Figure 1.
Flow chart of the experimental procedure. For the ZIKV infection experiments, 715 mosquitoes were collected in Nopporo forest park between July and September in both 2017 and 2018. All 715 mosquitoes were orally challenged with an IBM containing 105 FFU/mL or 106 FFU/mL of the ZIKV strain PRVABC59, and 158 mosquitoes ingested an IBM. A part of IBM-ingested Ae. punctor, Ae. japonicus and Ae. vexans were used for another preliminary experiments. The virus titers and the amount of virus RNA were determined by FFA and qRT-PCR, respectively. Conventional RT-PCR was used to confirm the qRT-PCR results, and the sequence of entire viral E gene of RT-PCR positive samples was determined. In another study 3170 mosquitoes were collected between April and October in 2016 to 2020 to screen the naturally infected Flaviviruses. Three hundred two pools including 10 species were screened by RT-PCR and the sequence of amplicons were determined.