Fig. 3
a Immunization protocols. Two different vaccination regimens (control, G1; experimental, G2) were followed using 7 mice per group. Two DNA recombinants were used for priming (pVAXgp, pVAXzenv, respectively), and two viral recombinants expressing the same genes were used for the boost (FPgp, FPzenv, respectively). The DNAgp and FPgp recombinants that contained the HIV-1 gag/pro genes were used as irrelevant immunogens. Each plasmid was administered in vivo by electroporation (10 + 50 µg/recombinant/mouse), and each virus was administered subcutaneously or intranasally (1 × 106 PFU/recombinant/mouse). The challenge with ZIKV was administered subcutaneously at 1 × 105 PFU/mouse. The mice were bled before each immunization, just before the ZIKV challenge (T6) and at further times after the challenge. b Analysis of the humoral immune response. The anti-Env antibody response was determined by ELISA, where Vero cells were infected with FPzenv and then lysed, as the plate-bound antigen. Serum was obtained from all of the mice at different times before each immunization, as well as before and after the ZIKV challenge. Each line represents an individual animal. Total IgG ELISA titres are shown. An anti-ZIKV Env-specific binding antibody response was seen soon after vaccination (G2, T4). It can be noted that at 10 weeks postvaccination, after boosting the animals by the intranasal route, the antibody titer was significantly higher as compared to the control mice (G2 vs G1, T5; AUC, p < 0.05). OD450 is expressed after subtraction of the T0 values for each mouse. c Neutralizing activity using 1:50 serum dilution. Viral neutralization activity was determined using for each animal the pre-immune serum (T0) and sera from bleedings after the last immunization (T6). No inhibition of viral infectivity was found. Plaque numbers did not decrease when using hyper-immune or pre-immune sera (T6 vs T0) in the experimental vs the control animals (G2 vs G1)