Figure 3.

Nitazoxanide acts at postentry level. (A,B) MRC-5 cells mock-infected or infected with HCoV-OC43 (A) or HCoV-229E (B) (0.5 TCID₅₀/cell) were treated with NTZ 1 μg/mL [(A,B) top; filled bars], NTZ 2.5 μg/mL [(A,B) bottom; filled bars] or vehicle (empty bars) 2 h before infection (Pre), after the adsorption period (Post), or 2 h before infection and treatment was continued during and after the adsorption period (Pre + Post). Virus yield was determined at 24 h p.i. by TCID₅₀ infectivity assay. (C) MRC-5 cells mock-infected or infected with HCoV-OC43 or HCoV-229E were treated with NTZ (1 μg/mL) or vehicle only during the adsorption period (Ads). Virus yield was determined at 24 h p.i. by infectivity assay. (A–C) Data represent the mean ± S.D. of duplicate samples. *p < 0.01; Student’s t-test. (D) Schematic representation of HCoV genomic RNA transfection assay. (E) MRC-5 cells were transfected with HCoV-OC43 or HCoV-229E genomic RNA for 4 h and treated with NTZ (2.5 μg/mL) or vehicle for 24 h. Virus yield was determined at 24 h after treatment in the supernatant of transfected cells by TCID₅₀ infectivity assay. Nucleocapsid (N) protein levels in HCoV-OC43 or HCoV-229E RNA-transfected cells are shown (insets). (F) MRC-5 cells infected with HCoV-OC43 were treated with NTZ (1 μg/mL) or vehicle at the indicated times after infection. Virus yield was determined at 24 h p.i. by infectivity assay. (E,F) Data represent the mean ± S.D. of duplicate samples. *p < 0.01; Student’s t-test.