Figure 1

Antiviral activity of manassantin B against coxsackievirus B3 in vitro. (a) Vero cells were infected with coxsackievirus B3 (CVB3) at 50% cell culture infective dose (CCID₅₀) and treated with manassantin B (Man B). The cytopathic effect on virus-infected Vero cells was analysed 2 days after infection. The antiviral activity was calculated based on cell viability. (b) Replication of CVB3 in Vero cells at 48 h after infection with CVB3 in the presence of Man B (10 µg/mL) was detected by RT-PCR of the CVB3 genome. (c) Quantitative real-time PCR was conducted to measure the copy number of CVB3 viral RNA in Vero cells after treatment with Man B (2 and 10 μg/mL). (d) Western blotting was performed to determine the effect of different concentrations of Man B on the production of CVB3 VP1 protein. Vero cells (1 × 10⁶) were seeded in a 6-well plate and incubated overnight. Cells were infected with CVB3 at an MOI of 0.8 and treated with 0.08, 0.4, 2, and 10 μg/mL of Man B. After 48 h, cells were harvested. (e) To evaluate the antiviral activity of Man B at multiple time points, cells were harvested at 10, 20, and 30 h after CVB3 infection and treatment with 2 µg/mL of Man B. Thirty micrograms of total cellular proteins from CVB3-infected Vero cells was electrophoresed and subjected to western blot. α-Tubulin was used as a loading control for each set of samples. The samples were derived from the same experiment, and the gels/blots were processed in parallel. Results are shown as means ± SEM of 3 independent experiments. ^###P < 0.001 for comparison with non-infected control group (Ctrl); ***P < 0.001 for comparison with CVB3-infected vehicle group (Veh), Bonferroni’s multiple comparison test (ANOVA).