Search

In the complex interplay of beneficial bacteria with the host, there are few examples of bacterial metabolites and effector molecules that have been consistently identified. Protective effects on the intestinal epithelium have been ascribed to P40 and P75, two well characterized cell wall muramidases, present in the culture supernatant of strains belonging to the taxon Lactobacillus casei/paracasei/rhamnosus. This work reports that Lactobacillus casei BL23 extracellular vesicles (BL23 EVs) have a small size (17–20 nm or 24–32 nm, depending on the method used) and contain lipoteichoic acid (LTA). Interestingly, all detected P40 and most of P75 were associated to EVs and possibly located at their external surface, as shown by proteinase K digestion. Biosensor assays showed that both proteins bind LTA and vesicles, suggesting that they could bind to ligands like LTA present on BL23 EVs. Native BL23 EVs have a moderate proinflammatory effect and they were able to induce phosphorylation of the epidermal growth factor receptor (EGFR), showing an effect similar to purified P40 and P75 and leading to the conclusion that the activity described in the supernatant (postbiotic) of these bacteria would be mainly due to P40 and P75 bound to EVs., Grant of the Spanish Ministry of Science and Universities RTI2018-097982-B-I00 and the CONSOLIDER Proposal nber. 25506, that provided the Biacore T100 instrument

Proteinase PrtP (EC:3.4.21.96) is a cell envelope proteinase (CEP) highly expressed in the probiotic strain Lactobacillus paracasei BL312(VSL#3) that accounts for its anti-inflammatory properties. The main aim of this work is to understand differences in CEP expression between this strain and L. paracasei BL23. Hence, differences in the regulation by amino acid sources of four proteinase related genes (prtP, prsA, prtR1 and prtR2) were determined by RT-qPCR in BL312(VSL#3) and BL23 using as a reference BL368, a BL23 derepressed mutant lacking the response regulator (RR) PrcR. BL312(VSL#3) showed greater expression of prtP (2- to 3-fold) than BL23, and prtP was highly repressed by peptone in both strains. Two other putative CEP genes, prtR1 and prtR2, showed a low expression profile. Interestingly, when the prsA-prtP promoter region from both strains, and deleted mutants, were cloned in vector pT1GR, expression of the gfp and mrfp fluorescent reporters was always repressed in BL23 (high or low peptone) and derepressed in BL368, revealing an interesting mechanism of regulation affecting specifically to this promoter. In conclusion, BL312(VSL#3) has higher expression of prtP and other CEP related genes than BL23, that could respond to a natural deregulation in this strain, possibly independent from the RR PrcR., Grants of the Spanish Ministry of Science and Universities AGL2010‐1567, AGL2015-70487-P, RTI2018-097982-B-I00, and from the Generalitat Valenciana ACOMP2012/137. Support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI).

In this study the rheological properties of aqueous suspensions of three microalgae species, Nannochloropsis gaditana, Scenedesmus almeriensis and Spirulina platensis, were investigated as a function of solids content, and related to their composition and microstructure. In addition, the impact of ultrasound processing on their structuring ability was also studied. The less rigid character of the Spirulina platensis cell walls (with very low carbohydrate contents) and the presence of extracellular components promoted cell-cell interactions, yielding suspensions which showed a shear thinning behaviour at lower concentrations than Nannochloropsis gaditana and Scenedesmus almeriensis. It is noteworthy that the three species showed different viscoelastic properties at 25 wt.% total solids. Spirulina platensis suspensions showed a more elastic behaviour and lower frequency dependence, characteristic of weak gels, whilst Nannochloropsis gaditana and Scenedesmus almeriensis behaved more like viscous liquids. The ultrasound treatment did not affect the cell wall integrity, but it promoted the release of intracellular components (some of which could have been partially degraded) and disrupted physical interparticle interactions in Nannochloropsis gaditana and Scenedesmus almeriensis. This has an impact on the rheological properties, increasing the viscosity of Nannochloropsis gaditana suspensions, whilst the viscosity of Scenedesmus almeriensis suspensions was reduced. The outcomes of this work give insights into the exploitation of these microalgae species in soft materials for food, pharma and other technological applications., This work was supported by the COST Action ES1408 European network for algal-bioproducts (EUALGAE).

Lactobacillus casei and Lactobacillus rhamnosus proteins P40 and P75 belong to a large family of secreted cell wall proteins that contain a carboxy(C)-terminal CHAP or NlpC/P60 superfamily domains. In addition to their peptidoglycan hydrolases activity, proteins in this family are specific antigens of pathogens, frequently responsible of interactions with the host. L. rhamnosus GG and L. casei BL23 purified P40 and P75 proteins have antiapoptotic activity by inducing the EGF/Akt pathway. The aim of this work was to study the genetics, phylogeny and dissemination of this family of proteins in the genus Lactobacillus as well as their characteristics and likely function. The scrutiny of their DNA encoding sequences revealed the presence of minisatellite DNA in the P75 encoding gene of L. casei/paracasei strains (cmuB) with intraspecific indels that gave raise to four different alleles (cmuB1–4), which are exclusive of this species. Phylogenic analyses suggest that both proteins are present mainly in the L. casei and Lactobacillus sakei phylogenomic groups. A P40 ancestral gene was possibly present in the common ancestor of Enterococcaceae, Lactobacillaceae and Streptococcaceae. P75 is also present in L. casei and L. sakei groups, but its evolution is difficult to explain only by vertical transmission. Antibodies raised against the N-terminal regions of P40 and P75 improved their immunological detection in culture supernatants as they recognized almost exclusively proteins of L. casei/paracasei/rhamnosus strains, highlighting their structural similarity, that allowed to detect them in different fermented dairy products that contained probiotic L. casei strains. Purified P40 and P75 proteins showed no evident lytic activity but they complemented L. casei BL23 cmuA and cmuB defective mutants, respectively, thus proving that they actively participate in cell division., We wish to thank the Grants of the Spanish Ministry of Science and Universities AGL2013-47420-R and AGL2015-70487-P. GA and SS wish to thank the grant from the L. N. Gumilyov Eurasian National University. We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI)

The synthesis of the inorganic polymer polyphosphate (poly-P) in bacteria has been linked to stress survival and to the capacity of some strains to sequester heavy metals. In addition, synthesis of poly-P by certain strains of probiotic lactobacilli has been evidenced as a probiotic mechanism due to the homeostatic properties of this compound at the intestinal epithelium. We analyzed the link between poly-P synthesis, stress response, and mercury toxicity/accumulation by comparing wild-type strains of

Fucosyl-N-acetylglucosamine disaccharides are important core structures that form part of human mucosal and milk glyco-complexes. We have previously shown that AlfB and AlfC α-L-fucosidases from Lactobacillus casei are able to synthesize fucosyl-α-1,3--N-acetylglucosamine (Fuc-α1,3-GlcNAc) and fucosyl-α-1,6-N-acetylglucosamine (Fuc-α1,6-GlcNAc), respectively, in transglycosylation reactions. Here, these reactions were performed in a semipreparative scale, and the produced disaccharides were purified. The maximum yields obtained of Fuc-α1,3-GlcNAc and Fuc-α1,6-GlcNAc were 4.2 and 9.3 g/l, respectively. The purified fucosyl-disaccharides were then analyzed for their prebiotic effect in vitro using strains from the Lactobacillus casei/paracasei/rhamnosus group and from Bifidobacterium species. The results revealed that 6 out of 11 L. casei strains and 2 out of 6 L. rhamnosus strains tested were able to ferment Fuc-α1,3-GlcNAc, and L. casei BL87 and L. rhamnosus BL327 strains were also able to ferment Fuc-α1,6-GlcNAc. DNA hybridization experiments suggested that the metabolism of Fuc-α1,3-GlcNAc in those strains relies in an α-L-fucosidase homologous to AlfB. Bifidobacterium breve and Bibidobacterium pseudocatenolatum species also metabolized Fuc-α1,3-GlcNAc. Notably, L-fucose was excreted from all the Lactobacillus and Bifidobacterium strains fermenting fucosyl-disaccharides, except from strains L. rhamnosus BL358 and BL377, indicating that in these latest strains, L-fucose was catabolized. The fucosyl-disaccharides were also tested for their inhibitory potential of pathogen adhesion to human colon adenocarcinoma epithelial (HT29) cell line. Enteropathogenic Escherichia coli (EPEC) strains isolated from infantile gastroenteritis were used, and the results showed that both fucosyl-disaccharides inhibited adhesion to different extents of certain EPEC strains to HT29 cells in tissue culture.

This work is aimed at evaluating the ability of the microalgae to partially substitute biodegradable materials with improved physicochemical properties. To this end, starch films containing the microalga Nannochloropsis gaditana sp. (N. gaditana) have been developed and characterized. Initially, different ultrasound treatments were evaluated for microalgae cell wall disruption. Although surfactant-aided sonication was the most efficient disruption method, the presence of the surfactant hampered continuous film formation. Subsequently, the ability of intact and ultrasound-treated N. gaditana cells to improve barrier properties of starch films was evaluated. Combined small and wide angle X-ray scattering experiments (SAXS/WAXS, respectively) evidenced slight nanostructural and crystallinity changes induced by the presence of N. gaditana cells. Incorporation of intact or ultrasound-treated microalgae into starch led to more hydrophobic films, with enhanced barrier properties for most of the formulations. However, the films containing the microalgae treated with the greatest ultrasound intensity, did not show any barrier improvement due to the plasticization promoted by the cell components (probably lipids) released during the ultrasound treatment, as suggested by SAXS/WAXS and the mechanical properties., Synchrotron experiments were performed at NCD beamline at ALBA Synchrotron with the collaboration of ALBA staff (2016021658 project). M.J. Fabra, Marta Martinez-Sanz and L.G. Gómez-Mascaraque are recipients of a Ramon y Cajal (RYC-2014-158), Juan de la Cierva (IJCI-2015-23389) and predoctoral (call 2013) contracts from the Spanish Ministry of Economy, Industry and Competitiveness, respectively. Authors would also like to thank the Central Support Service for Experimental Research (SCSIE) of the University of Valencia for the electronic microscopy service.

Summary The effect of the addition of different types of inulin (long-chain, native and short-chain inulin) at four concentrations (0, 2.5, 5 and 7.5% w/w) on flow properties of semisolid dairy desserts formulated with either skimmed or whole milk and three modified tapioca starch concentrations (2.5, 3.25 and 4% w/w) was studied. All samples displayed shear-thinning flow behaviour. The addition of the three types of inulin to whole milk samples with low starch concentrations (2.5 and 3.25% w/w) led to combined hysteresis loops: samples showed antithixotropy at low shear rate values, but thixotropy at high shear rate values. The remaining samples showed thixotropic behaviour. The effect of the different types of inulin on the flow properties was not only highly dependent on the inulin concentration but also on milk type and starch concentration. On adding 7.5% of long-chain inulin, which significantly increased consistency index values and significantly decreased flow index values in both types of milk and for all starch concentrations.

Inulin is a natural component of several fruits and vegetables with beneficial effects on health and interesting technological properties, which are linked to its average chain length. The objective of this work was to study how the addition of different types of inulin (average long-chain, native and average short-chain inulin) at four concentrations (0%, 2.5%, 5% and 7.5% w/w) may affect the viscoelastic properties of semisolid dairy systems formulated with either skimmed or whole milk and two modified tapioca starch concentrations (3.25% or 4% w/w). The mechanical spectra obtained for most of the samples were typical of gelled materials. A significant effect of the interactions between the inulin type, the inulin concentration and the starch concentration on the storage and loss moduli (G′and G″), on the loss tangent angle (tan δ) and on the complex viscosity (η*) values at 1 Hz was observed in both whole- and skimmed-milk samples. The effect of the different types of inulin on the viscoelastic properties of the systems was dependent not only on the inulin concentration added to them but also on the sample composition. The most remarkable effects were observed when adding 7.5% of long-chain inulin, which significantly increased G′, G″ and η* values, and significantly decreased tan δ values in both types of milk and both starch concentrations.