Evaluation of the assays for detection of West Nile virus from mosquitoes in Japan

Abstract

In Japan, the invasion of the West Nile virus (WNV) has not been reported. However, in order to develop a rapid- and sensitive-detection system for WNV, reverse transcriptase-polymerase chain reaction (RT-PCR), TaqMan RT-PCR, and VecTest^<TM> WNV assays were evaluated. VecTest^<TM> produced results within 1 hour and no specialized training was required. However it showed the lowest sensitivity to detect WNV (> 10^5 plaque-forming units; PFU/tube) and was costly. The TaqMan RT-PCR assay was the highest sensitive assay (>0.25 PFU/tube) and was followed by the RT-PCR assay (>20 PFU/tube), although it took 10 hours for investigators to get the results of both assays and their protocols are more complex than that of the VecTest^<TM>. In addition, TaqMan RT-PCR is simpler than RT-PCR. These results suggest that the TaqMan RT-PCR assay may be the most effective for WNV survey at present in Japan. We hereby recommend one of the most preferable WNV surveillance systems at present in Japan as follows. First, the mosquito pool supernatants are screened for the presence of WN viral RNA by using the TaqMan RT-PCR assay with WNV primer-probe sets. Second, the viral RNA-positive pools are used for the virus isolation using cell culture assay. Finally, the isolated viruses are characterized for the viral genome by using molecular biological techniques.