Lactobacillus johnsonii NCC 533 is a member of the acidophilus group of intestinal lactobacilli that has been extensively studied for their ''probiotic'' activities that include, pathogen inhibition, epithelial cell attachment, and immunomodulation. To gain insight into its physiology and identify genes potentially involved in interactions with the host, we sequenced and analyzed the 1.99-Mb genome of L. johnsonii NCC 533. Strikingly, the organism completely lacked genes encoding biosynthetic pathways for amino acids, purine nucleotides, and most cofactors. In apparent compensation, a remarkable number of uncommon and often duplicated amino acid permeases, peptidases, and phosphotransferase-type transporters were discovered, suggesting a strong dependency of NCC 533 on the host or other intestinal microbes to provide simple monomeric nutrients. Genome analysis also predicted an abundance (>12) of large and unusual cell-surface proteins, including fimbrial subunits, which may be involved in adhesion to glycoproteins or other components of mucin, a characteristic expected to affect persistence in the gastrointestinal tract (GIT). Three bile salt hydrolases and two bile acid transporters, proteins apparently critical for GIT survival, were also detected. In silico genome comparisons with the >95% complete genome sequence of the closely related Lactobacillus gasseri revealed extensive synteny punctuated by clear-cut insertions or deletions of single genes or operons. Many of these regions of difference appear to encode metabolic or structural components that could affect the organisms competitiveness or interactions with the GIT ecosystem.

The aim of this work was to identify Lactobacillus johnsonii NCC533 (La1) surface molecules mediating attachment to intestinal epithelial cells and mucins. Incubation of Caco-2 intestinal epithelial cells with an L. johnsonii La1 cell wall extract led to the recognition of elongation factor Tu (EF-Tu) as a novel La1 adhesin-like factor. The presence of EF-Tu at the surface of La1 was confirmed by analysis of purified outer surface protein extract by immunoblotting experiments, by electron microscopy, and by enzyme-linked immunosorbent assays of live bacteria. Furthermore, tandem mass spectrometry analysis proved that EF-TU was expressed at the La1 surface as an intact molecule. Using recombinant La1 EF-Tu protein, we were able to determine that its binding to intestinal cells and to mucins is pH dependent. Competition experiments suggested that EF-Tu has an important role in La1 mucin binding capacity. In addition, immunomodulation studies performed on HT29 cells showed that EF-Tu recombinant protein can induce a proinflammatory response in the presence of soluble CD14. Our in vitro results indicate that EF-Tu, through its binding to the intestinal mucosa, might participate in gut homeostasis.Probiotic bacteria, mainly lactic acid bacteria and bifidobacteria, have been shown to have beneficial effects on the immune defenses and to alleviate or prevent diverse intestinal disorders (3,4,16,25,27,30,39,42,47). Several in vitro studies have shown that one of them, Lactobacillus johnsonii NCC533 (La1), is able to bind to epithelial cell lines (5,8,9,21) and can induce the secretion of different cytokines in coculture systems (9, 24). Furthermore, human and animal studies have demonstrated that La1 has immune adjuvant effects (40, 44) and can also act as a modulator of nonspecific immune responses (6,14,29,48,53,62).The mechanisms underlying these beneficial effects are not completely understood, but it is believed that the maximum probiotic effects can be achieved if the organisms adhere to mucus and/or intestinal epithelial cells (31, 62). It has recently been shown that lipoteichoic acid (LTA), a molecule associated with the surface of La1 bacteria, participates in their adhesion to intestinal cells (21) and has an immunomodulatory effect on gut homeostasis (64). However, competition experiments indicated that LTA is not the only surface molecule mediating La1 binding to epithelial cells (21). Indeed, it had already been suggested by Bernet et al. (5) that proteinaceous compounds are involved in the attachment of bacteria to these cells. This observation is in accordance with recent studies showing that surface proteins of other lactobacilli participate in adhesion to epithelial cell lines, gastrointestinal mucins, or extracellular matrix proteins (1,26,58,60).In this work, therefore, we have investigated the ability of La1 surface proteins to attach to intestinal epithelial cells and mucoproteins. We have identified the elongation factor Tu as a novel surface protein possessing the characteristics of an adhesion ...

We have investigated a new class of food-grade particles, whey protein microgels, as stabilisers of triglyceride-water emulsions. The sub-micron particles stabilized oil-in-water emulsions at all pH with and without salt. All emulsions creamed but exhibited exceptional resistance to coalescence. Clear correlations exist between the properties of the microgels in aqueous dispersion and the resulting emulsion characteristics. For conditions in which the particles were uncharged, fluid emulsions with relatively large drops were stabilised, whereas emulsions stabilized by charged particles contained smaller flocculated drops. A combination of optical microscopy of the drops and spectrophotometry of the resolved aqueous phase allowed us to estimate the interfacial adsorption densities of the particles using the phenomenon of limited coalescence. We deduce two classes of particle arrangement. Complete adsorption of the particles was obtained when they were neutral or when their charges were screened by salt resulting in at least one particle monolayer at the interface. By contrast, only around 50% of the particles adsorbed when they were charged with emulsion drops being covered by less than half a monolayer. These findings were supported by direct visualization of drop interfaces using cryo-scanning electron microscopy. Uncharged particles were highly aggregated and formed a continuous 2-D network at the interface. Otherwise particles organized as individual aggregates separated by particle-free regions. In this case, we suggest that some particles spread at the interface leading to the formation of a continuous protein membrane. Charged particles displayed the ability to bridge opposing interfaces of neighbouring drops to form dense particle disks protecting drops against coalescence; this is the main reason for the flocculation and stability of emulsions containing sparsely covered drops.