Project description:Diet-microbe interactions play a crucial role in infant development and modulation of the early-life microbiota. The genus Bifidobacterium dominates the breast-fed infant gut, with strains of B. longum subsp. longum (B. longum) and B. longum subsp. infantis (B. infantis) particularly prevalent within the early-life microbiota. Although, transition from milk to a more diversified diet later in infancy initiates a shift to a more complex microbiome, with concurrent reductions in Bifidobacterium abundance, specific strains of B. longum may persist in individual hosts for prolonged periods of time. Here, we sought to investigate the adaptation of B. longum to the changing infant diet during the early-life developmental window. Genomic characterisation of 75 strains isolated from nine either exclusively breast- or formula-fed infants in the first 18 months of their lives revealed subspecies- and strain-specific intra-individual genomic diversity with respect to glycosyl hydrolase families and enzymes, which corresponded to different dietary stages. Complementary phenotypic growth studies indicated strain-specific differences in human milk oligosaccharide and plant carbohydrate utilisation profiles between and within individual infants, while proteomic profiling identified proteins involved in metabolism of selected carbohydrates. Our results indicate a strong link between infant diet and B. longum subspecies/strain genomic and carbohydrate utilisation diversity, which aligns with a changing nutritional environment i.e. moving from breast milk to a solid food diet. These data provide additional insights into possible mechanisms responsible for the competitive advantage of this bifidobacterial species and their long-term persistence in a single host and may contribute to rational development of new dietary therapies for this important development window.
Project description:This work aimed to investigate the ability of two human-derived bifidobacterial strains, i.e. Bifidobacterium breve UCC2003 and Bifidobacterium longum NCIMB 8809, to utilize various oligosaccharides (i.e., 4-galactosyl-kojibiose, lactulosucrose, lactosyl-oligofructosides, raffinosyl-oligofructosides and lactulose-derived galacto-oligosaccharides) synthesized by means of microbial glycoside hydrolases. With the exception of raffinosyl-oligofructosides, these biosynthetic oligosaccharides were shown to support growth of at least one of the two studied strains. Short-chain fatty acid (SCFA) analysis by HPLC corroborated the suitability of most of the studied novel oligosaccharides as growth substrates for the two bifidobacterial strains, showing that acetate is the main metabolic end product followed by lactic and formic acids. Transcriptomic and functional genomic approaches carried out for B. breve UCC2003 allowed the identification of key genes encoding glycoside hydrolases and protein transport systems involved in the metabolism of 4-galactosyl-kojibiose and lactulosucrose. In particular, the role of β-galactosidases in the hydrolysis of these particular trisaccharides was demonstrated, highlighting their importance in oligosaccharide metabolism by human bifidobacterial strains.
Project description:Members of the genus Bifidobacterium are typically found in the gastrointestinal tract (GIT) of humans and other mammals. Bifidobacterium breve strains are numerically prevalent among the gut microbiota of healthy, breast-fed infants. The metabolism of sialic acid, a ubiquitous monosaccharide in the infant and adult gut, by B. breve UCC2003 is dependent on a large gene cluster, designated the nan/nag cluster. This study describes the transcriptional regulation of the nan/nag cluster and thus sialic acid metabolism in B. breve UCC2003. Insertion mutagenesis and transcriptome analysis revealed that the nan/nag cluster is regulated by a GntR family transcriptional repressor, designated NanR. NanR was shown to bind to two promoter regions within this cluster, each of which containing an imperfect inverted repeat that is believed to act as the NanR operator sequence. Formation of the DNA-NanR complex is prevented in the presence of sialic acid, which we had previously shown to induce transcription of this gene cluster. NanR represents the first GntR-type transcriptional regulator to be characterised from bifidobacteria.
Project description:Members of the genus Bifidobacterium are common inhabitants of the gastrointestinal tract of humans and other mammals, where they ferment many diet-derived carbohydrates that cannot be digested by their host. To extend our understanding of bifidobacterial carbohydrate utilisation, we investigated the molecular mechanisms by which various strains of Bifidobacterium breve metabolize four distinct α-glucose and/or α-galactose-containing oligosaccharides, namely raffinose, stachyose, melibiose and melezitose. Here we demonstrate that all B. breve strains examined possess the ability to utilise raffinose, stachyose and melibiose. However, the ability to metabolize melezitose was not ubiquitous for all tested B. breve strains. Transcriptomic and functional genomic approaches identified a gene cluster dedicated to the metabolism of α-galactose-containing carbohydrates, while an adjacent gene cluster, dedicated to the metabolism of α-glucose-containing melezitose, was identified, yet being present only in those B. breve strains that were able to support growth on this carbohydrate.
Project description:Bifidobacterium breve represents one of the most abundant (bifido)bacterial species in the gastro-intestinal tract of (breast-fed) infants, where their presence is believed to be beneficial. In the present study whole genome sequencing, employing PacBio’s Single Molecule, Real-Time (SMRT) sequencing platform, combined with comparative genome analysis allowed the most extensive genetic investigation of this taxon. Our findings demonstrate that Restriction/Modification (R/M) systems constitute a substantial part of the B. breve variome. Availing of the methylome data generated by SMRT sequencing, combined with targeted Illumina bisulfite sequencing (BiSeq) and comparative genome analysis, we were able to detect methylation recognition motifs and assign these to identified B. breve R/M systems, where in several cases such assignments were confirmed by restriction analysis. Furthermore, cloning of a selected methyltransferase-encoding gene validated the activity of the corresponding R/M system, and was shown to overcome the barrier they impose to genetic accessibility, thus allowing future genetic manipulation of members of this species.
Project description:The purpose of this course is to study the relationship between the primary structure of exopolysaccharides and the gene expression level of Bifidobacterium longum W13 and W13R, and to better explain the differences between different strains from the level of gene transcription.
Project description:Transcriptional profiling of Bifidobacterium longum mutant versus wt strain in exponentional phase Keywords: Characterization of natural mutant