Extraction of the same novel homoglycan mixture from two different strains of Bifidobacterium animalis and three strains of Bifidobacterium breve

Abstract

Three strains of Bifidobacterium breve (JCM 7017, JCM 7019 and JCM 2258) and two strains of Bifidobacterium animalis subsp. lactis (AD011 and A1dOxR) were grown in broth cultures or on plates, and a standard exopolysaccharide extraction method was used in an attempt to recover exocellular polysaccharides. When the extracted materials were analysed by NMR it was clear that mixtures of polysaccharides were being isolated including exopolysaccharides (EPS) cell wall polysaccharides and intracellular polysaccharides. Treatment of the cell biomass from the B. breve strains, or the B. animalis subsp. lactis AD011 strain, with aqueous sodium hydroxide provided a very similar mixture of polysaccharides but without the EPS. The different polysaccharides were partially fractionated by selective precipitation from an aqueous solution upon the addition of increasing percentages of ethanol. The polysaccharides extracted from B. breve JCM 7017 grown in HBM media supplemented with glucose (or isotopically labelled D-glucose-1-C) were characterised using 1D and 2D-NMR spectroscopy. Addition of one volume of ethanol generated a medium molecular weight glycogen (Mw=1×10 Da, yield 200 mg/l). The addition of two volumes of ethanol precipitated an intimate mixture of a low molecular weight β-(1→6)-glucan and a low molecular weight β-(1→6)-galactofuranan which could not be separated (combined yield 46 mg/l). When labelled D-glucose-1-C was used as a carbon supplement, the label was incorporated into >95% of the anomeric carbons of each polysaccharide confirming they were being synthesised in situ. Similar H NMR profiles were obtained for polysaccharides recovered from the cells of B. animalis subsp. lactis AD011and A1dOxR (in combination with an EPS), B. breve JCM 7017, B. breve JCM 7019, B. breve JCM 2258 and from an EPS (-ve) mutant of B. breve 7017 (a non-EPS producer).

Bifidobacterium breve UCC2003 Exopolysaccharide Modulates the Early Life Microbiota by Acting as a Potential Dietary Substrate

“…Previous studies on B. breve EPS has identified glucose and galactose as main components of EPS, which is similar to that proposed for UCC2003 EPS structure (includes glucose, galactose and/or the N-acetylated versions of these two sugars in different ratios or composition) [12,[14][15][16]. The large molecular weight and structure of these EPSs may prevent host-associated digestion, allowing them to act as prebiotics within the colon (similar to inulin and fructooligosaccharide) [18,33,34].…”

“…Bacterial EPS are polymerized mono-or oligo-saccharides which form a diverse range of homo-or hetero-polysaccharides, that can be linked to the bacterial cell wall or secreted. Analyses of several Bifidobacterium species has revealed the presence of EPS gene clusters [13], with chemical analysis indicating glucose and galactose as major components, with very low levels of rhamnose also present throughout these structures [14,15]. To date, most studies have focused on EPS-host interactions including: adhesion to host cells, and modulation of epithelial and immune responses [16][17][18].…”

Bifidobacterium breveUCC2003 exopolysaccharide modulates the early life microbiota by acting as a dietary substrate

“…Previous studies on B. breve EPS has identified glucose and galactose as main components of EPS, which is similar to that proposed for UCC2003 EPS structure (includes glucose, galactose and/or the N-acetylated versions of these two sugars in different ratios or composition) [13, 15-17]. The large molecular weight and structure of these EPS may prevent host-associated digestion, allowing them to act as prebiotics within the colon (similar to inulin and fructooligosaccharide) [19, 29, 30].…”

“…Bacterial EPS are polymerised mono- or oligo-saccharides which form a diverse range of homo- or hetero-polysaccharides, that can be linked to the bacterial cell wall or secreted. Analyses of several Bifidobacterium species has revealed the presence of EPS gene clusters [14], with chemical analysis indicating glucose and galactose as major components, with very low levels of rhamnose also present throughout these structures [15, 16]. To date, most studies have focused on EPS-host interactions including; adhesion to host cells, and modulation of epithelial and immune responses [17-20].…”

Bifidobacterium breve Exopolysaccharide Blocks Dendritic Cell Maturation and Activation of CD4+ T Cells

“…Furthermore, the administration of EPS deficient B. breve to mice increased susceptibility to Citrobacter rodentium infection and reduced persistence of B. breve in the murine GIT ( Fanning et al, 2012 ). Although many studies have used purified EPS to characterize host responses, this approach is susceptible to contamination with additional MAMPs derived from components of bacterial cell walls, membranes, or even cytoplasms ( Alhudhud et al, 2018 ). An alternative reductionist approach is to investigate the effects of EPS using EPS deficient isogenic mutants of EPS producers and compare immune cell responses to both WT EPS proficient and mutant EPS deficient strains.…”