Zika Virus Infection of Human Postnatal Thymic Epithelial Cells

The human postnatal TEC line^{17 (link)} was maintained in RPMI-1640 medium - supplemented with 10% fetal calf serum, 2 g/L sodium bicarbonate, 2 g/L HEPES, 1X antimycotic solution (Sigma-Aldrich) and 5 µg/mL of plasmocin prophylactic (InvivoGen) at 37 °C, in an atmosphere containing 5% CO₂.
The ZIKV strain used in this study was RIO-U1, which was isolated in 2016 from urine of a patient from Rio de Janeiro state, Brazil (GenBank Accession number: KU926309), as described elsewhere^{49 (link)}. Viral stocks of ZIKV Rio-U1 were prepared by infecting Vero cell monolayers. The titer (PFU/ml) and the genome integrity were determined as described elsewhere^{49 (link)}. Samples of the stock were used in the TEC infection assays. Vero cell cultures were also applied to test the infectivity of virus particles derived from infected TEC.
Unless described elsewhere, growing human TEC was plated in T-25 flasks (10⁵ cells) and infected 24 h later with 0.1 or 1.0 multiplicity of infection (MOI) or with 1 mL of infected TEC-derived supernatants for 1 h at 37°C in an atmosphere containing 5% CO₂. After infection, culture medium was changed and cells were maintained in culture for 72 h. Controls (MOCK group) corresponded to cell grown under the same culture conditions but not infected by ZIKV.