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SHORT REPORT SIMPLE RNA EXTRACTION METHOD FOR REVERSE TRANSCRIPTION LOOP-MEDIATED AMPLIFICATION DETECTION OF SARS-COV-2

Authors

  • Yee Ling Lau Department of Parasitology, Faculty of Medicine, University of Malaya
  • Nur Zulaikha Zulkefli Department of Parasitology, Faculty of Medicine, Universiti Malaya, 50603 Kuala Lumpur, Malaysia
  • Fatma Diyana Mohd Bukhari Department of Parasitology, Faculty of Medicine, Universiti Malaya, 50603 Kuala Lumpur, Malaysia
  • Meng Yee Lai Department of Parasitology, Faculty of Medicine, Universiti Malaya, 50603 Kuala Lumpur, Malaysia
  • Jeyanthi Suppiah Virology Unit, Infectious Disease Research Centre, Institute for Medical Research, National Institutes of Health, Ministry of Health Malaysia
  • Ravindran Thayan Virology Unit, Infectious Disease Research Centre, Institute for Medical Research, National Institutes of Health, Ministry of Health Malaysia

Keywords:

COVID-19, rapid detection, LAMP assay, SARS-CoV-2, Chelex resin

Abstract

Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a robust and cost-effective assay for rapid diagnosis of SARS-CoV-2 compared to reverse transcription quantitative (RT-q) PCR. The study evaluated the performance of RT-LAMP technique that incorporated a simple Chelex 100 resin-based RNA extraction step for SARS-CoV-2 detection targeting virus E (encoding envelope protein) and RdRP (encoding RNA-dependent RNA polymerase). Using primer sets for E and RdRP, the developed RT-LAMP assay had a limit of detection (LOD) of 1 copy/µl transcribed RNA. For nasopharyngeal and oropharyngeal swab samples (n = 58), in comparison to the gold standard RT-qPCR (amplifying E and RdRP) sensitivity, specificity, positive predictive value, and negative predictive value of SARS-CoV-2 RT-LAMP assay targeting E gene was 88% (95% confidence interval (CI): 75-96%), 87% (95% CI: 59-98%), 99% (95% CI: 97-100%), and 28% (95% CI: 14-48%), respectively and for RdRP gene was 67% (95% CI: 51-98%), 87% (95% CI: 59-98%), 100% (95% CI: 96-100%), and 12% (95% CI : 8-18%), respectively. The whole process of RT-LAMP assay was completed within ~60 minutes. This developed RT-LAMP method for on-site COVID-19 detection should be useful in resource limited settings.

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Published

2022-07-23

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