Direct Detection of SARS-CoV-2 Using Reverse Transcription Loop-Mediated Isothermal Amplification (RT-LAMP) Without RNA Purification (preprint)

Coronavirus disease 2019 (COVID-19) is an acute respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) which has spread globally. Shortage of nucleic acid extraction kits may delay the speed of diagnosis. In order to address this need, we propose a simple method for direct RNA extraction from nasopharyngeal swab samples without RNA purification. Here, we describe a reverse transcription loop-mediated isothermal amplification technique with 95.4% sensitivity (95% CI: 84.2-99.4%) and 100% specificity (95% CI: 78-100%). Combined with a simple RNA extraction step, this optimized RT-LAMP method is able to amplify SARS-CoV-2 directly from clinical nasopharyngeal swab samples in viral transport media in 50 minutes. This method provides an opportunity to facilitate SARS-CoV-2 detection especially in resource limited settings where nucleic acid extraction kits are few.Funding: This study was supported by Prototype Research Grant Scheme (PRGS), PR001-2020B (PRGS/2/2020/SKK09/UM/02/1) from the Ministry of Education, Malaysia.Declaration of Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this manuscript.Ethics Approval Statement: Ethical approvals for this study was obtained from UMMC 85 Medical Ethics Committee (202041-8418) and Medical Research Ethics Committee 86 (MREC) Ministry of Health Malaysia (NMRR-20-2344-56994).

Whole Genome Sequence Analysis of SARS-CoV-2 Strains Circulating in Malaysia During First Wave and Early Second Wave of Infections. (preprint)

Background: Since December 2019, the outbreak of COVID-19 has raised a great public health concern globally. Here, we report the whole genome sequencing analysis of SARS-CoV-2 strains in Malaysia isolated from six patients diagnosed with COVID-19.Methods: The SARS-CoV-2 viral RNA extracted from clinical specimens and isolates were subjected to whole genome sequencing using NextSeq 500 platform. The sequencing data were assembled to full genome sequences using Megahit and phylogenetic tree was constructed using Mega X software.Results: Six full genome sequences of SARS-CoV-2 comprising of strains from 1st wave (25th January 2020) and 2nd wave (27th February 2020) infection were obtained. Downstream analysis demonstrated diversity among the Malaysian strains with several synonymous and non-synonymous mutations in four of the six cases, affecting the genes M, orf1ab, and S of the SARS-CoV-2 virus. The phylogenetic analysis revealed viral genome sequences of Malaysian SARS-CoV-2 strains clustered under the ancestral Type B.Conclusion: This study comprehended the SARS-CoV-2 virus evolution during its circulation in Malaysia. Continuous monitoring and analysis of the whole genome sequences of confirmed cases would be crucial to further understand the genetic evolution of the virus.