Figure 4. Members of the gut microbiota respond transcriptionally to S-BMO in vivo and degrade S-BMO in vitro.
(A) Relative abundance of E. coli (mean±SEM) over time in the fecal microbiota of gnotobiotic mice colonized with the stunted Malawian infant’s culture collection and fed M8 ± S-BMO. *p<0.05 two-tailed, unpaired Student’s t-test. (B) Volcano plot of bacterial gene expression in the cecal microbiota of gnotobiotic mice. Expression is plotted on the x-axis as the log2 fold difference between S-BMO-supplemented mice and unsupplemented mice. Significantly up- or downregulated genes are labeled in black or by bacterial species of origin. −Log10(p-values) are plotted on the y-axis (Benjamini-Hochberg corrected negative binomial test). (C) Heatmap of select differentially expressed E. coli genes, grouped by KEGG functional pathway (see Table S6 for a full list). Red asterisks denote that differences in expression are statistically significant after Benjamini-Hochberg correction for multiple hypotheses (α=0.1). (D) Effects of S-BMO on expression of genes in a B. fragilis PUL. Expression is plotted on the y-axis as the log2 fold difference (mean±SEM) between S-BMO-supplemented and control mice. *p<0.05, **p<0.01 (two-tailed, unpaired Student’s t-test). (E) Abundance (mean±SEM) of siallyllactose and sialic acid in E. coli or B. fragilis monoculture supernatants after a 24-hour incubation with 5% S-BMO. Control incubations contained uninoculated PBS buffer. ***p<0.001 two-tailed, unpaired Student’s t-test. (F) OD600 (mean±SEM) of E. coli grown in minimal medium containing various single carbon sources. ‘Conditioned S-BMO’ refers to filter-sterilized supernatant from a 24-hour monoculture of B. fragilis with S-BMO. (G) Mouse weights (mean±SEM), normalized to values at time of colonization with the entire Malawian infant culture collection or with only B. fragilis and E. coli (Bf + Ec). Mice were fed M8 ± S-BMO (n=5 mice/group).