Insights into ß-manno-oligosaccharide uptake and metabolism in Bifidobacterium adolescentis DSMZ 20083 from whole-genome microarray analysis.

Metabolism of non-digestible dietary glycans directly influences the structure and composition of human gut microbiota and, in turn, the host health. ß-Mannans form an integral component of the modern diet as naturally occurring dietary fibre or additives in processed foods. In the present study, in vitro fermentation and TLC studies were used to determine the ability of adult-associated Bifidobacterium adolescentis DSMZ 20083 to utilise ß-manno-oligosaccharides from guar gum, locust bean gum, konjac root, and copra meal generated using GH26 endo-ß-mannanase (ManB-1601). Further, to gain insights into the underlying molecular mechanism, a whole-genome microarray analysis, RT-qPCR, and molecular docking studies were employed to reconstruct the copra meal ß-manno-oligosaccharides (CM-ß-MOS) utilisation pathway in B. adolescentis DSMZ 20083. B. adolescentis DSMZ 20083 grew appreciably (O.D600 nm up to 0.8) on all tested ß-manno-oligosaccharides but maximally on CM-ß-MOS. CM-ß-MOS having DP2-3 were found to deplete from the fermentation media. Whole-genome transcriptome analysis, RT-qPCR, and molecular docking studies suggested that in B. adolescentis DSMZ 20083, ABC & MFS transporters are possibly involved in the uptake of DP ≥ 2 and DP ≥ 3 linear CM-ß-MOS, respectively, while GH1 ß-glucosidase, and GH32 ß-fructofuranosidase possibly cleave linear CM-ß-MOS into monosaccharides. Sugar absorption and utilisation pathways; Bifid shunt, ABC transport system, pyruvate metabolism, glycolysis/gluconeogenesis, pentose, and glucouronate inter-conversions were also found up-regulated following the growth on CM-ß-MOS. This is the first study reporting on possible molecular determinants used by B. adolescentis DSMZ 20083 to utilise ß-manno-oligosaccharides. Our studies can prove resourceful to food and nutraceutical industries, aiming at precision microbiome modulation using ß-manno-oligosaccharides.

Co-culture fermentations suggest cross-feeding among Bacteroides ovatus DSMZ 1896, Lactiplantibacillus plantarum WCFS1 and Bifidobacterium adolescentis DSMZ 20083 for utilizing dietary galactomannans.

Galactomannans from sources like guar, fenugreek, locust bean, and copra form an important part of human diet. In this study, we have attempted to understand the cross-feeding and resource sharing between a generalist degrader and probiotic utilizers for utilizing dietary galactomannans. In mono-cultures, Bacteroides ovatus DSMZ 1896 grew maximally on substituted galactomannans and produced high amount of succinate. Polysaccharide break down products [ß-manno-oligosaccharides; degree of polymerization (DP) 2-4] left after the growth of B. ovatus DSMZ 1896 in galactomannan supplemented media supported the growth of Lactiplantibacillus plantarum WCFS1 (DP2 and DP3) and Bifidobacterium adolescentis DSMZ 20083 (majorly DP3) and led to the production of lactate and acetate, respectively as the major end products. Co-cultures (bi- and tri-cultures) studies demonstrated cross-feeding being used as a strategy for resource sharing among B. ovatus DSMZ 1896, L. plantarum WCFS1 and B. adolescentis DSMZ 20083 while foraging galactomannans. Structure and DP of galactomannan substrates altered the SCFA and organic acid production patterns in co-cultures.

Soluble and Cross-Linked Aggregated Forms of α-Galactosidase from Vigna mungo Immobilized on Magnetic Nanocomposites: Improved Stability and Reusability.

α-Galactosidases hold immense potential due to their biotechnological applications in various industrial and functional food sectors. In the present study, soluble and covalently cross-linked aggregated forms of a low molecular weight, thermo-labile α-galactosidase from Vigna mungo (VM-αGal) seeds were immobilized onto chitosan-coated magnetic nanoparticles for improved stability and repeated usage by magnetic separation. Parameters like precipitants (type, amount, and ratio), glutaraldehyde concentration, and enzyme load were optimized for the preparation of chitosan-coated magnetic nanocomposites of cross-linked VM-αGal (VM-αGal-MC) and VM-αGal (VM-αGal-M) resulted in 100% immobilization efficiency. Size and morphology of VM-αGal-M were studied through dynamic light scattering (DLS) and scanning electron microscopy (SEM), while Fourier transform infrared spectroscopy (FTIR) was used to study the chemical composition of VM-αGal-MC and VM-αGal-M. VM-αGal-MC and VM-αGal-M were found more active in a broad range of pH (3-8) and displayed optimal temperatures up to 25 °C higher than VM-αGal. Addition of non-ionic detergents (except Tween-40) improved VM-αGal-MC activity by up to 44% but negatively affected VM-αGal-M activity. Both VM-αGal-MC (15% residual activity after 21 min at 85 °C, Ed 92.42 kcal/mol) and VM-αGal-M (69.0% residual activity after 10 min at 75 °C, Ed 39.87 kcal/mol) showed remarkable thermal stability and repeatedly hydrolyzed the substrate for 10 cycles.

Structural diversity and prebiotic potential of short chain ß-manno-oligosaccharides generated from guar gum by endo-ß-mannanase (ManB-1601).

Size exclusion chromatography of short chain ß-manno-oligosaccharides (GG-ß-MOS) produced after endo-mannanase (ManB-1601) hydrolysis of guar gum resulted in seven (P1-P7) peaks. Electron spray ionization mass-spectrometry (ESI-MS) revealed P3, P4, P5 and P6 peaks as pentasaccharide (DP5), tetrasaccharide (DP4), trisaccharide (DP3) and disaccharide (DP2), respectively. DP2 and DP3 GG-ß-MOS were structurally characterized by NMR (1H and 13C), FTIR and XRD. DP2 GG-ß-MOS was composed of two species (A) mannopyranose ß-1,4 mannopyranose and (B) α-1,6-galactosyl-mannopyranose while, DP3 oligosaccharide showed presence of three species i.e. (A) α-d-galactosyl-ß-d-mannobiose (galactosyl residue at reducing end), (B) α-d-galactosyl-ß-d-mannobiose (galactosyl residue at non-reducing end) and (C) mannopyranose ß-1,4 mannose ß-1,4 mannopyranose. In batch fermentation, DP2 GG-ß-MOS was preferred over DP3 by all Lactobacillus sp. except Lactobacillus casei var rhamnosus. DP2/DP3 and GG-ß-MOS mixture inhibited the growth of enteropathogens in monoculture and co-culture fermentations, respectively. Fermentation of GG-ß-MOS mixture by Lactobacillus sp. produced short chain fatty acids.