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Direct Viral RNA Detection of SARS-CoV-2 and DENV in Inactivated Samples by Real-Time RT-qPCR: Implications for Diagnosis in Resource Limited Settings with Flavivirus Co-Circulation
The RT-qPCR method remains the gold standard and first-line diagnostic method for the detection of SARS-CoV-2 and flaviviruses, especially in the early stage of viral infection. Rapid and accurate viral detection is a starting point in the containment of the COVID-19 pandemic and flavivirus outbreak...
Autores principales: | , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8705679/ https://www.ncbi.nlm.nih.gov/pubmed/34959513 http://dx.doi.org/10.3390/pathogens10121558 |
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author | Mao, Zhan Qiu Fukuta, Mizuki Balingit, Jean Claude Nguyen, Thi Thanh Ngan Nguyen, Co Thach Inoue, Shingo Nguyen, Thi Thu Thuy Nguyen, Le Khanh Hang Minakawa, Noboru Morita, Kouichi Le, Thi Quynh Mai Hasebe, Futoshi Moi, Meng Ling |
author_facet | Mao, Zhan Qiu Fukuta, Mizuki Balingit, Jean Claude Nguyen, Thi Thanh Ngan Nguyen, Co Thach Inoue, Shingo Nguyen, Thi Thu Thuy Nguyen, Le Khanh Hang Minakawa, Noboru Morita, Kouichi Le, Thi Quynh Mai Hasebe, Futoshi Moi, Meng Ling |
author_sort | Mao, Zhan Qiu |
collection | PubMed |
description | The RT-qPCR method remains the gold standard and first-line diagnostic method for the detection of SARS-CoV-2 and flaviviruses, especially in the early stage of viral infection. Rapid and accurate viral detection is a starting point in the containment of the COVID-19 pandemic and flavivirus outbreaks. However, the shortage of diagnostic reagents and supplies, especially in resource-limited countries that experience co-circulation of SARS-CoV-2 and flaviviruses, are limitations that may result in lesser availability of RT-qPCR-based diagnostic tests. In this study, the utility of RNA-free extraction methods was assessed for the direct detection of SARS-CoV-2 and DENV-2 in heat-inactivated or chemical-inactivated samples. The findings demonstrate that direct real-time RT-qPCR is a feasible option in comparison to conventional real-time RT-qPCR based on viral genome extraction-based methods. The utility of heat-inactivation and direct real-time RT-qPCR for SARS-CoV-2, DENV-2 viral RNA detection was demonstrated by using clinical samples of SARS-CoV-2 and DENV-2 and spiked cell culture samples of SARS-CoV-2 and DENV-2. This study provides a simple alternative workflow for flavivirus and SARS-CoV-2 detection that includes heat inactivation and viral RNA extraction-free protocols, with aims to reduce the risk of exposure during processing of SARS-CoV-2 biological specimens and to overcome the supply-chain bottleneck, particularly in resource limited settings with flavivirus co-circulation. |
format | Online Article Text |
id | pubmed-8705679 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-87056792021-12-25 Direct Viral RNA Detection of SARS-CoV-2 and DENV in Inactivated Samples by Real-Time RT-qPCR: Implications for Diagnosis in Resource Limited Settings with Flavivirus Co-Circulation Mao, Zhan Qiu Fukuta, Mizuki Balingit, Jean Claude Nguyen, Thi Thanh Ngan Nguyen, Co Thach Inoue, Shingo Nguyen, Thi Thu Thuy Nguyen, Le Khanh Hang Minakawa, Noboru Morita, Kouichi Le, Thi Quynh Mai Hasebe, Futoshi Moi, Meng Ling Pathogens Article The RT-qPCR method remains the gold standard and first-line diagnostic method for the detection of SARS-CoV-2 and flaviviruses, especially in the early stage of viral infection. Rapid and accurate viral detection is a starting point in the containment of the COVID-19 pandemic and flavivirus outbreaks. However, the shortage of diagnostic reagents and supplies, especially in resource-limited countries that experience co-circulation of SARS-CoV-2 and flaviviruses, are limitations that may result in lesser availability of RT-qPCR-based diagnostic tests. In this study, the utility of RNA-free extraction methods was assessed for the direct detection of SARS-CoV-2 and DENV-2 in heat-inactivated or chemical-inactivated samples. The findings demonstrate that direct real-time RT-qPCR is a feasible option in comparison to conventional real-time RT-qPCR based on viral genome extraction-based methods. The utility of heat-inactivation and direct real-time RT-qPCR for SARS-CoV-2, DENV-2 viral RNA detection was demonstrated by using clinical samples of SARS-CoV-2 and DENV-2 and spiked cell culture samples of SARS-CoV-2 and DENV-2. This study provides a simple alternative workflow for flavivirus and SARS-CoV-2 detection that includes heat inactivation and viral RNA extraction-free protocols, with aims to reduce the risk of exposure during processing of SARS-CoV-2 biological specimens and to overcome the supply-chain bottleneck, particularly in resource limited settings with flavivirus co-circulation. MDPI 2021-11-29 /pmc/articles/PMC8705679/ /pubmed/34959513 http://dx.doi.org/10.3390/pathogens10121558 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Mao, Zhan Qiu Fukuta, Mizuki Balingit, Jean Claude Nguyen, Thi Thanh Ngan Nguyen, Co Thach Inoue, Shingo Nguyen, Thi Thu Thuy Nguyen, Le Khanh Hang Minakawa, Noboru Morita, Kouichi Le, Thi Quynh Mai Hasebe, Futoshi Moi, Meng Ling Direct Viral RNA Detection of SARS-CoV-2 and DENV in Inactivated Samples by Real-Time RT-qPCR: Implications for Diagnosis in Resource Limited Settings with Flavivirus Co-Circulation |
title | Direct Viral RNA Detection of SARS-CoV-2 and DENV in Inactivated Samples by Real-Time RT-qPCR: Implications for Diagnosis in Resource Limited Settings with Flavivirus Co-Circulation |
title_full | Direct Viral RNA Detection of SARS-CoV-2 and DENV in Inactivated Samples by Real-Time RT-qPCR: Implications for Diagnosis in Resource Limited Settings with Flavivirus Co-Circulation |
title_fullStr | Direct Viral RNA Detection of SARS-CoV-2 and DENV in Inactivated Samples by Real-Time RT-qPCR: Implications for Diagnosis in Resource Limited Settings with Flavivirus Co-Circulation |
title_full_unstemmed | Direct Viral RNA Detection of SARS-CoV-2 and DENV in Inactivated Samples by Real-Time RT-qPCR: Implications for Diagnosis in Resource Limited Settings with Flavivirus Co-Circulation |
title_short | Direct Viral RNA Detection of SARS-CoV-2 and DENV in Inactivated Samples by Real-Time RT-qPCR: Implications for Diagnosis in Resource Limited Settings with Flavivirus Co-Circulation |
title_sort | direct viral rna detection of sars-cov-2 and denv in inactivated samples by real-time rt-qpcr: implications for diagnosis in resource limited settings with flavivirus co-circulation |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8705679/ https://www.ncbi.nlm.nih.gov/pubmed/34959513 http://dx.doi.org/10.3390/pathogens10121558 |
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