AG129 mice59 were bred in a specific-pathogen-free facility at Institut Pasteur’s animal facility, and C57BL/6 mice were obtained from Charles River Laboratories. Infected adult mice were housed in BSL-3 level isolators and handled in compliance with the Animal Committee regulations and guidelines of Institut Pasteur Paris France, under the 2010/63 European Union Council directive. Animal protocols were approved by the Ethics Committee on Animal Experimentation (CETEA) under dossier number dap160116/CHCST 18.176 and the USAMRMC Animal Care and Use Review Office (ACURO), under the protocol number DARPA-5417.04. Mice were monitored daily, with food and water supplied ad libitum. Endpoints were defined and mice humanely euthanized when these were reached.

4-6 week old AG129 or C57BL/6 female mice were given a mix of Ketamine(10 mg/mL)/Xylazine (1 mg/mL) by intraperitoneal injection. Once anesthetized, mice were inoculated by a subcutaneous (footpad) route with vehicle (DMEM media), 104 PFU of Zika virus alone, or complemented with DVG-containing VLPs (TIPs). C57BL/6 mice were treated with 2 mg of an IFNAR1 blocking mouse MAb (MAR-5A3, Euromedex) by intraperitoneal injection one day prior to infection. Weight loss and viremia were monitored at the indicated times after infection. Blood was collected from the facial vein in Microtainer blood collection tubes (BD) and allowed to clot at room temperature. Serum was separated by centrifugation and stored at −80 °C. Viremia was quantified either by plaque assay or RT-qPCR on unextracted and diluted serum samples, as described previously60. Infected mice were euthanized by cervical dislocation. Spleen, ovaries, brain, and injected footpads were harvested and homogenized with 600 μl DMEM supplemented with 2% FBS in Precellys tubes containing ceramic beads, using a Precellys 24 homogenizer (Bertin Technologies) at 5000 rpm for 2 cycles of 20 seconds. Homogenates were cleared by centrifugation, and the supernatant stored at −80 °C until virus titration or RT-qPCR. Viral burden in organs was measured either by plaque assay (expressed as PFU/g for organs) or by RT-qPCR (expressed as PFU equivalents relative to GAPDH) following RNA extraction using Direct-zol-96 (Zymo). The number of DVG or WT RNA genomes in mouse organs was normalized to GAPDH genomes, derived from a standard curve using mouse GAPDH primers (Supplementary Table 3) and RNA extracted from the respective organ of uninfected mice.

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