Fig. 3: Optimal mucosal efficacy of STm^Aux is viability dependent.

a Germ-free mice were enterally conditioned with four doses of 10¹⁰ CFU STm^Aux (filled red triangles, n = 10 animals), PAA-killed STm^Aux (filled blue diamonds, n = 11 animals), or were left untreated (gray-filled circles, n = 10 animals). Four weeks after the first treatment (day 0) mice were challenged with wild-type STm (10⁴ CFU). Mice were sacrificed at day 1 and day 4 after challenge with wild-type STm, respectively. Each symbol represents one individual. b STm-specific titer (logEC₅₀) of intestinal secretory IgA determined by live bacterial flow cytometry. c, d Lipocalin-2 concentration in feces (c) and cecal content (d) at day 0–4 after challenge. (e) Cecal histopathology score at days 1 and 4 after challenge, respectively. Each symbol represents one individual. Data points depicted with an arrow are shown in panel f. f Cecal histology at day 1 and day 4 after challenge, respectively. H&E staining of cryosections. Scale bar = 100 μm. g Bacterial burden of wild-type STm in mLNs, spleens, and livers at days 1 and 4 after challenge, respectively. h Fecal colonization of wild-type STm at days 0–4 after challenge (inset graph: quantification of STm^Aux in feces at day 0 confirming germ-free status at day 0). i Cecal luminal colonization of wild-type STm at day 4 after challenge. Statistics: bars indicate mean (b–d, g–i) or median values (e). Horizontal, dotted lines indicate the detection limit (b–d, g–i). Panel b was analyzed with an unpaired two-tailed t-test (control group excluded from test). Panels c, d, and g were analyzed with ordinary one-way ANOVA and Dunnett’s post hoc test (comparison to control group). Panel e was analyzed with a two-sided multicomparisons Kruskal–Wallis test and Dunn’s post hoc test. Source data are provided as a Source Data file. Detailed statistical metrics are available in the Supplementary Statistical Analysis file.