Evaluating the prebiotic activity of arabinogalactan on the human gut microbiota using 16S rRNA gene sequencing and Raman-activated cell sorting.

Abstract

Background: Arabinogalactan is a complex plant-derived polysaccharide proposed to function as a selective prebiotic, yet the microbial taxa directly involved in its metabolism and the cooperative dynamics within the gut microbiota remain incompletely defined. Methods: Here, we combined community-level sequencing with targeted single-cell activity profiling to investigate how arabinogalactan shapes gut microbial composition and function. Fecal samples from ten healthy individuals were incubated ex vivo with arabinogalactan, and microbial responses were assessed using 16S rRNA gene amplicon sequencing alongside Raman-activated cell sorting (RACS) and coculture experiments. Results: Arabinogalactan consistently enriched Bifidobacterium and Gemmiger across donors, with Bifidobacterium also responding to galactose and Gemmiger and Blautia stimulated by arabinose, the two monosaccharide components of arabinogalactan. RACS enabled the selective isolation of metabolically active arabinogalactan responders, including Bifidobacterium longum (B. longum) and Faecalibacterium prausnitzii, along with other strains from the phyla Actinomycetota, Bacteroidota, and Bacillota. Notably, coculture experiments revealed that B. longum not only degraded arabinogalactan efficiently but also supported the growth of non-degrading species via metabolic cross-feeding. These cooperative interactions highlight B. longum as a keystone species in arabinogalactan utilization and suggest broader community-level benefits from its activity. Conclusion: Together, our findings demonstrate arabinogalactan's bifidogenic effect and its potential to promote functionally important microbes within the gut ecosystem. This study also highlights the utility of RACS for linking microbial identity to function, enabling the targeted recovery of active strains from complex communities.