In this work we analyzed the immune activation properties of different Bifidobacterium strains in order to establish their ability as inductors of specific effector (Th) or regulatory (Treg) responses. First, we determined the cytokine pattern induced by 21 Bifidobacterium strains in peripheral blood mononuclear cells (PBMCs). Results showed that four Bifidobacterium bifidum strains showed the highest production of IL-17 as well as a poor secretion of IFNγ and TNFα, suggesting a Th17 profile whereas other Bifidobacterium strains exhibited a Th1-suggestive profile. Given the key role of Th17 subsets in mucosal defence, strains suggestive of Th17 responses and the putative Th1 Bifidobacterium breve BM12/11 were selected to stimulate dendritic cells (DC) to further determine their capability to induce the differentiation of naïve CD4+ lymphocytes toward different Th or Treg cells. All selected strains were able to induce phenotypic DC maturation, but showed differences in cytokine stimulation, DC treated with the putative Th17 strains displaying high IL-1β/IL-12 and low IL-12/IL-10 index, whereas BM12/11-DC exhibited the highest IL-12/IL-10 ratio. Differentiation of naïve lymphocytes confirmed Th1 polarization by BM12/11. Unexpectedly, any B. bifidum strain showed significant capability for Th17 generation, and they were able to generate functional Treg, thus suggesting differences between in vivo and vitro responses. In fact, activation of memory lymphocytes present in PBMCS with these bacteria, point out the presence in vivo of specific Th17 cells, supporting the plasticity of Treg/Th17 populations and the key role of commensal bacteria in mucosal tolerance and T cell reprogramming when needed.

ABSTRACTThe ability of bifidobacteria to establish in the intestine of mammals is among the main factors considered to be important for achieving probiotic effects. The role of surface molecules fromBifidobacterium bifidumtaxon in mucin adhesion capability and the aggregation phenotype of this bacterial species was analyzed. Adhesion to the human intestinal cell line HT29 was determined for a collection of 12B. bifidumstrains. In four of them—B. bifidumLMG13195, DSM20456, DSM20239, and A8—the involvement of surface-exposed macromolecules in the aggregation phenomenon was determined. The aggregation ofB. bifidumA8 and DSM20456 was abolished after treatment with proteinase K, this effect being more pronounced for the strain A8. Furthermore, a mucin binding assay ofB. bifidumA8 surface proteins showed a high adhesive capability for its transaldolase (Tal). The localization of this enzyme on the surface ofB. bifidumA8 was unequivocally demonstrated by immunogold electron microscopy experiments. The gene encoding Tal fromB. bifidumA8 was expressed inLactococcus lactis, and the protein was purified to homogeneity. The pure protein was able to restore the autoaggregation phenotype of proteinase K-treatedB. bifidumA8 cells. A recombinantL. lactisstrain, engineered to secrete Tal, displayed a mucin- binding level more than three times higher than the strain not producing the transaldolase. These findings suggest that Tal, when exposed on the cell surface ofB. bifidum, could act as an important colonization factor favoring its establishment in the gut.

Probiotics are live microorganisms that when administered in adequate amounts confer a health benefit on the host. They are mainly bacteria from the genera Lactobacillus and Bifidobacterium. Traditionally, functional properties of lactobacilli have been studied in more detail than those of bifidobacteria. However, many recent studies have clearly revealed that the bifidobacterial population in the human gut is far more abundant than the population of lactobacilli. Although the 'beneficial gut microbiota' still remains to be elucidated, it is generally believed that the presence of bifidobacteria is associated with a healthy status of the host, and scientific evidence supports the benefits attributed to specific Bifidobacterium strains. To carry out their functional activities, bifidobacteria must be able to survive the gastrointestinal tract transit and persist, at least transiently, in the host. This is achieved using stress response mechanisms and adhesion and colonization factors, as well as by taking advantage of specific energy recruitment pathways. This review summarizes the current knowledge of the mechanisms involved in facilitating the establishment, colonization, and survival of bifidobacteria in the human gut.

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Bifidobacteria are widely used as probiotics in several commercial products; however, to date there is little knowledge about their carbohydrate metabolic pathways. In this work, we studied the metabolism of glucose and lactose in the widely used probiotic strain Bifidobacterium animalis subsp.

The use of beneficial microorganisms, the so-called probiotics, to improve human health is gaining popularity. However, not all of the probiotic strains trigger the same responses and they differ in their interaction with the host. In spite of the limited knowledge on mechanisms of action some of the probiotic effects seem to be exerted through maintenance of the gastrointestinal barrier function and modulation of the immune system. In the present work, we have addressed in vitro the response of the intestinal epithelial cell line HT29 to the strain Bifidobacterium breve IPLA20004. In the array of 84 genes involved in inflammation tested, the expression of 12 was modified by the bifidobacteria. The genes of chemokine CXCL6, the chemokine receptor CCR7, and, specially, the complement component C3 were upregulated. Indeed, HT29 cells cocultivated with B. breve produced significantly higher levels of protein C3a. The proteome of HT29 cells showed increased levels of cytokeratin-8 in the presence of B. breve. Altogether, it seems that B. breve IPLA20004 could favor the recruitment of innate immune cells to the mucosa reinforcing, as well as the physical barrier of the intestinal epithelium.

Bacillus cereus CH is a probiotic strain used in human nutrition whose adhesion to mucin is dependent on its surface-associated flagellin. Flagellins from the surface of several probiotic Bacillus strains were efficiently extracted with 5 M LiCl and identified by peptide fingerprinting. Based on the proteomic analysis, cloning of the gene coding for the flagellin of B. cereus CH was performed in the lactococcal vector pNZ8110 under the control of a nisin-inducible promoter. The resulting strain, Lactococcus lactis CH, produced a surface-associated flagellin after 6 h of induction with nisin. The recombinant Lactococcus strain adhered strongly to mucin-coated polystyrene plates, whilst inhibiting competitively the adhesion of the pathogens Escherichia coli LMG2092 and Salmonella enterica ssp. enterica LMG15860 to the same molecule. Strain CH could be used in further experimentation for the characterization of the molecular mechanism of action of this probiotic B. cereus CH flagellin.