Dose response of ZIKVLPs in AG129 mice.

<p>5AB: PRNT₅₀ values (+/- SD) of serum samples taken from AG129 mice administered a prime and boost of 0.45 μg (5A) or a prime only of 3.0 μg (5B) ZIKVLPs pre and post challenge. 5C: Survival of 11 week old AG129 after ID challenge with 200 PFU ZIKV over a 14 day period.</p

ZIKVLP serum transfer to naïve AG129 mice.

<p>4A: Average weight loss (+/- SD) of 8 week AG129 transferred serum from mice vaccinated with ZIKVLPs after ID challenge with 20 PFU of ZIKV over a 14 day period. 4B: Survival of AG129 after challenge with ZIKV over a 14 day period.</p

Protection of ZIKVLPs in AG129 mice.

<p>3A: Neutralizing antibody titers (+/- SD) of vaccinated AG129 mice pre boost and pre challenge. 3B: Average weight loss (+/- SD) of AG129 after ID challenge with 200 PFU ZIKV over a 14 day period. 3C: Survival of 11 week old AG129 after ID challenge with 200 PFU ZIKV over a 14 day period. 3D: Viremia (+/- SD) in serum samples from mice two days post challenge by qRT-PCR. Values are total RNA copies per reaction. 3E. Viremia (+/- SD) in serum samples from mice two days post challenge by TCID₅₀. 3F: PRNT₅₀ values (+/- SD) of serum samples taken from ZIKVLP vaccinated AG129 mice post challenge, and pre challenge serum from PBS/alum mice.</p

LD50 of ZIKV in AG129 mice.

<p>Survival of AG129 after ZIKV over a 14 day period.</p

Protection of ZIKVLPs in BALB/c mice.

<p>6A: PRNT₅₀ values (+/- SD) of serum samples taken from BALB/c mice administered a prime only of 3.0 μg ZIKVLPs post challenge. 6B: Viremia (+/- SD) in serum samples from mice two days post challenge by qRT-PCR. Values are total RNA copies per reaction. 6C: Average weight loss (+/- SD) of BALB/c mice after ID challenge with 200 PFU ZIKV over a 14 day period.</p

<i>In vitro</i> characterization of Zika virus like particles.

<p>1A: Schematic of pCMV-prM/E expression cassette. 1B: Western blot analysis of Zika virus like particles. Lanes are, 1) Bio-rad precision plus kaleidoscope protein standards. 2): pCMV-prM/E transfection pre sucrose cushion purification supe. 3) 3.5x10⁴ PFU ZIKV positive control. 4) pCMV-prM/E transfection post sucrose cushion purification pt. 5) pCMV-GFP transfection post sucrose cushion purification pt. 1C-E: Sucrose cushion purified Zika VLPs observed using transmission electron microscopy. 1C: VLPs stained with Tungsten. Diameter is indicated. Background protein staining also apparent. 1D: VLP stained with Tungsten. Membrane proteins visible on the surface of VLP are indicated with arrow. Background protein staining apparent. 1E: VLP stained with Uranyl acetate. Membrane proteins visible on the surface of VLP are indicated with an arrow.</p

Mice have high serum viremia early during infection.

<p>(<b>A</b>) Viral titers in the blood of young vs. adult mice receiving 10⁵ PFU of ZIKV at days two, four, and six PI. (<b>B</b>) Viral titers in the blood of mice receiving different doses of ZIKV at days two, four, and six PI. Symbols indicate the mean value between individual mice (n = 3 for young mice and n = 6 for adult mice) ± standard deviation.</p

ZIKV causes mortality and morbidity in AG129 mice.

<p>Kaplan-Meier curves illustrate the susceptibility of AG129 mice to ZIKV. Young mice were inoculated f.p. with several doses (n = 3 for doses 10⁵−10³ PFU and n = 6 for doses 10 and 1 PFU; 10² PFU was done as two separate independent replicates with n = 3 for each) of ZIKV (<b>A</b>). All mice challenged with 10⁴−1 PFU succumbed 8 d PI. Young (n = 4) and adult mice (n = 6) were inoculated f.p. with 10⁵ PFU of ZIKV (<b>B</b>). Mice were monitored until day 10 PI. Changes in weight were calculated daily for ZIKV- and mock-infected mice (<b>C</b>). Error bars represent standard error of the mean.</p

Comparative histological imaging of skeletal muscle and brain after mock infection and infection with ZIKV.

<p>Musculature from the posterior rear limb of a ZIKV-infected mouse revealing nuclear rowing as well as degenerate muscle fibers and infiltrating inflammatory cells (<b>A</b>). Hippocampal section from a ZIKV-infected mouse revealing neutrophilic infiltration (<b>B</b>). Musculature from the same site in a mock-infected mouse (<b>C</b>) A section of hippocampus in the brain of mock-infected mouse (<b>D</b>). Scale bar, 20 μm. Data are representative of two independent experiments (n = 4 and 5).</p

Brain histopathology after ZIKV infection.

<p>Cortical tissue revealing meningeal infiltration by a mixture of neutrophils and mononuclear cells. A small amount of a similar infiltrate is seen surrounding an adjacent vessel (<b>A</b>). Cerebral neuropil section will cellular pyknosis, scattered neutrophils, and perivascular neutrophilic infiltration (<b>B</b>). Apparent necrosis and neutrophil invasion of primordial germ cell region (<b>C</b>). Scale bar, 20 μm. Data are representative of two independent experiments (n = 4 and 5).</p