Fig 2.
A. Replication of BgV and WNVKUN in vertebrate cells. Cells were inoculated at MOI 1 with BgV or WNVKUN in triplicate for 1h 30min, the inoculum removed and the cells washed before incubating at 34°C (black), 37°C (red) or 28°C (blue) with fresh medium. The inoculated cell culture supernatants were harvested five days p.i., titrated on C6/36 cells and analysed by fixed-cell ELISA. The dotted line represents the lower limit of detection, * stands for p < 0.05. Error bars represent the standard deviation. B. Thermostability of BgV and WNVKUN at various temperatures over three days. Triplicates of 105 TCID50 IU of BgV and WNVKUN were held at 4°C, 28°C, 34°C and 37°C in culture medium with 2% FBS in the absence of cells, harvested at 30min, 24h, 48h and 72h after the start of incubation and stored at -80°C. These samples were titrated on C6/36 cells and the titres determined by fixed-cell ELISA. Error bars represent the standard deviation. C. Replication kinetics of BgV, BgV/WNV-prME, WNVKUN and WNV/BgV-prME, in BSR (left) and Vero cells (right) over five days at 34°C and 37°C. The cells were inoculated at MOI 0.1 in triplicates at 34°C or 37°C for 1h 30min, the inoculum removed and the cells washed before incubating at either temperature with fresh medium. The supernatants were harvested at 2, 24, 72 and 120 hours p.i., stored at -80°C until they were titrated on C6/36 cells and the titres determined by fixed-cell ELISA. Error bars represent the standard deviation and the dotted lines represent the lower limit of detection.