FIG 2.

Growth of C. difficile 630 on FOS in vitro and differential impacts of FOS and inulin on SCFA production by the microbiome in C. difficile 630-infected mice. (A) C. difficile 630 was grown in PETC-F minimal medium (MM, black lines), MM + FOS (purple lines), and MM + inulin (red lines) (n = 5 biological replicates per strain), for 24 h, and culture density (OD₆₀₀) was monitored (left). Lines and error bars represent mean OD₆₀₀ readings and standard deviation at each time point, respectively. Statistically significant differences between the final OD₆₀₀s of these cultures were determined by Mann-Whitney test (*, P < 0.05). (B) Filtered supernatants from these cultures were analyzed using high-performance anion-exchange chromatography and a pulsed amphoteric detector (HPAEC-PAD). Representative chromatograms are shown for uninoculated media (left, green) and for spent media (right, purple). See Fig. S1 in the supplemental material for a reference chromatogram, demonstrating that the metabolites depleted by C. difficile are monosaccharides that contaminate the FOS preparation rather than FOS itself. (C) The SCFAs acetate, propionate, and butyrate were quantified in the cecal contents of mice described in the legend to Fig. 1, collected after euthanasia at 19 days postinfection. Individual data points represent SCFA concentrations measured via GC-MS, and bars represent mean concentrations. Blue bars represent SCFAs quantified in mice fed the MAC-deficient diet, purple bars represent SCFAs quantified in mice fed the FOS-containing diet, and red bars represent SCFAs quantified in mice fed the inulin-containing diet. Statistical significance was determined by one-way analysis of variance with Tukey’s multiple-comparison test (*, P < 0.05; ***, P < 0.001; ****, P < 0.0001). See also Fig. S1.