Abstract
Dengue hemorrhagic fever (DHF) is caused by infection with dengue virus (DENV). Four different serotypes (DENV1-4) co-circulate in dengue endemic areas. The viral RNA genome-based reverse-transcription PCR (RT-PCR) is the most widely used method to identify DENV serotypes in patient specimens. However, the non-structural protein 1 (NS1) antigen as a biomarker for DENV serotyping is an emerging alternative method. We modified the serotyping-NS1-enzyme linked immunosorbent assay (stNS1-ELISA) from the originally established assay which had limited sensitivity overall and poor specificity for the DENV2 serotype. Here, four biotinylated serotype-specific antibodies were applied, including an entirely new design for detection of DENV2. Prediction of the infecting serotype of retrospective acute-phase plasma from dengue patients revealed 100% concordance with the standard RT-PCR method for all four serotypes and 78% overall sensitivity (156/200). The sensitivity of DENV1 NS1 detection was greatly improved (from 62% to 90%) by the addition of a DENV1/DENV3 sub-complex antibody pair. Inclusive of five antibody pairs, the stNS1-ELISA (plus) method showed an overall increased sensitivity to 85.5% (171/200). With the same clinical specimens, a commercial NS1 rapid diagnostic test (NS1-RDT) showed 72% sensitivity (147/200), significantly lower than the stNS1-ELISA (plus) performance. In conclusion, the stNS1-ELISA (plus) is an improved method for prediction of DENV serotype and for overall sensitivity. It could be an alternative assay not only for early dengue diagnosis, but also for serotype identification especially in remote resource-limited dengue endemic areas.
Highlights
Dengue virus (DENV), the cause of the mosquito-borne disease dengue hemorrhagic fever (DHF), comprises four different serotypes (DENV1, DENV2, DENV3 and DENV4) co-circulating in tropical and subtropical endemic areas worldwide
Secondary infection with a different DENV serotype is beleived to involve with severe dengue disease
We have previously established an alternative protein-based non-structural protein 1 (NS1) assay for DENV serotyping namely, a serotyping-NS1-ELISA, with the use of serotype-specific monoclonal antibodies (Mabs) to NS1 protein
Summary
Dengue virus (DENV), the cause of the mosquito-borne disease dengue hemorrhagic fever (DHF), comprises four different serotypes (DENV1, DENV2, DENV3 and DENV4) co-circulating in tropical and subtropical endemic areas worldwide. The increasing time between sequential infection was associated with greater severity of the second detected infection [5] Serotypes of dengue virus were found to be associated with disease severity under certain conditions such as immune status, geographical areas and epidemic years. A metaanalysis of relevant dengue over 15,000 cases in 31 studies over the past 50 years was found that primary infection with DENV3 and secondary infection with DENV2, DENV3 and DENV4 increased the risk of dengue severity in Southeast Asia [8]. Laboratory diagnosis of DENV infection in patient specimens is currently performed using three different approaches, i.e. detection of DENV RNA genome (by reverse-transcription polymerase chain reaction, RT-PCR), DENV non-structural protein 1 (NS1) protein (by NS1 ELISA or rapid test) and antibody responses to DENV (by anti-dengue IgM/IgG ELISA or rapid test) [11]. Realtime or quantitative RT-PCR method is more sensitive, but it is complicated and costly, and requires a special laboratory and well-trained technicians, which is not suitable for resource-poor areas where dengue is endemic
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