Figure 2.
FMT experiments demonstrate the causal role of microbiome in the induction of type 2 cytokines in the proximal colon, which correlates with the enrichment of B. thetaiotaomicron in the microbiome. (A and B) IL-25, IL-5, and IL-13 concentrations were quantified in the (A) proximal colon or (B) in the ileum (for IL-25) of C57BL/6 WT mice (see Materials and methods) receiving FMTs from mice treated with vancomycin or left untreated. Prior to FMT, mice were either untreated or treated with AVNM to eliminate the resident microbiota (Background). Protein concentration in the lysate (pg/ml) was normalized by the total protein mass generated in the sample (in mg) as in Buonuomo et al. (2016). Data are representative of independent experiments repeated twice, using n = 4–6 female mice per treatment group. ANOVA with Tukey post-hoc test was run to determine significant differences. Statistical significance was estimated at an FDR of 0.05. (C) 16S rRNA sequencing was conducted on input FMT and fecal samples collected from mice after FMT. (D) Volcano plots were generated following differential analysis using DeSeq2 on fecal microbiome sequencing samples from mice treated with AVNM and receiving FMT from either vancomycin-treated or untreated mice. Amplicon SVs from two succinate-producing species (B. thetaiotaomicron and E. faecalis) were found to be enriched in AVNM-treated mice receiving FMT from vancomycin-treated animals. Significance was determined based on an FDR of 0.05. (E) Permutation importance analysis followed by accumulated local effects calculations was carried out on the results of RFR modeling, which aimed to predict colonic IL-25 concentrations based on microbiome species abundance. Sequence variants associated with B. thetaiotaomicron and E. faecalis were identified as positive predictors of IL-25 concentrations in the colon. (F) The significance and directionality inferred by the RFR model were confirmed through Elastic Net Regression modeling and Bayesian Variable Selection Linear Regression. SVs linked to B. thetaiotaomicron and E. faecalis were identified as significant predictors of IL-25 accumulation in the colon. Note: Animals assigned to different FMT treatments were first co-housed based on microbiome background to homogenize the microbiome and then separated according to FMT type.

F MT experiments demonstrate the causal role of microbiome in the induction of type 2 cytokines in the proximal colon, which correlates with the enrichment of B . thetaiotaomicron in the microbiome. (A and B) IL-25, IL-5, and IL-13 concentrations were quantified in the (A) proximal colon or (B) in the ileum (for IL-25) of C57BL/6 WT mice (see Materials and methods) receiving FMTs from mice treated with vancomycin or left untreated. Prior to FMT, mice were either untreated or treated with AVNM to eliminate the resident microbiota (Background). Protein concentration in the lysate (pg/ml) was normalized by the total protein mass generated in the sample (in mg) as in Buonuomo et al. (2016). Data are representative of independent experiments repeated twice, using n = 4–6 female mice per treatment group. ANOVA with Tukey post-hoc test was run to determine significant differences. Statistical significance was estimated at an FDR of 0.05. (C) 16S rRNA sequencing was conducted on input FMT and fecal samples collected from mice after FMT. (D) Volcano plots were generated following differential analysis using DeSeq2 on fecal microbiome sequencing samples from mice treated with AVNM and receiving FMT from either vancomycin-treated or untreated mice. Amplicon SVs from two succinate-producing species (B. thetaiotaomicron and E. faecalis) were found to be enriched in AVNM-treated mice receiving FMT from vancomycin-treated animals. Significance was determined based on an FDR of 0.05. (E) Permutation importance analysis followed by accumulated local effects calculations was carried out on the results of RFR modeling, which aimed to predict colonic IL-25 concentrations based on microbiome species abundance. Sequence variants associated with B. thetaiotaomicron and E. faecalis were identified as positive predictors of IL-25 concentrations in the colon. (F) The significance and directionality inferred by the RFR model were confirmed through Elastic Net Regression modeling and Bayesian Variable Selection Linear Regression. SVs linked to B. thetaiotaomicron and E. faecalis were identified as significant predictors of IL-25 accumulation in the colon. Note: Animals assigned to different FMT treatments were first co-housed based on microbiome background to homogenize the microbiome and then separated according to FMT type.

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