Figure 3: Lactulose-mediated Bifidobacteria expansion is associated with significant changes in bioactive fecal metabolites.

(A) Volcano plot (log₂fold change vs. log₁₀p-value) of qualitative metabolites comparing samples with low (< 10%) vs. high (≥10%) Bifidobacteria abundance after lactulose exposure. P-values were calculated using a two-tailed Wilcoxon rank sum test and are corrected for multiple comparisons using the Benjamini-Hochberg procedure. Values with log2 fold-change > 1 (corresponding to a 2-fold change with a p-value < 0.05) were considered significant. (B) select SCFA and BA were quantified. Units for SCFA are mM, and units for BA derivatives are in μg/mL. (C) BA conversion from conjugated-primary BA to primary BA and then to secondary BAs was tested for each sample. Each point represents a molar ratio for an individual sample. For all comparisons (A-C), there is one sample per patient that was chosen based on the highest relative abundance of Bifidobacteria, and sample size was n=87 (lactulose exposure <10% Bifidobacteria) and n=72 (lactulose exposure with ≥10% Bifidobacteria). For panels B and C, each point represents a single sample. Median and interquartile range are indicated by the horizontal line and box, respectively. The lower vertical line depicts Q1 – 1.5*IQR and the upper vertical line depicts Q3 + 1.5*IQR. Statistical comparisons between individual groups were analyzed using a two-tailed Wilcoxon rank sum test. P-values are adjusted for multiple comparisons using the Benjamini-Hochberg procedure and represented as follows: *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001. For both panels B and C, Samples were grouped by whether they had expanded Bifidobacteria in the presence of lactulose. Median and interquartile range are indicated by the line and box, respectively. The lower vertical line depicts Q1 – 1.5*IQR and the upper vertical line depicts Q3 + 1.5*IQR. CA: cholic acid; GCA: glycocholic acid; DCA: deoxycholic acid.