Glycobiology of commensal bacteria with emphasis on cell surface adhesins and exopolysaccharides
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The human gastro-intestinal tract (GIT) is an elaborate ecosystem specific to an individual and his physiological stage that involves inter- and intra-relationships between the bacteria composing the microbiota and between the host and the microbes. The idea of a communication system or a 'crosstalk' between the host and the microbiota has emerged after bacterial-derived molecules, such as polysaccharide A in Bacteroides fragilis, were shown to be essential for the maturation of the host immune system (Mazmanian et al., 2007). Understanding the molecular nature of the interaction between host gut epithelial cells and commensal microbes is essential for promotion of the mutualistic relationship with all its associated benefits for the host and prevention of the deleterious effects of infection by pathogens. Considerable efforts have been directed at the exploration of the lectin - adhesins of pathogenic bacteria and their relevant glycan receptors, with a view to identification of potential candidates for vaccines. To date, very little is known about adhesion mechanisms in gut commensal species. In particular, the lectin-like adhesins and their associated carbohydrate ligands have not been described. Recent technological advances in glycomics (omic databases, synthesis of glyco-conjugates and glycan microarrays) have provided new tools to study lectin/adhesin interactions with glycans. In this thesis, an in silico approach was used to identify surface-exposed proteins and putative adhesins of human commensal species, from the genera, Faecalibacterium, Bifidobacterium and Lactobacillus, that could represent potential probiotic strains. In vitro expression studies supported the in silico work, especially for the Lactobacillus species. Selected strains and their extracted pili were also incubated on mucin and neo-glycoconjugate microarrays and glycan ligands identified in F. prausnitzii, B. bifidum PRL2010 and L. paracasei subsp. paracasei. The potential of these novel approaches for the study of adhesins was thus demonstrated. The beneficial effects attributed to commensals in the GIT are in many cases exerted through exopolysaccharides (EPSs). Bacteria assemble a variety of glycoconjugates and polysaccharides as part of their cell walls. These include anchored molecules such as bacterial peptidoglycan, capsules and EPSs. Several functions, including health benefits for the host, have been attributed to EPSs. However, their isolation and analysis is a lengthy process that requires several analytical steps. The study of mammalian glycosylation has benefited significantly from recent developments in microarray technology, which enables the simultaneous analysis of the interactions between glycans and immobilised carbohydrate-binding proteins, such as plant lectins. Here, lectin microarray technology was successfully used for the first time to profile and differentiate fluorescently-labelled EPSs from commensal strains, as confirmed by monosaccharide analyses. Thus will help to move our understanding of EPS molecules ahead more quickly and promote their exploitation.