FIG 4.

Characterization of the recombinant DENV-1 I64V mutant. (A) BHK-21 cells were transfected with 10 μg of WT or NS4B mutant genome-length RNA of DENV-1 (West Pacific strain). The transfected cells were monitored for viral E protein expression by IFA at days 2 and 4 posttransfection. (Left) Anti-E monoclonal antibody 4G2 and Alexa Fluor 488 goat anti-mouse IgG were used as the primary and secondary antibodies, respectively. (Middle) Plaque morphology of WT DENV-1 and the I64V mutant. (Right) Viral titers in the culture fluids at day 5 p.t. were determined by a plaque assay. (B) Conversion of DENV-1 to sensitivity to compound 1a inhibition by an I64V mutation. Recombinant DENV-1 containing the I64V mutation was generated. The effect of compound 1a on the DENV-1 I64V mutant was determined by a viral titer reduction assay in A549 cells. The logarithm (log₁₀) values of average viral titers from triplicates are plotted against the compound 1a concentration. Average results and standard deviations (n = 3) are presented.