Molecular and epidemiological aspects of dengue virus infection in Hong Kong.

摘要

Dengue has emerged as a global public health concern and the incidence has increased dramatically in recent decades. In the present study, the molecular and epidemiological aspects of dengue virus infection were investigated.;Two diagnostic tools, a molecular and a serological assay for dengue virus (DV), have been developed. For the molecular assay, a one-step reverse transcription-polymerase chain reaction (RT-PCR) LightCycler assay was developed for simultaneous detection and typing of DV in 2 hours. The assay's strategy for detection was based on colour and melting temperature (Tm) multiplexing. By using in-house RT-PCR cocktail replacement, an economical in-house assay was modified based on the same detection strategy. Tm profiles and detection limits of the in-house assay were comparable to the original kit-based assay. This in-house assay was validated with clinical sera.;Three recombinant fusion proteins, prM, ED3 and prM-ED3, of DEN-2 were produced in Escherichia coli for the development of a serological assay and investigation of their potential as subunit vaccine candidates. A serological assay based on the three recombinant proteins as detection antigens was set up. Performance of the assay was compared with a commercially-available ELISA kit. Interestingly, prM as capture antigen was able to detect dengue IgG-positive sera with the highest frequency. To investigate their potential as vaccine candidates, purified recombinant proteins were administrated into rabbits for polyclonal antibody production. The neutralisation potential of antisera were investigated by means of an inhibition assay to determine DEN-2 recombinant subviral particle (RSP) binding to Vero E6 cells using flow cytometry. Antiserum against prM-ED3 chimeric protein showed the strongest inhibition of RSP binding among the four tested.;Two epidemiological aspects were investigated in the present study. For vector epidemiology study, no DV was detected from Aedes mosquitoes (593) collected in the present study which were screened for DV by our in-house RT-PCR assay and a conventional nested RT-PCR. For the seroepidemiology study, the overall prevalence of DV was 1.61% among 685 subjects recruited in Hong Kong. It was observed that seropositivity was significantly associated with increased risk for subjects who were not born and did not grown up locally in Hong Kong.