Antiviral activity in vitro of ribavirin and eicar against Betanodavirus

The aim of this study was to evaluate the antiviral activity and the citotoxicity of ribavirin (RBV) and eicar, a broad spectrum antiviral agents against Betanodavirus. To determine the toxicity (CC50) of the compounds, medium with serial dilutions of RBV and eicar was added in SSN-1 cells in replication. After 3 days the cell number per well was counted with a Coulter Counter. To test the antiviral activity, confluent monolayers of SSN-1 were infected and after 2 hs the inoculum was removed and, medium containing serial dilution of RBV and eicar was added. After 3 days the cytopathic effect of the virus was recorded and the inhibitory concentration (IC50) and the selectivity index (SI) were calculated. In order to detect the reduction of extracellular and cell-associated virions at 24,48 and 72 hs post infection, the virus yield from supernatants and cell monolayer treated with 60, 40, 20, 5, 0 g/ml of ribavirin and 40, 20, 5, 2, 0 g/ml of eicar was quantified by real-time PCR and titration. Ribavirin and eicar resulted not toxic at the highest tested concentration (0,5 mg/m and 50g/ml respectively) The IC50 of RBV was 23 μg/ml + 10,9, with a SI>21,7. The antiviral activity of the compuonds was confirmed by the inhibition of the extracellular (48, 72 h) and the intracellular virus yield (24, 48 and 72 hs). In particular, the analysis showed that the reduction of the virus titre and the viral RNA quantification was statistically significant at the three time point for each concentration of ribavirin and eicar compared to the virus control. In conclusion the compounds proved to be active against the replication of Betanodavirus and it will be interesting to elucidate the mechanism of action of ribavirin and eicar against Betanodavirus.