A comparative retrospective study of RT-PCR-based liquid hybridization assay for early, definitive diagnosis of dengue

Hapugoda, Menaka D.; de Silva, Nilanthi R.; Khan, Baldip; Damsiri Dayanath, M.Y.; Gunesena, Sunethra; Prithimala, L.D.; Abeyewickreme, Wimaladharma

doi:10.1016/j.trstmh.2009.11.001

« Previous Next »Transactions of the Royal Society of Tropical Medicine and Hygiene
Volume 104, Issue 4 , Pages 279-282, April 2010

A comparative retrospective study of RT-PCR-based liquid hybridization assay for early, definitive diagnosis of dengue

Menaka D. Hapugoda
- Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, P.O. Box 06, Ragama, Sri Lanka
Nilanthi R. de Silva
- Department of Parasitology, Faculty of Medicine, University of Kelaniya, P.O. Box 06, Ragama, Sri Lanka
Baldip Khan
- Division of Human Health, International Atomic Energy Agency, P.O. Box 100, Wagramerstrasse 5, Vienna A-1400, Austria
M.Y. Damsiri Dayanath
- Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, P.O. Box 06, Ragama, Sri Lanka
Sunethra Gunesena
- Department of Virology, Medical Research Institute, Colombo 08, Sri Lanka
L.D. Prithimala
- Department of Virology, Medical Research Institute, Colombo 08, Sri Lanka
Wimaladharma Abeyewickreme
- Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, P.O. Box 06, Ragama, Sri Lanka
- Department of Parasitology, Faculty of Medicine, University of Kelaniya, P.O. Box 06, Ragama, Sri Lanka
- Corresponding author. Tel.: +94 11 2960483; fax: +94 11 2953412.

Received 21 February 2007; received in revised form 3 November 2009; accepted 3 November 2009.

Summary

Dengue is an important flaviviral infection in tropical and subtropical regions. Early diagnosis of dengue infection helps in monitoring the disease, determining when hospital admission is necessary and reducing case fatalities. The objective of this study was to carry out a retrospective comparison of an RT-PCR-based liquid hybridization (RT-PCR-LH) assay with PCR amplification, virus isolation and serological techniques for laboratory diagnosis of dengue infection. Amplified products of non-structural 3 gene were hybridized with a mixture of four dengue type-specific DNA probes in liquid phase. The assay was validated in a comparative retrospective study using acute serum samples collected from 119 fever patients with or without dengue, confirmed by haemagglutination inhibition (HAI) assay, the gold standard assay for diagnosis of dengue infection. The RT-PCR-LH assay was highly specific for dengue and, as an early laboratory diagnostic method, had 100% sensitivity in detecting dengue patients confirmed by HAI assay. A high analytical sensitivity of two fluorescent focus units of dengue virus/reaction was achieved. This RT-PCR-LH assay using a single serum specimen offers distinct advantages of specificity and sensitivity over other diagnostic techniques for early definitive laboratory diagnosis of dengue infection when serological methods are of little value.

Keywords: Dengue, Diagnosis, Clinical laboratory techniques, PCR, Liquid hybridization, Haemagglutination inhibition assay