Figure 3.
The rTS-all vaccine candidate strain is highly attenuated and shows high immunogenicity in Syrian hamsters
(A) Scheme for the evaluation of the risk of virulent reversion by their ability to replicate at lower temperatures.
(B) The number of CPE-positive wells (upper line) and that of those that changed back to the wild-type sequence (lower line).
(C) Growth dynamics of the rTS-all and rB-1 strains at different temperatures. Vero cells were infected with each strain at MOI = 0.01. Infectious viruses in the supernatants were evaluated by TCID50 in Vero cells. Symbols represent the average of three independent experiments, and error bars represent SDs. For samples below the LOD, the assay’s LOD was used to calculate the mean. The LOD is 2.65 log10 TCID50/mL, indicated in the x axis. For statistical analysis, two-way ANOVA was performed (∗p < 0.05, ∗∗∗∗p < 0.0001).
(D) Overview of the investigation of the pathogenicity and immunogenicity of the rTS-all strain (n = 5).
(E) Weight change in the recombinant viruses-infected hamsters. The average weight change is plotted, and error bars represent the SDs. For statistical analysis, two-way ANOVA was performed (∗∗∗∗p < 0.0001).
(F and G) The viral titers in lung homogenates (F) and nasal wash specimens (G) three days post-infection. Viral titration was performed by plaque formation assays. Bars depict the mean values, and symbols represent individual viral titers. Error bars represent mean SDs. The LOD is indicated in the x axis. For statistical analysis, one-way ANOVA was performed (∗∗p < 0.01 and ∗∗∗p < 0.001).
(H)Neutralization titers in the sera of rB-1 or rTS-all-infected and mock-treated hamsters. Symbols represent individual neutralization titers. Bars show median values and error bars represent 95% CIs. For statistical analysis, the Kruskal-Wallis test was performed (n.s.: not significant). The LOD is indicated in the x axis.
(I)Weight change in Syrian hamsters after reinfection with the B-1 virus. The average weight change is plotted, and error bars represent mean SDs. To compare weight change on the same day between the naïve and the other groups, two-way ANOVA was performed (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗∗p < 0.0001).