Biology subjects
Publications and source records attributed to Lagos, L..
Withdrawal StatementThis manuscript has been withdrawn by the authors as it was submitted and made public without the full consent of all the authors. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.
The study of specific glycan uptake and metabolism has been shown to be an effective tool in aiding with the continued unravelling of the complexities in the human gut microbiome. To this aim fluorescent labelling of glycans may provide a powerful route towards target. In this study, we successfully used the fluorescent label 2-aminobenzamide (2-AB), most commonly employed for enhancing the detection of protein anchored glycans, to monitor and study microbial degradation of labelled glycans. Both single strain and co-cultured fermentations of microbes from the common human-gut derived Bacteroides genus, were able to grow when supplemented with 2-AB labelled glycans of different monosaccharide composition, degrees of acetylation and polymerization. Utilizing a multifaceted approach that combines chromatography, mass spectrometry, microscopy and flow cytometry techniques, it was possible to comprehensively track the metabolism of the labelled glycans in both supernatants and at a single cell level. We envisage this combination of complimentary techniques will help further the understanding of substrate specificity and the role it plays within microbial communities. IMPORTANCEInformation on how bacterial consortia utilize polysaccharides at strain level, whilst progressing rapidly in recent years still lacks a suitable way to study the vast range of ornamentations and structural motifs found in the natural glycans we consume in everyday life. As multi-omic approaches commonly require complex and costly analysis, a screening platform, as described in our work, could be seen as both a complementary and essential new tool in the understanding of microbial polysaccharide metabolism. Our study demonstrates a fast and efficient glycan labelling technique composed of several integrated procedures and advanced analytical methodologies. Chromatography and mass spectrometry are applied in the tracking of metabolized labelled glycans whilst microscopy and flow cytometry are used in the visualization of labelled bacteria at a single cell level.
Beneficial modulation of the gut microbiome has high-impact implications not only in humans, but also in livestock that sustain our current societal needs. In this context, we have tailored an acetylated galactoglucomannan (AcGGM) fibre to match unique enzymatic capabilities of Roseburia and Faecalibacterium species, both renowned butyrate-producing gut commensals. The accuracy of AcGGM was tested within the complex endogenous gut microbiome of pigs, wherein we resolved 355 metagenome-assembled genomes together with quantitative metaproteomes. In AcGGM-fed pigs, both target populations differentially expressed AcGGM-specific polysaccharide utilization loci, including novel, mannan-specific esterases that are critical to its deconstruction. However, AcGGM-inclusion also manifested a "butterfly effect", whereby numerous metabolic changes and interdependent cross-feeding pathways were detected in neighboring non-mannolytic populations that produce short-chain fatty acids. Our findings show that intricate structural features and acetylation patterns of dietary fibre can be customized to specific bacterial populations, with potential to create greater modulatory effects at large.