Fig. 4.

Immunization with pZIKV-3′UTR-∆20 protects male mice from ZIKV-induced damages to testis. (a) Experimental design. The bottom panel shows three mouse groups (I, II, and III) with different immunizing agents and challenge conditions. Six-week-old male A129 mice were vaccinated with pZIKV-3′UTR-Δ20 (1 μg) or DPBS (sham) using TriGrid™. On day 29 post-immunization, the mice were challenged with 10⁶ PFU of epidemic ZIKV strain PRVABC59 or DPBS controls. (b) Neutralizing antibody titers on day 29 before challenge and on day 14 post-challenge (equivalent to day 43 post-immunization). Paired t-test was performed to indicate no significant difference (n.s.) between the pre-challenge (day 29) and post-challenge (day 43) neutralizing antibody titers. (c) Viremia on day 2 post-challenge (equivalent to day 31 post-immunization). On day 21 post-challenge, the mice were sacrificed to determine viral loads in brain (d), spleen (e), and testis (f) using a focus-forming assay. (g) Testis weight on day 21 post-challenge. (h) Representative images of testes from each group collected on day 21 post-challenge. The epididymis was harvested for total sperm counts (i) and motile sperm counts (j). Individual mice are indicated by different colors and symbols. A one-way ANOVA test was performed to determine statistically significant differences among groups.