Figure 3. PRRSV RNA and protein synthesis are strongly inhibited by Cryptoporus volvatus extract.

(A) Cryptoporus volvatus extract inhibits PRRSV RNA synthesis. Marc-145 cells were infected with PRRSV Ch1a at an MOI of 0.01. Twenty-four hours post infection, cells were then treated with various concentrations of the extract or IFN-α (10 units/µl). Whole-cell RNA was isolated from the cells at 6, 12, 24, 48, or 72 hours after treatment and analyzed for PRRSV RNA using a quantitative real-time RT-PCR assay. (B) PRRSV protein synthesis is inhibited by Cryptoporus volvatus extract. Marc-145 cells were infected with PRRSV Ch1a (MOI = 5) and then treated with Cryptoporus volvatus extract at 1 mg/ml or 3 mg/ml. Thirty-six hours later, whole-cell extracts were prepared for western blot analysis, and PRRSV protein was analyzed using anti-PRRSV N monoclonal antibody (SDOW17). (C) Cryptoporus volvatus extract directly inhibits the PRRSV RNA dependent RNA polymerase activity. PRRSV nsp9/RdRp was incubated with Cryptoporus volvatus extract at a final concentration of 0, 0.05, 0.25, or 0.5 mg/ml, and the activity of the PRRSV nsp9/RdRp was examined using filter-binding assays to monitor incorporation of [³²P]ATP by using a poly(U)₁₈ RNA template. (D) Cordycepin inhibits PRRSV RdRp activity but IFN-α not. Similar experiment as C was done with IFN-α, Cryptoporus volvatus extract, and Cordycepin at final concentrations of 10 units/µl, 0.5 mg/ml, 1 mM, respectively. Data are representative of three independent experiments (mean ± SD). Statistical significance was analyzed by Student’s t test; *P<0.05; **P<0.01; ***P<0.001.