Analysis of metabolic interactions in an artificial consortium including butyrogenic gut microbes

Cross-feeding interactions shape gut microbial communities and contribute to the establishment of bacteria associated with beneficial fermentation products such as butyrate. However, many gut microbes involved in fiber degradation and butyrate-associated trophic networks remain poorly characterized. Here, we combined genomic analysis of candidate strains, genome-scale metabolic modeling, in vitro cultivation, and transcriptomics to investigate metabolic interactions among gut bacteria during growth on inulin. Genomic analyses identified genes associated with inulin utilization, fermentation enzymes, and amino acid synthesis and transport. In vitro screening of different bacterial combinations on inulin identified Clostridium sp. HGF2 as an inulin-degrading, butyrate-producing strain, leading us to focus on its interaction with Bifidobacterium animalis subsp. lactis PT33. Co-culture assays, transwell experiments, and batch bioreactor analyses supported metabolic interactions between both strains during growth on inulin. Although PT33 did not enhance butyrate production, the combined results were consistent with competition for readily available carbohydrates, while transcriptomics and model integration suggested that HGF2 degrades inulin and releases simple carbohydrates that can be utilized by PT33, while PT33 produces acetate and lactate that support HGF2 metabolism. Altogether, this work shows a methodology to identify and evaluate candidate cross-feeding interactions in artificial gut communities, in this case, revealing a competitive rather than synergistic interaction, and characterizes HGF2, identified here as Clostridium innocuum, as an understudied butyrate-producing, inulin-degrading strain.

Understanding microbial and metabolic interactions that support butyrate producers is important for developing strategies to promote beneficial gut bacteria. However, most cross-feeding studies have focused on butyrate producers from the genera Faecalibacterium, Eubacterium, Anaerostipes, and Roseburia, leaving many other butyrate-producing bacteria poorly characterized. Here, we characterize the interaction between Clostridium sp. HGF2 and Bifidobacterium animalis subsp. lactis PT33 using RNA-seq and genome-scale metabolic modeling. Starting from a pool of candidate butyrate producers selected through genomic analysis and in vitro experiments, we identified HGF2 as a strain of interest and explored its metabolic interaction with PT33.