protocols have a limit of detection of 10 copies/25 µl reaction with positive amplification within 35 minutes. Over the 4 weeks of monitoring, the colorimetric protocol detected a prevalence of 12.5%, when 1 µl of extracted RNA with 92.7(± 28.2) ng/µl concentration was analysed. When the RNA template was increased by fivefold, the prevalence increased to 44%. The fluorescent RT-LAMP had a prevalence of 31% and 47% for starting templates of 92.7(± 28.2) ng/µl and 480(± 134.5) ng/µl of the extracted RNA, respectively. All samples were positive for SARS-CoV-2 when analysed with droplet digital PCR, with viral loads ranging from 18.1 to 195.6 gc/ml of wastewater. The RT-ddPCR, therefore, confirms the presence of the viral RNA in the wastewater samples, albeit at low concentrations. Additionally, the RT-LAMP protocols positively detected SARS-CoV-2 in wastewater samples with copies as low as 20.7 gc/ml. The results obtained in our study show the potential application of RT-LAMP for the detection of SARS-CoV-2 in wastewater, which could provide a cheaper and faster alternative to RT-qPCR or RT-ddPCR for wastewater-based epidemiological monitoring of COVID-19 and other viral infections.
(© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)
Databáze: MEDLINE
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