Diet influences the functions of the human intestinal microbiome.

Gut microbes programme their metabolism to suit intestinal conditions and convert dietary components into a panel of small molecules that ultimately affect host physiology. To unveil what is behind the effects of key dietary components on microbial functions and the way they modulate host-microbe interaction, we used for the first time a multi-omic approach that goes behind the mere gut phylogenetic composition and provides an overall picture of the functional repertoire in 27 fecal samples from omnivorous, vegan and vegetarian volunteers. Based on our data, vegan and vegetarian diets were associated to the highest abundance of microbial genes/proteins responsible for cell motility, carbohydrate- and protein-hydrolyzing enzymes, transport systems and the synthesis of essential amino acids and vitamins. A positive correlation was observed when intake of fiber and the relative fecal abundance of flagellin were compared. Microbial cells and flagellin extracted from fecal samples of 61 healthy donors modulated the viability of the human (HT29) colon carcinoma cells and the host response through the stimulation of the expression of Toll-like receptor 5, lectin RegIIIα and three interleukins (IL-8, IL-22 and IL-23). Our findings concretize a further and relevant milestone on how the diet may prevent/mitigate disease risk.

Beneficial Plant Microorganisms Affect the Endophytic Bacterial Communities of Durum Wheat Roots as Detected by Different Molecular Approaches.

This study aimed at characterising the endophytic bacterial communities living in durum wheat roots, as affected by wheat cultivar and inoculation of the Arbuscular mycorrhizal fungus Funneliformis mosseae IMA1 and the wheat root endophytic bacterium Lactobacillus plantarum B.MD.R.A2. These microorganisms were inoculated, alone or in combination, in durum wheat (cultivars Odisseo and Saragolla). Non-inoculated plants of each cultivar represented the controls. Forty-three days after sowing, roots were deprived of the epiphytic microbiota and subjected to DNA extraction. The DNA was used as template in PCR-DGGE analysis of the 16S rRNA gene (variable region V3-V5) and 16S (region V1-V3) metagenetics. Odisseo and Saragolla root endophytic bacterial biotas differed for number of OTUs and composition. In detail, Pseudomonas was higher in Odisseo than in Saragolla. The inoculation of F. mosseae and L. plantarum increased the abundance of Pseudomonas, some Actinobacteria (e.g., Streptomyces, Microbacterium, two genera including several plant growth promoting (PGP) strains) and Bacteroidetes in both cultivars. However, the endophytic bacterial biota of Saragolla roots inoculated just with lactobacilli did not differ from that of the control. The inoculation of Saragolla with F. mosseae, alone or in combination with lactobacilli, led to higher abundance of Rhodococcus, belonging to Actinobacteria and encompassing PGP strains. First, this work showed that F. mosseae and L. plantarum shape the endophytic bacterial biota of durum wheat roots. Abundance of some OTUs was affected by the microbial inoculation, depending on the cultivar. This result represents a starting point for exploitation of beneficial endophytes of wheat roots.

Microbial cell-free extracts affect the biochemical characteristics and sensorial quality of sourdough bread.

This study aimed to improve the sensorial quality of sourdough wheat bread by the addition of cell-free enzyme extracts (CFEs) from Lactobacillus sanfranciscensis (SF), Hafnia alvei (HF) and Debaryomyces hansenii (DH). CFEs were suitable sources of peptidases, glutamate dehydrogenase and cystathionine γ-lyase. The concentration of free amino acids (FAA) in the sourdoughs containing CFEs was higher than the control sourdough, produced without addition of CFEs. The community-level catabolic profiles showed that the highest number of carbohydrates, polymers and carboxylic acids were consumed in the SF sourdough. Breads produced with CFEs were characterized by higher specific volume than the control. The use of CFEs impacted on the profile of volatile organic compounds. Overall, positive correlations were found between some key-aroma compounds and enzyme activities/precursor FAA. The SF bread, characterized by highest level of alcohols, received the highest score for aroma and sweetness in the sensory analysis.

Selected Probiotic Lactobacilli Have the Capacity To Hydrolyze Gluten Peptides during Simulated Gastrointestinal Digestion.

The aim of this study was to demonstrate the capacity of probiotic lactobacilli to hydrolyze immunogenic gluten peptides. Eighteen commercial strains of probiotic lactobacilli with highly variable peptidase activity (i.e., aminopeptidase N, iminopeptidase, prolyl endopeptidyl peptidase, tripeptidase, prolidase, prolinase, and dipeptidase), including toward Pro-rich peptides, were tested in this study. Ten probiotic strains were selected on the basis of their specific enzyme activity. When pooled, these 10 strains provided the peptidase portfolio that is required to completely degrade the immunogenic gluten peptides involved in celiac disease (CD). The selected probiotic mixture was able to completely hydrolyze well-known immunogenic epitopes, including the gliadin 33-mer peptide, the peptide spanning residues 57 to 68 of the α9-gliadin (α9-gliadin peptide 57-68), A-gliadin peptide 62-75, and γ-gliadin peptide 62-75. During digestion under simulated gastrointestinal conditions, the pool of 10 selected probiotic lactobacilli strongly hydrolyzed the wheat bread gluten (ca. 18,000 ppm) to less than 10 ppm after 360 min of treatment. As determined by multidimensional chromatography (MDLC) coupled to nanoelectrospray ionization (nano-ESI)-tandem mass spectrometry (MS/MS), no known immunogenic peptides were detected in wheat bread that was digested in the presence of the probiotics. Accordingly, the level of cytokines (interleukin 2 [IL-2], IL-10, and interferon gamma [IFN-γ]) produced by duodenal biopsy specimens from CD patients who consumed wheat bread digested by probiotics was similar to the baseline value (negative control). Probiotics that specifically hydrolyze gluten polypeptides could also be used to hydrolyze immunogenic peptides that contaminate gluten-free products. This could provide a new and safe adjunctive therapy alternative to the gluten-free diet (GFD).IMPORTANCE This study confirmed that probiotic Lactobacillus strains have different enzymatic abilities for hydrolyzing polypeptides, including the Pro-rich epitopes involved in the pathology of CD. Ten lactobacilli with complementary peptidase activities that hydrolyze gluten peptides during simulated gastrointestinal digestion were selected and tested. The results collected showed the potential of probiotic formulas as novel dietary treatments for CD patients.

Salivary and fecal microbiota and metabolome of celiac children under gluten-free diet.

Celiac disease (CD) is an inflammatory autoimmune disorder resulting from the combination of genetic predisposition and gluten ingestion. A life-long gluten free diet (GFD) is the only therapeutic approach. Dysbiosis, which can precede the CD pathogenesis and/or persist when subjects are on GFD, is reviewed and discussed. Salivary microbiota and metabolome differed between healthy and celiac children treated under GFD (T-CD) for at least two years. The type of GFD (African- vs Italian-style) modified the microbiota and metabolome of Saharawi T-CD children. Different studies showed bacterial dysbiosis at duodenal and/or fecal level of patients with active untreated CD (U-CD) and T-CD compared to healthy subjects. The ratio of protective anti-inflammatory bacteria such as Lactobacillus-Bifidobacterium to potentially harmful Bacteroides-Enterobacteriaceae was the lowest in U-CD and T-CD children. In agreement with dysbiosis, serum, fecal and urinary metabolome from U-CD and T-CD patients showed altered levels of free amino acids and volatile organic compounds. However, consensus across studies defining specific bacteria and metabolites in U-CD or T-CD patients is still lacking. Future research efforts are required to determine the relationships between CD and oral and intestinal microbiotas to improve the composition of GFD for restoring the gut dysbiosis as a preventative or therapeutic approach for CD.

Functional proteomics within the genus Lactobacillus.

Lactobacillus are mainly used for the manufacture of fermented dairy, sourdough, meat, and vegetable foods or used as probiotics. Under optimal processing conditions, Lactobacillus strains contribute to food functionality through their enzyme portfolio and the release of metabolites. An extensive genomic diversity analysis was conducted to elucidate the core features of the genus Lactobacillus, and to provide a better comprehension of niche adaptation of the strains. However, proteomics is an indispensable "omics" science to elucidate the proteome diversity, and the mechanisms of regulation and adaptation of Lactobacillus strains. This review focuses on the novel and comprehensive knowledge of functional proteomics and metaproteomics of Lactobacillus species. A large list of proteomic case studies of different Lactobacillus species is provided to illustrate the adaptability of the main metabolic pathways (e.g., carbohydrate transport and metabolism, pyruvate metabolism, proteolytic system, amino acid metabolism, and protein synthesis) to various life conditions. These investigations have highlighted that lactobacilli modulate the level of a complex panel of proteins to growth/survive in different ecological niches. In addition to the general regulation and stress response, specific metabolic pathways can be switched on and off, modifying the behavior of the strains.

Effect of Whole-Grain Barley on the Human Fecal Microbiota and Metabolome.

In this study, we compared the fecal microbiota and metabolomes of 26 healthy subjects before (HS) and after (HSB) 2 months of diet intervention based on the administration of durum wheat flour and whole-grain barley pasta containing the minimum recommended daily intake (3 g) of barley ß-glucans. Metabolically active bacteria were analyzed through pyrosequencing of the 16S rRNA gene and community-level catabolic profiles. Pyrosequencing data showed that levels of Clostridiaceae (Clostridium orbiscindens and Clostridium sp.), Roseburia hominis, and Ruminococcus sp. increased, while levels of other Firmicutes and Fusobacteria decreased, from the HSB samples to the HS fecal samples. Community-level catabolic profiles were lower in HSB samples. Compared to the results for HS samples, cultivable lactobacilli increased in HSB fecal samples, while the numbers of Enterobacteriaceae, total coliforms, and Bacteroides, Porphyromonas, Prevotella, Pseudomonas, Alcaligenes, and Aeromonas bacteria decreased. Metabolome analyses were performed using an amino acid analyzer and gas chromatography-mass spectrometry solid-phase microextraction. A marked increase in short-chain fatty acids (SCFA), such as 2-methyl-propanoic, acetic, butyric, and propionic acids, was found in HSB samples with respect to the HS fecal samples. Durum wheat flour and whole-grain barley pasta containing 3% barley ß-glucans appeared to be effective in modulating the composition and metabolic pathways of the intestinal microbiota, leading to an increased level of SCFA in the HSB samples.