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Am J Trop Med Hyg. 2019 Jun;100(6):1534-1540. doi: 10.4269/ajtmh.19-0138.

A Direct from Blood/Plasma Reverse Transcription-Polymerase Chain Reaction for Dengue Virus Detection in Point-of-Care Settings.

Author information

1
School of Public Health, University of Alberta, Edmonton, Canada.
2
Department of Cell Biology, University of Alberta, Edmonton, Canada.
3
Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Canada.
4
Section of Pediatric Infectious Diseases, Department of Pediatrics, Chong Hua Hospital, Cebu, Philippines.
5
Cebu Institute of Medicine, Cebu, Philippines.
6
Lebumfacil-Santa Ana Medical Center, Cebu, Philippines.
7
Tropical Disease Unit, The University Health Network-Toronto General Hospital, University of Toronto, Toronto, Canada.
8
Women and Children's Health Research Institute, Edmonton, Canada.
9
Stollery Science Lab, Edmonton, Canada.
10
Department of Pediatrics, University of Alberta, Edmonton, Canada.

Abstract

Infection with dengue virus (DENV) is widespread across tropical regions and can result in severe disease. Early diagnosis is important both for patient management and to differentiate infections that present with similar symptoms, such as malaria, chikungunya, and Zika. Rapid diagnostic tests that are used presently for point-of-care detection of DENV antigens lack the sensitivity of molecular diagnostics that detect viral RNA. However, no molecular diagnostic test for DENV is available for use in field settings. In this study, we developed and validated a reverse transcription-polymerase chain reaction (RT-PCR) for the detection of DENV adapted for use in field settings. Reverse transcription-polymerase chain reaction was performed directly from plasma samples without RNA extraction. The assay detected all four serotypes of DENV spiked into blood or plasma. Our RT-PCR does not cross-react with pathogens that cause symptoms that overlap with dengue infection. The test performed equally well in a conventional laboratory qPCR instrument and a small, low-cost portable instrument that can be used in a field setting. The lower limit of detection for the assay was 1 × 104 genome copy equivalents/mL in blood. Finally, we validated our test using 126 archived patient samples. The sensitivity of our RT-PCR was 76.7% (95% CI: 65.8-87.9%) on the conventional instrument, and 78.3% (95% CI: 65.8-87.9%) on the field instrument, when compared with the RealStar Dengue RT-PCR Kit 2.0. The molecular test described here is user-friendly, low-cost, and can be used in regions with limited laboratory capabilities.

PMID:
30994095
PMCID:
PMC6553918
[Available on 2020-06-01]
DOI:
10.4269/ajtmh.19-0138

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