Fig. 3. ZIKV intra-anal inoculation leads to systemic infection in mice.

a Illustration of the experiment. 3–4-week-old male Ifnar1^−/− mice were infected with ZIKV (10⁶ PFU/mouse) by the i.a. route (a–f). b The survival rate of the infected Ifnar1^−/− mice (n = 6 per group). **p < 0.01, log-rank (Mantel–Cox) test. c Viral RNA copies in sera were measured at the indicated time points by qRT-PCR assay (n = 5 per time point). d ZIKV RNA recovered from the brain, lung, liver, kidney, spleen, testes, small intestine, and rectum were quantified by qRT-PCR (n = 4 per group). e The size and weight of the testes from uninfected or ZIKV anally-infected Ifnar1^−/− mice were measured at 15 dpi (n = 6, mock; n = 8, ZIKV). Scale bar = 0.5 cm. ****p < 0.0001, unpaired Student’s t-test. f RNA in situ hybridization of organ tissues from mock-infected or ZIKV anally-infected Ifnar1^−/− mice with a ZIKV-specific probe at 5 dpi. Scale bar represents 100 μm. The data in (f) are representative of two independent experiments with at least two animals per group