Fig. 4.

Isolation and characterization of A/WSN/33 (H1N1) variants with reduced drug susceptibility. A Schematic representation of serial passage of the viruses for selecting ZX-7101-resistant virus. MDCK cells were initially infected with A/WSN/33 (H1N1) virus at an MOI of 0.1 and cultured with ZX-7101 at a concentration of 0.1 ​nmol/L for 3 days. The supernatants were diluted 10-fold and sequentially passaged through fresh MDCK cells in the presence of 0.1 ​nmol/L ZX-7101. B, C Susceptibility of variants to ZX-7101 and BXA by CellTiter-Glo® Luminescent Cell Viability Assay. MDCK cells were infected with 100 TCID₅₀ of H1N1 and H1N1-P15 variants. Then, different concentrations of ZX-7101 or BXA were added for coculture at 37 ​°C for 48 ​h. The inhibition potencies of the compounds were determined by CellTiter-Glo® Luminescent Cell Viability Assay. The results are presented as the mean of at least three replicates ​± ​SD. D Susceptibility of variants to ZX-7101 and BXA by TCID₅₀ assay. MDCK cells infected with H1N1 or the H1N1-P15 variant at an MOI of 0.01 were treated with ZX-7101 (1 ​nmol/L) or BXA (1 nmol/L). After 48 ​h, supernatants were collected for TCID₅₀ assay in MDCK cells. The results are presented as the mean of three replicates ± SD. The black dots show the values from each experiment. ∗∗∗P ​< ​0.001. One-way ANOVA was used for virus titer.