Development of a Singleplex Real-Time Reverse Transcriptase PCR Assay for Pan-Dengue Virus Detection and Quantification
1
Issued Date
2022-06-01
Resource Type
eISSN
19994915
Scopus ID
2-s2.0-85132198623
Pubmed ID
35746742
Journal Title
Viruses
Volume
14
Issue
6
Rights Holder(s)
SCOPUS
Bibliographic Citation
Viruses Vol.14 No.6 (2022)
Suggested Citation
Songjaeng A., Thiemmeca S., Mairiang D., Punyadee N., Kongmanas K., Hansuealueang P., Tangthawornchaikul N., Duangchinda T., Mongkolsapaya J., Sriruksa K., Limpitikul W., Malasit P., Avirutnan P. Development of a Singleplex Real-Time Reverse Transcriptase PCR Assay for Pan-Dengue Virus Detection and Quantification. Viruses Vol.14 No.6 (2022). doi:10.3390/v14061271 Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/84978
Title
Development of a Singleplex Real-Time Reverse Transcriptase PCR Assay for Pan-Dengue Virus Detection and Quantification
Other Contributor(s)
Abstract
Dengue virus (DENV) infection is a significant global health problem. There are no specific therapeutics or widely available vaccines. Early diagnosis is critical for patient management. Viral RNA detection by multiplex RT-PCR using multiple pairs of primers/probes allowing the simultaneous detection of all four DENV serotypes is commonly used. However, increasing the number of primers in the RT-PCR reaction reduces the sensitivity of detection due to the increased possibility of primer dimer formation. Here, a one tube, singleplex real-time RT-PCR specific to DENV 3′-UTR was developed for the detection and quantification of pan-DENV with no cross reactivity to other flaviviruses. The sensitivity of DENV detection was as high as 96.9% in clinical specimens collected at the first day of hospitalization. Our assay provided equivalent PCR efficiency and RNA quantification among each DENV serotype. The assay’s performance was comparable with previously established real-time RT-PCR targeting coding sequences. Using both assays on the same specimens, our results indicate the presence of defective virus particles in the circulation of patients infected with all serotypes. Dual regions targeting RT-PCR enhanced the sensitivity of viral genome detection especially during the late acute phase when viremia rapidly decline and an incomplete viral genome was clinically evident.