Fig 1: Inactivated SARS-CoV-2 + aluminum hydroxide (iCoV2 + Alum) vaccination causes enhanced subclinical disease and type 2 inflammation during infection by a SARS-related coronavirus.iFLU = inactivated influenza virus vaccine; iCoV2 = inactivated SARS-CoV-2 vaccine; Alum = aluminum hydroxide adjuvant; RIBI = Sigma Adjuvant System adjuvant; DPI = days post-infection; IC80 = 80% serum reciprocal inhibitory dilution titer; pfu = plaque-forming units; EPX = Eosinophil peroxidase; WBP = whole body plethysmography; SEM = standard error of the mean; SD = standard deviation; LOD = limit of detection; PenH = Enhanced Pause; Rpef = Rate of Peak Expiratory Flow; TP = Time of PauseVaccinated mice were challenged with Rs-SHC014-CoV (SHC014) at either 4 weeks (a-g) or 10.5 months (h) post-boost. Pulmonary function was measured 1–4 DPI using WBP and pulmonary tissue was collected from vaccinated mice at 2 and 5 DPI (results displayed for 5 DPI if not specified).(a) Post-boost serum samples were collected prior to challenge and neutralizing antibody titers against SHC014 were measured using a luminescence-based microneutralization assay. Log-transformed results reported as IC80. Combined results from 4 independent replicates. (b) Pulmonary function of vaccinated mice was measured by whole-body plethysmography before (baseline) and during SHC014 infection. PenH, Rpef, and TP were measured 1–4 DPI. (c) Pulmonary viral titers at 2 and 5 DPI were quantified by plaque assay and log-transformed results reported as log10 pfu. (d, e) H&E-stained pulmonary specimens were analyzed by a blinded pathologist to evaluate ALI (d) and DAD (e). (f, g) Pulmonary eosinophils (EPX+ cells) at 5 DPI were measured using immunohistochemical staining (representative micrographs, 100X magnification in f) and quantified (g). (h) Pulmonary viral titers, ALI and DAD were measured in samples collected at 5 DPI following challenge at 10.5 months post-boost vaccination.(a, c-e, g, h) Individual data points represent biological replicates. Results analyzed by Kruskal-Wallis test with Dunn’s multiple comparisons correction (alpha = 0.05). Solid horizontal lines overlaying data represent group means. Error bars represent group SD. Dotted lines represent assay LOD. Solid horizontal lines above data represent pairwise comparisons with P values. Results from 1 (c, h), 3 (d, e, and g), or 4 (a) independent replicates (if multiple replicates, effects reproduced in each replicate).(b) Combined results from 2 independent replicates Reported as group mean ± SEM. Analyzed by two-way ANOVA with Tukey’s multiple comparisons (alpha = 0.05). Asterisks (*) denote P values (pairwise comparison between proximate experimental value and control iFLU + Alum value) using the following scheme: (*) = 0.01 – 0.05, (**) = 0.001 – 0.01, (***) 0.0001 – 0.001, (****) < 0.0001. Number of animals per group: iFLU + Alum (12), iCoV2 + Alum (24), iCoV2 + RIBI (12).