Fig 3.

Replication rates of BTV-1/NS3mCherry. (A to C) In vitro virus growth curves were performed in BSR (A), BFAE (B), or KC (C) cells infected with wt BTV-1 or BTV-1/NS3mCherry. Multiple time points postinfection were generated by sampling and titrating cell supernatants. The BTV-1/NS3mCherry growth curve was more similar to that of wt BTV-1 in BSR cells than in BFAE or KC cells, where the increase in BTV-1/NS3mCherry titer was slower. (D to E) The graphs display the ratio between the intracellular versus extracellular virus titers in BSR (D) or BFAE (E) cells at 24 h p.i. BTV-1/NS3mCherry ratios are significantly higher than the wt BTV-1 in BFAE (P < 0.05) but not in BSR cells. (F) To determine whether BTV-1/NS3mCherry was able to replicate in vivo, C. sonorensis females were injected intrathoracically with BTV-1/NS3mCherry (or Schneider's medium as a negative control, referred to as “mock”). The wt BTV was used in this experiment as a positive control. After 10 days of incubation, C. sonorensis were individually homogenized and titrated by dilution assays on BSR cells. As indicated by the graph, BTV-1/NS3mCherry is able to replicate in C. sonorensis.