Fig. 3. IBIS vaccination induces a potent antibody response and protects mice against lethal SARS‑CoV-2 infection.

a Schematic timeline of vaccination and viral challenge. Six to ten week-old male K18-hACE2 transgenic mice were vaccinated with 2 doses of either intranasal IBIS vaccine (1×10⁶ PFU/dose; n = 12) or with intramuscular BioNTech (BNT) BNT162b2 mRNA vaccine (1 µg/dose; n = 6) on day −28 and day −14. Mouse sera were collected at 14 days after each dose of vaccination. Neutralizing antibody titer of the sera was determined by focus-reduction neutralization test with 50% reduction cut-off (FRNT₅₀) or 75% cut-off (FRNT₇₅) against authentic SARS-CoV‑2 (ancestral) virus (b). The titer of IgG antibodies specific to SARS-CoV‑2 spike-receptor binding domain (RBD) and nucleoprotein (N) was determined by ELISA (c, d); n = 6 per group. Center line, median; box limits, upper and lower quartiles; whiskers, minima and maxima. On day 0, mock- and IBIS‑vaccinated mice were lethally challenged by intranasal inoculation of SARS‑CoV‑2 virus (ancestral) (1×10³ PFU). Body weight change (e) and survival (f) of the infected mice were monitored for 14 days. Mock‑infected, n = 3; Mock‑vaccinated/SARS‑CoV‑2 infected, n = 7; IBIS‑vaccinated/SARS‑CoV‑2 infected, n = 6. Lungs of mock‑vaccinated mice (n = 4) and IBIS‑vaccinated mice (n = 5) were harvested at day 2 post‑infection to determine the lung viral load by plaque assay (g). A group of mice were also vaccinated with 1×10⁶ PFU of IBIS and then sacrificed at day 2 post‑vaccination to confirm the absence of IBIS productive infection in vivo (n = 3). Data are presented as mean ± standard deviation (SD). PFU/g, plaque‑forming unit per gram of tissue; N.D., not detected. h–i Histology of infected mouse lungs. Mock- or IBIS‑vaccinated mice challenged by SARS‑CoV‑2 (ancestral) (1×10³ PFU) were sacrificed at 2 days post-infection. Lungs of the infected mice were harvested and PFA‑fixed for histological examination by H&E staining (upper panels) and immunofluorescence (IF) staining for SARS‑CoV‑2 N (green). Nuclei were counterstained by Hoechst 33258 (blue). PBS-vaccinated, n = 4; IBIS-vaccinated, n = 5. White scale bar=0.2 cm. Two-sided non-parametric Mann‑Whitney test was used to calculate statistical significance. ***, p < 0.001. Source data are provided as a Source Data file.