Fig. 3: A. muciniphila-mediated neuroinflammation is associated with increased caecal concentrations of γ-amino butyric acid.

a–d, GF C57BL/6N mice were colonized with A. muciniphila only (SM01), SM13, SM14 or remained GF. Caecal contents were collected 25 d after colonization (−EAE) or 30 d after EAE induction (+EAE) and subjected to CE–TOF/MS-based metabolomics analysis. a, Principal component analysis (PCA) of log₂-normalized metabolite concentrations, faceted by colonization. b, Ward hierarchical clustering based on scaled group means of log₂-normalized metabolite concentrations. c, Statistically significant positive (PCor) or negative (NCor) Spearman correlations by linear regression across all samples (both −EAE and +EAE mice) and groupwise comparison criteria. Correlations referring to EAE-associated readouts (abbreviations as in Fig. 1) were calculated from +EAE mice only. Groupwise comparisons include significantly different metabolites based on unpaired t-tests of log₂-normalized concentrations with P-value adjustment using the Benjamini–Hochberg method. Barplots indicate the total number of metabolites that fulfil each criterion. Of 175 measured metabolites, only the 14 metabolites that demonstrate a significant correlation with AUC are displayed. Grey squares indicate that a given metabolite fulfilled a specific criterion, while white squares indicate failure to fulfil a given criterion. d, Boxplots showing median, quartiles and 1.5 × IQR of log₂-concentrations of GABA −EAE or +EAE conditions. One-way ANOVA followed by Tukey’s post-hoc test. e, Multidimensional reduction of caecal metatranscriptome profiles of −EAE SM14- and SM13-colonized mice. In SM14-colonized mice, transcripts attributed to A. muciniphila were removed and counts were renormalized to allow for fair comparison with metatranscriptome profiles of SM13-colonized mice. Two dots from SM13 −EAE overlap in the plot. f, Volcano plot showing log₂(fold change, FC) of gene product-annotated transcript abundances in SM14- vs SM13-colonized mice (x axis) and −log₁₀(P value) (y axis), calculated using an exact test in edgeR. Dashed line represents significance threshold. Yellow and blue dots represent transcripts found only in SM14- or SM13-colonized mice, respectively, while grey dots represent transcripts found in both groups. g, Two left columns: transcripts found only in SM14-colonized or SM13-colonized mice. Two right columns: transcripts up- or downregulated in SM14- vs SM13-colonized mice but present in both groups. Mouse numbers are indicated on the respective panels and treated as biological replicates. All statistical tests were two-sided.

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