Moreover, the single mutation P246A in EV71-C4 and EV71-2242 (KE strains) rendered them E151 resistant
Moreover, the single mutation P246A in EV71-C4 and EV71-2242 (KE strains) rendered them E151 resistant. was Kaempferol-3-rutinoside 1,870?M. Food azo dyes, including Kaempferol-3-rutinoside E151, interacted with the vertex of the 5-fold axis of EV71 and prevented viral entry. Their efficacy in viral inhibition was regulated by amino acids Kif2c at VP1-98, VP1-145, and/or VP1-246. Dye E151 not only prevented EV71 attachment but also eluted attached viruses in a concentration-dependent manner. Moreover, E151 inhibited the interaction between EV71 and its cellular uncoating factor cyclophilin A. studies demonstrated that E151 at a dose of 200?mg/kg of body weight/day given on the initial 4 days of challenge protected AG129 mice challenged with 10 the 50% lethal dose of wild-type EV71 isolates. Taken together, these data highlight E151 as a promising antiviral agent against EV71 infection. IMPORTANCE Human enterovirus 71 (EV71) is one of the causative agents of hand, foot, and mouth disease in children and is responsible for thousands of deaths in the past 20?years. Food azo dyes have been widely used since the nineteenth century; however, their biological effects on humans and microbes residing in humans are poorly understood. Here, we discovered that one of these dyes, brilliant black BN (E151), was particularly effective in inhibiting the infectivity of EV71 in both cell culture and mouse model studies. Mechanistic studies demonstrated that these sulfonated dyes mainly competed with EV71 attachment factors for viral binding to block viral attachment/entry to host cells. As no commercial antiviral drugs against EV71 are currently available, our findings open an avenue to exploit the development of permitted food dye E151 as a potential anti-EV71 agent. in the family (Fig. 1). Open in a separate window FIG 1 Effects of food dyes on infection of RD cells by EV71-GFP. EV71-GFP viruses were incubated with different dyes at 300 M in cell culture medium at 37C for 1?h and then inoculated into RD cells in 96-well plates at a multiplicity of infection (MOI) of 1 1. The GFP signals were observed and recorded at 24?h postinfection. Antiviral activity of dyes E122, E123, E129, and E151 values of 0.01 (one-way ANOVA with Dunnetts posttest in panel A and two-tailed Students test in panel B). (C) Inhibitory effect of E151 on the virus progeny yields of 5 enteroviruses. RD cells were infected with the virus at an MOI of 0.1 in the presence of Kaempferol-3-rutinoside various concentrations of E151 for 48 h. The titers of the respective virus progeny were determined in RD cells and are presented as percentages of the titer without E151 treatment. The average values standard errors of triplicated experiments are shown. The inhibitory effects of the dye E151 on human enteroviruses varied. PSGL-1 binding EV71 strains with VP1-98E,145G/Q, such as EV71-B2, EV71-B4, EV71-B5, EV71-C2, EV71-C5, and EV71-202 (Fig. 4A), were highly sensitive to E151, with titer reductions of 105, while non-PSGL-1 binding strains with VP1-98K,145E, such as EV71-C1, EV71-C4, EV71-252, and EV71-2242 (Fig. 4A), were slightly sensitive to E151, with titer reductions of 102 in the presence of 100?M E151 (Fig. 3A and ?andB).B). In order to determine whether the amino acids at VP1-98 and/or VP1-145 affect the sensitivity of EV71 to E151 and the other 3 dyes, variants of EV71-B5 Kaempferol-3-rutinoside and EV71-C4 with different Kaempferol-3-rutinoside amino acid combinations at VP1-98 and VP1-145 were generated using a reverse genetics (RG) system (34). In the titer reduction assay, the EE variants (VP1-98E,145E) of RG/EV71-B5 and RG/EV71-C4 were slightly sensitive to E122, E123, and E151 but resistant to E129. The KE variants were only slightly sensitive to E151 and resistant to E122 and E123. The EG and EQ variants were highly sensitive to all 4 dyes, while the KG and KQ variants were highly sensitive only.