Journal
PATHOGENS
Volume 10, Issue 12, Pages -Publisher
MDPI
DOI: 10.3390/pathogens10121558
Keywords
SARS-CoV-2; DENV; virus co-circulation; direct RT-qPCR; virus inactivation; biosafety
Categories
Funding
- Japan Agency for Medical Research and Development (AMED) [JP21wm0125006, JP21wm0225018]
- Research on Emerging and Re-emerging Infectious Diseases [21fk0108109h0003, 21fk0108123h1102]
- Nagasaki University
- [LC522975.1]
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This study demonstrates that direct real-time RT-qPCR is a feasible option for the detection of SARS-CoV-2 and DENV-2, compared to traditional real-time RT-qPCR based on viral genome extraction methods. This approach can provide faster viral detection, aiding in addressing viral outbreaks in resource-limited settings.
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.
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