Fig. 5: fgTiO₂-Salmonella interactions.

Mouse Study 2 used a 16-week fgTiO₂ feeding period before switching all animals to a normal diet (i.e., without fgTiO₂-supplementation) then orally inoculating half with attenuated, ΔaroA-Salmonella. Tissues were harvested +3 or +28 days after infection. a–c Z-stack maximum projections showing immunofluorescent labelling for flagellin (FliC) to detect the ΔaroA-Salmonella in the Peyer’s patch SED region. Marked accumulation of both fgTiO₂ and ΔaroA-Salmonella was observed in autofluorescent, phagocytic mononuclear cells at the +3 timepoint. d Faecal ELISA studies confirmed a Salmonella-specific IgA response at the +28 timepoint (bars represent medians with interquartile range error-bars, n = 3–6 samples per treatment group). e–g Protein expression analyses of ileal tissue digests by proximity extension assay (OLINK mouse exploratory panel). e Comparison of ileal tissues taken from wild-type (WT) mice unexposed to fgTiO₂ but with (WT +Sal) or without (WT) ΔaroA-Salmonella infection confirmed significant increases key cytokines/chemokines associated with a Th1 immune response (P ≤ 0.05, two-sided Wilcoxon rank-sum test, n = 5–6 tissues per group). f Inflammation-associated protein expression analyses of ileal tissue digests. At each timepoint, no significant differences in protein expression were observed between the WT and WT +TiO₂ (P ≥ 0.53, two-sided Wilcoxon rank-sum test, n = 3 animals per group) or WT +Sal and WT +TiO₂ +Sal groups (P ≥ 0.10, two-sided Wilcoxon rank-sum test, n = 6 animals per group). Full dataset is shown in Supplementary Fig. 12. g Peyer’s patch-focussed inflammation-associated protein expression analyses using carefully-excised patch digests in mice treated with either ΔaroA-Salmonella alone (WT +Sal) or with fgTiO₂ and ΔaroA-Salmonella (WT +TiO₂ +Sal) at the +3 or +28 timepoints. At each timepoint, no significant differences were observed between the two groups (P ≥ 0.20, two-sided Wilcoxon rank-sum test, n = 6 animals per group) demonstrating no interaction of fgTiO₂ and ΔaroA-Salmonella despite (a–c) heavy loading into the same cells (full dataset shown in Supplementary Fig. 13). OLINK protein abbreviations are defined in Supplementary Table 1. All p-values are available for download at the BioStudies database under accession number S-BSST875. Scale bars: a–c = 10 μm.