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Development of a Laboratory-safe and Low-cost Detection Protocol for SARS-CoV-2 of the Coronavirus Disease 2019 (COVID-19).

ABSTRACT: The severe acute respiratory coronavirus 2 (SARS-CoV-2), which emerged in December 2019 in Wuhan, China, has spread rapidly to over a dozen countries. Especially, the spike of case numbers in South Korea sparks pandemic worries. This virus is reported to spread mainly through person-to-person contact via respiratory droplets generated by coughing and sneezing, or possibly through surface contaminated by people coughing or sneezing on them. More critically, there have been reports about the possibility of this virus to transmit even before a virus-carrying person to show symptoms. Therefore, a low-cost, easy-access protocol for early detection of this virus is desperately needed. Here, we have established a real-time reverse-transcription PCR (rtPCR)-based assay protocol composed of easy specimen self-collection from a subject via pharyngeal swab, Trizol-based RNA purification, and SYBR Green-based rtPCR. This protocol shows an accuracy and sensitivity limit of 1-10 virus particles as we tested with a known lentivirus. The cost for each sample is estimated to be less than 15 US dollars. Overall time it takes for an entire protocol is estimated to be less than 4 hours. We propose a cost-effective, quick-and-easy method for early detection of SARS-CoV-2 at any conventional Biosafety Level II laboratories that are equipped with a rtPCR machine. Our newly developed protocol should be helpful for a first-hand screening of the asymptomatic virus-carriers for further prevention of transmission and early intervention and treatment for the rapidly propagating virus.

SUBMITTER: Won J

PROVIDER: S-EPMC7237269 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature

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Development of a Laboratory-safe and Low-cost Detection Protocol for SARS-CoV-2 of the Coronavirus Disease 2019 (COVID-19).

Won Joungha J Lee Solji S Park Myungsun M Kim Tai Young TY Park Mingu Gordon MG Choi Byung Yoon BY Kim Dongwan D Chang Hyeshik H Kim V Narry VN Lee C Justin CJ

Experimental neurobiology 20200401 2

The severe acute respiratory coronavirus 2 (SARS-CoV-2), which emerged in December 2019 in Wuhan, China, has spread rapidly to over a dozen countries. Especially, the spike of case numbers in South Korea sparks pandemic worries. This virus is reported to spread mainly through person-to-person contact via respiratory droplets generated by coughing and sneezing, or possibly through surface contaminated by people coughing or sneezing on them. More critically, there have been reports about the possi ...[more]

PMID: 32156101

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Project description:The emergence of 2019 novel coronavirus (2019-nCoV) is of global concern and might have emerged from RNA recombination among existing coronaviruses. CoV spike (S) protein which is crucial for receptor binding, membrane fusion via conformational changes, internalization of the virus, host tissue tropism and comprises crucial targets for vaccine development, remain largely uncharacterized. Therefore, the present study has been planned to determine the sequence variation, structural and antigenic divergence of S glycoprotein which may be helpful for the management of 2019-nCoV infection. The sequences of spike glycoprotein of 2019-nCoV and SARS coronavirus (SARS-CoV) were used for the comparison. The sequence variations were determined using EMBOSS Needle pairwise sequence alignment tools. The variation in glycosylation sites was predicted by NetNGlyc 1.0 and validated by N-GlyDE server. Antigenicity was predicted by NetCTL 1.2 and validated by IEDB Analysis Resource server. The structural divergence was determined by using SuperPose Version 1.0 based on cryo-EM structure of the SARS coronavirus spike glycoprotein. Our data suggests that 2019-nCoV is newly spilled coronavirus into humans in China is closely related to SARS-CoV, which has only 12.8% of difference with SARS-CoV in S protein and has 83.9% similarity in minimal receptor-binding domain with SARS-CoV. Addition of a novel glycosylation sites were observed in 2019-nCoV. In addition, antigenic analysis proposes that great antigenic differences exist between both the viral strains, but some of the epitopes were found to be similar between both the S proteins. In spite of the variation in S protein amino acid composition, we found no significant difference in their structures. Collectively, for the first time our results exhibit the emergence of human 2019-nCoV is closely related to predecessor SARS-CoV and provide the evidence that 2019-nCoV uses various novel glycosylation sites as SARS-CoV and may have a potential to become pandemic owing its antigenic discrepancy. Further, demonstration of novel Cytotoxic T lymphocyte epitopes may impart opportunities for the development of peptide based vaccine for the prevention of 2019-nCoV.

Dataset Information

Development of a Laboratory-safe and Low-cost Detection Protocol for SARS-CoV-2 of the Coronavirus Disease 2019 (COVID-19).

Publications

Development of a Laboratory-safe and Low-cost Detection Protocol for SARS-CoV-2 of the Coronavirus Disease 2019 (COVID-19).

Similar Datasets

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