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Purpose Plasma trimethylamine-N-oxide (TMAO) levels have been shown to correlate with increased risk of metabolic diseases including cardiovascular diseases. TMAO exposure predominantly occurs as a consequence of gut microbiota-dependent trimethylamine (TMA) production from dietary substrates including choline, carnitine and betaine, which is then converted to TMAO in the liver. Reducing microbial TMA production is likely to be the most effective and sustainable approach to overcoming TMAO burden in humans. Current models for studying microbial TMA production have numerous weaknesses including the cost and length of human studies, differences in TMA(O) metabolism in animal models and the risk of failing to replicate multi-enzyme/multi-strain pathways when using isolated bacterial strains. The purpose of this research was to investigate TMA production from dietary precursors in an in-vitro model of the human colon. Methods TMA production from choline, l-carnitine, betaine and γ-butyrobetaine was studied over 24–48 h using an in-vitro human colon model with metabolite quantification performed using LC–MS. Results Choline was metabolised via the direct choline TMA-lyase route but not the indirect choline–betaine-TMA route, conversion of l-carnitine to TMA was slower than that of choline and involves the formation of the intermediate γ-BB, whereas the Rieske-type monooxygenase/reductase pathway for l-carnitine metabolism to TMA was negligible. The rate of TMA production from precursors was choline > carnitine > betaine > γ-BB. 3,3-Dimethyl-1-butanol (DMB) had no effect on the conversion of choline to TMA. Conclusion The metabolic routes for microbial TMA production in the colon model are consistent with observations from human studies. Thus, this model is suitable for studying gut microbiota metabolism of TMA and for screening potential therapeutic targets that aim to attenuate TMA production by the gut microbiota. Trial registration number NCT02653001 (http://www.clinicaltrials.gov), registered 12 Jan 2016.

Background New developments in next-generation sequencing technologies and massive data received from this approach open wide prospects for personalised medicine and nutrition studies. Metagenomic analysis of the gut microbiota is paramount for the characterization of human health and wellbeing. Despite the intensive research, there is a huge gap and inconsistency between different studies due to the non-standardised and biased pipeline. Methodical and systemic understanding of every stage in the process is necessary to overcome all bottlenecks and grey zones of gut microbiota studies, where all details and interactions between processes are important. Results Here we show that an inexpensive, but reliable iSeq 100 platform is an excellent tool to perform the analysis of the human gut microbiota by amplicon sequencing of the 16 S rRNA gene. Two commercial DNA extraction kits and different starting materials performed similarly regarding the taxonomic distribution of identified bacteria. DNA/RNA Shield reagent proved to be a reliable solution for stool samples collection, preservation, and storage, as the storage of faecal material in DNA/RNA Shield for three weeks at different temperatures and thawing cycles had a low impact on the bacterial distribution. Conclusions Altogether, a thoroughly elaborated pipeline with close attention to details ensures high reproducibility with significant biological but not technical variations.

Epidemiological and clinical evidence highlight the benefit of dietary fibre consumption on body weight. This benefit is partly attributed to the interaction of dietary fibre with the gut microbiota. Dietary fibre possesses a complex food structure which resists digestion in the upper gut and therefore reaches the distal gut where it becomes available for bacterial fermentation. This process yields short chain fatty acids (SCFAs) which stimulate the release of appetite suppressing hormones Glucagon-like peptide-1 (GLP-1) and peptide YY (PYY). Food structures can further enhance the delivery of fermentable substrates to the distal gut by protecting the intracellular nutrients during upper gastro intestinal digestion. Domestic and industrial processing can disturb these food structures that act like barriers towards digestive enzymes. This leads to more digestible products that are better absorbed in the upper gut. As a result, less resistant material (fibre) and intracellular nutrients may reach the distal gut, thus reducing substrates for bacterial fermentation and its subsequent benefits on the host metabolism including appetite suppression. Understanding this link is essential for the design of diets and food products that can promote appetite suppression and act as a successful strategy towards obesity management. This article reviews the current evidence in the interplay between food structure, bacterial fermentation and appetite control.

National screening programs use dried blood specimens to detect metabolic disorders or aberrant protein functions that are not clinically evident in the neonatal period. Similarly, gut microbiota metabolites and immunological acute-phase proteins may reveal latent immune aberrations. Microbial metabolites interact with xenobiotic receptors (i.e., aryl hydrocarbon and pregnane-X) to maintain gastrointestinal tissue health, supported by acute-phase proteins, functioning as sensors of microbial immunomodulation and homeostasis. The delivery (vaginal or cesarean section) shapes the microbial colonization, which substantially modulates both the immune system’s response and mucosal homeostasis. This study profiled microbial metabolites of the kynurenine and tryptophan pathway and acute-phase proteins in 134 neonatal dried blood specimens. We newly established neonatal blood levels of microbial xenobiotic receptors ligands (i.e., indole-3-aldehyde, indole-3-butyric acid, and indole-3-acetamide) on the second day of life. Furthermore, we observed diverse microbial metabolic profiles in neonates born vaginally and via cesarean section, potentially due to microbial immunomodulatory influence. In summary, these findings suggest the supportive role of human gut microbiota in developing and maintaining immune system homeostasis.

Bifidobacterium longum subsp. longum is a prevalent group in the human gut microbiome. Its persistence in the intestinal microbial community suggests a close host-microbe relationship according to age. The subspecies adaptations are related to metabolic capabilities and genomic and functional diversity. In this study, 154 genomes from public databases and four new Chilean isolates were genomically compared through an in silico approach to identify genomic divergence in genes associated with carbohydrate consumption and their possible adaptations to different human intestinal niches. The pangenome of the subspecies was open, which correlates with its remarkable ability to colonize several niches. The new genomes homogenously clustered within subspecies longum, as observed in phylogenetic analysis. B. longum SC664 was different at the sequence level but not in its functions. COG analysis revealed that carbohydrate use is variable among longum subspecies. Glycosyl hydrolases participating in human milk oligosaccharide use were found in certain infant and adult genomes. Predictive genomic analysis revealed that B. longum M12 contained an HMO cluster associated with the use of fucosylated HMOs but only endowed with a GH95, being able to grow in 2-fucosyllactose as the sole carbon source. This study identifies novel genomes with distinct adaptations to HMOs and highlights the plasticity of B. longum subsp. longum to colonize the human gut microbiota.

How to study the unculturable bacteria in the laboratory is one of the major challenges in human gut microbiota research. The resulting lack of microbiology knowledge of this “dark matter” greatly hinders further understanding of our gut microbiota. Here, to characterize the in vivo growth and division of human gut bacteria, we report the integrative use of STAMP (sequential tagging with D-amino acid–based metabolic probes) and fluorescence in situ hybridization (FISH) in a human microbiota-associated mouse model. After stable colonization of the human fecal microbiotas in germ-free mice, two fluorescent D-amino acid probes were sequentially administered by gavage, and the dually labeled peptidoglycan of the bacteria provided a chronological recording of their cell wall syntheses. Following taxonomic identification with FISH staining, the growth patterns of 32 species, including 5 currently unculturables, were identified. Surprisingly, we found that many bacterial species in the human microbiota were significantly shorter than those in the mouse gut microbiota. An imaging database for gut bacteria – Microbiome Atlas was built for summarizing STAMP imaging of bacteria from different microbiotas, which can be contributed by the microbiota research community worldwide. This integrative imaging strategy and the database will promote our understanding of the bacterial cytology in gut microbiotas and facilitate communications among cellular microbiologists.

Although the vast majority of women encounters at least one vaginal infection during their life, the amount of microbiome-related research performed in this area lags behind compared to alternative niches such as the intestinal tract. As a result, effective means of diagnosis and treatment, especially of recurrent infections, are limited. The role of the metabolome in vaginal health is largely elusive. It has been shown that lactate produced by the numerous lactobacilli present promotes health by limiting the chance of infection. Short chain fatty acids (SCFA) have been mainly linked to dysbiosis, although the causality of this relationship is still under debate. In this review, we aim to bring together information on the role of the vaginal metabolome and microbiome in infections caused by Candida. Vulvovaginal candidiasis affects near to 70% of all women at least once in their life with a significant proportion of women suffering from the recurrent variant. We assess the role of fatty acid metabolites, mainly SCFA and lactate, in onset of infection and virulence of the fungal pathogen. In addition, we pinpoint where lack of research limits our understanding of the molecular processes involved and restricts the possibility of developing novel treatment strategies. ispartof: FRONTIERS IN MICROBIOLOGY vol:12 ispartof: location:Switzerland status: published

Background Culturomics can ascertain traces of microorganisms to be cultivated using different strategies and identified by matrix-assisted laser desorption/ionization–time-of-flight mass spectrometry or 16S rDNA sequencing. However, to cater to all requirements of microorganisms and isolate as many species as possible, multiple culture conditions must be used, imposing a heavy workload. In addition, the fast-growing bacteria (e.g., Escherichia) surpass the slow-growing bacteria in culture by occupying space and using up nutrients. Besides, some bacteria (e.g., Pseudomonas) suppress others by secreting antibacterial metabolites, making it difficult to isolate bacteria with lower competence. Applying inhibitors to restrain fast-growing bacteria is one method to cultivate more bacterial species from human feces. Results We applied CHIR-090, an LpxC enzyme inhibitor that has antibacterial activity against most Gram-negative bacteria, to culturomics of human fresh feces. The antibacterial activity of CHIR-090 was first assessed on five Gram-negative species of bacteria (Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Proteus vulgaris, and Bacteroides vulgatus), all of which are commonly isolated from the human gut. Then, we assessed suitable concentrations of the inhibitor. Finally, CHIR-090 was applied in blood culture bottles for bacterial cultivation. In total, 102 species from five samples were identified. Of these, we found one new species, two species not reported previously in the human gut, and 11 species not previously isolated from humans. Conclusions CHIR-090 can suppress E. coli, P. aeruginosa, K. pneumoniae, Pro. vulgaris, but not B. vulgatus. Compared with the non-inhibitor group, CHIR-090 increased bacteria isolation by 23.50%, including four species not reported in humans and one new species. Application of LpxC enzyme inhibitor in culturomics increased the number of species isolated from the human gut.

Efficient capture of glycans, the prime metabolic resources in the human gut, confers a key competitive advantage for gut microbiota members equipped with extracellular glycoside hydrolases (GHs) to target these substrates. The association of glycans to the bacterial cell surface is typically mediated by carbohydrate binding modules (CBMs). Here, we report the structure of RiCBM86 appended to a GH family 10 xylanase from Roseburia intestinalis. This CBM represents a new family of xylan binding CBMs present in xylanases from abundant and prevalent healthy human gut Clostridiales. RiCBM86 adopts a canonical β-sandwich fold, but shows structural divergence from known CBMs. The structure of RiCBM86 has been determined with a bound xylohexaose, which revealed an open and shallow binding site. RiCBM86 recognizes only a single xylosyl ring with direct hydrogen bonds. This mode of recognition is unprecedented amongst previously reported xylan binding type-B CBMs that display more extensive hydrogen-bonding patterns to their ligands or employ Ca2+ to mediate ligand-binding. The architecture of RiCBM86 is consistent with an atypically low binding affinity (KD about 0.5 mm for xylohexaose) compared to most xylan binding CBMs. Analyses using NMR spectroscopy corroborated the observations from the complex structure and the preference of RiCBM86 to arabinoxylan over glucuronoxylan, consistent with the largely negatively charged surface flanking the binding site. Mutational analysis and affinity electrophoresis established the importance of key binding residues, which are conserved in the family. This study provides novel insight into the structural features that shape low-affinity CBMs that mediate extended bacterial glycan capture in the human gut niche. Databases: Structural data are available in the protein data bank database under the accession number 6SGF. Sequence data are available in the GenBank database under the accession number EEV01588.1. The assignment of the Roseburia intestinalis xylan binding module into the CBM86 new family is available in the CAZy database (http://www.cazy.org/CBM86.html).

The human gut can be viewed as a flow-through system with a short residence time, a high turnover rate and a spatial gradient of physiological conditions. As a consequence, the gut microbiota is exposed to highly fluctuating environmental determinants presented by the host and diet. Here, we assessed the fermentation and colonisation of insoluble wheat bran by faecal microbiota of three individuals at an unprecedented sampling intensity. Time-resolved 16S rRNA gene amplicon sequencing, revealed a dynamic microbial community, characterised by abrupt shifts in composition, delimiting states with a more constant community, giving rise to a succession of bacterial taxa alternately dominating the community over a 72 h timespan. Early stages were dominated by Enterobacteriaceae and Fusobacterium species, growing on the carbohydrate-low, protein rich medium to which wheat bran was supplemented. The onset of wheat bran fermentation, marked by a spike in short chain fatty acid production with an increasing butyrate proportion and an increased endo-1,4-β-xylanase activity, corresponded to donor-dependent proportional increases of Bacteroides ovatus/stercoris, Prevotella copri and Firmicutes species, which were strongly enriched in the bran-attached community. Literature and database searches provided novel insights into the metabolic and growth characteristics underlying the observed succession and colonisation, illustrating the potency of a time-resolved analysis to increase our understanding of gut microbiota dynamics upon dietary modulations.

To investigate the relationship between intestinal microbiota and SARS-CoV-2-mediated pathogenicity in a United States, majority African American cohort. We prospectively collected fecal samples from 50 SARS-CoV-2 infected patients, 9 SARS-CoV-2 recovered patients, and 34 uninfected subjects seen by the hospital with unrelated respiratory medical conditions (controls). 16S rRNA sequencing and qPCR analysis was performed on fecal DNA/RNA. The fecal microbial composition was found to be significantly different between SARS-CoV-2 patients and controls (PERMANOVA FDR-P = .004), independent of antibiotic exposure. Peptoniphilus, Corynebacterium and Campylobacter were identified as the three most significantly enriched genera in COVID-19 patients compared to controls. Actively infected patients were also found to have a different gut microbiota than recovered patients (PERMANOVA FDR-P = .003), and the most enriched genus in infected patients was Campylobacter, with Agathobacter and Faecalibacterium being enriched in the recovered patients. No difference in microbial community structure between recovered patients and uninfected controls was observed, nor a difference in alpha diversity between the three groups. 24 of the 50 COVID-19 patients (48%) tested positive via RT-qPCR for fecal SARS-CoV-2 RNA. A significant difference in gut microbial composition between SARS-CoV-2 positive and negative samples was observed, with Klebsiella and Agathobacter being enriched in the positive cohort. No significant associations between microbiome composition and disease severity was found. The intestinal microbiota is sensitive to the presence of SARS-CoV-2, with increased relative abundance of genera (Campylobacter, Klebsiella) associated with gastrointestinal (GI) disease. Further studies are needed to investigate the functional impact of SARS-CoV-2 on GI health.

The human gastrointestinal tract is colonized by a diverse microbial community, which plays a crucial role in human health. In the gut, a protective mucus layer that consists of glycan structures separates the bacteria from the host epithelial cells. These host-derived glycans are utilized by bacteria that have adapted to this specific compound in the gastrointestinal tract. Our study investigated the close interaction between two distinct gut microbiota members known to use mucus glycans, the generalist Bacteroides thetaiotaomicron and the specialist Akkermansia muciniphila in vitro and in vivo. The in vitro study, in which mucin was the only nutrient source, indicated that B. thetaiotaomicron significantly upregulated genes coding for Glycoside Hydrolases (GHs) and mucin degradation activity when cultured in the presence of A. muciniphila. Furthermore, B. thetaiotaomicron significantly upregulated the expression of a gene encoding for membrane attack complex/perforin (MACPF) domain in co-culture. The transcriptome analysis also indicated that A. muciniphila was less affected by the environmental changes and was able to sustain its abundance in the presence of B. thetaiotaomicron while increasing the expression of LPS core biosynthesis activity encoding genes (O-antigen ligase, Lipid A and Glycosyl transferases) as well as ABC transporters. Using germ-free mice colonized with B. thetaiotaomicron and/or A. muciniphila, we observed a more general glycan degrading profile in B. thetaiotaomicron while the expression profile of A. muciniphila was not significantly affected when colonizing together, indicating that two different nutritional niches were established in mice gut. Thus, our results indicate that a mucin degrading generalist adapts to its changing environment, depending on available carbohydrates while a mucin degrading specialist adapts by coping with competing microorganism through upregulation of defense related genes.

β-Mannans are hemicelluloses that are abundant in modern diets as components in seed endosperms and common additives in processed food. Currently, the collective understanding of β-mannan saccharification in the human colon is limited to a few keystone species, which presumably liberate low-molecular-weight mannooligosaccharide fragments that become directly available to the surrounding microbial community. Here, we show that a dominant butyrate producer in the human gut, Faecalibacterium prausnitzii, is able to acquire and degrade various β-mannooligosaccharides (β-MOS), which are derived by the primary mannanolytic activity of neighboring gut microbiota. Detailed biochemical analyses of selected protein components from their two β-MOS utilization loci (F. prausnitzii β-MOS utilization loci [FpMULs]) supported a concerted model whereby the imported β-MOS are stepwise disassembled intracellularly by highly adapted enzymes. Coculturing experiments of F. prausnitzii with the primary degraders Bacteroides ovatus and Roseburia intestinalis on polymeric β-mannan resulted in syntrophic growth, thus confirming the high efficiency of the FpMULs' uptake system. Genomic comparison with human F. prausnitzii strains and analyses of 2,441 public human metagenomes revealed that FpMULs are highly conserved and distributed worldwide. Together, our results provide a significant advance in the knowledge of β-mannan metabolism and the degree to which its degradation is mediated by cross-feeding interactions between prominent beneficial microbes in the human gut. IMPORTANCE Commensal butyrate-producing bacteria belonging to the Firmicutes phylum are abundant in the human gut and are crucial for maintaining health. Currently, insight is lacking into how they target otherwise indigestible dietary fibers and into the trophic interactions they establish with other glycan degraders in the competitive gut environment. By combining cultivation, genomic, and detailed biochemical analyses, this work reveals the mechanism enabling F. prausnitzii, as a model Ruminococcaceae within Firmicutes, to cross-feed and access β-mannan-derived oligosaccharides released in the gut ecosystem by the action of primary degraders. A comprehensive survey of human gut metagenomes shows that FpMULs are ubiquitous in human populations globally, highlighting the importance of microbial metabolism of β-mannans/β-MOS as a common dietary component. Our findings provide a mechanistic understanding of the β-MOS utilization capability by F. prausnitzii that may be exploited to select dietary formulations specifically boosting this beneficial symbiont, and thus butyrate production, in the gut.

BackgroundAcetate is a short-chain fatty acid (SCFA) produced by gut bacteria, which has been implicated in cardio-metabolic health. Here we examine the relationships of circulating acetate levels with gut microbiome composition and diversity and with visceral fat in a large population-based cohort.ResultsMicrobiome alpha-diversity was positively correlated with circulating acetate levels (Shannon, Beta [95%CI] = 0.12 [0.06, 0.18], P = 0.002) after adjustment for covariates. Six serum acetate-associated bacterial genera were also identified, including positive correlations with Coprococcus, Barnesiella, Ruminococcus, and Ruminococcaceae NK4A21 and negative correlations were observed with Lachnoclostridium and Bacteroides. We also identified a correlation between visceral fat and serum acetate levels (Beta [95%CI] = −0.07 [−0.11, −0.04], P = 2.8 × 10–4) and between visceral fat and Lachnoclostridium (Beta [95%CI] = 0.076 [0.042, 0.11], P = 1.44 × 10–5). Formal mediation analysis revealed that acetate mediates ∼10% of the total effect of Lachnoclostridium on visceral fat. The taxonomic diversity showed that Lachnoclostridium and Coprococcus comprise at least 18 and 9 species, respectively, including novel bacterial species. By predicting the functional capabilities, we found that Coprococcus spp. present pathways involved in acetate production and metabolism of vitamins B, whereas we identified pathways related to the biosynthesis of trimethylamine (TMA) and CDP-diacylglycerol in Lachnoclostridium spp.ConclusionsOur data indicates that gut microbiota composition and diversity may influence circulating acetate levels and that acetate might exert benefits on certain cardio-metabolic disease risk by decreasing visceral fat. Coprococcus may play an important role in host health by its production of vitamins B and SCFAs, whereas Lachnoclostridium might have an opposing effect by influencing negatively the circulating levels of acetate and being involved in the biosynthesis of detrimental lipid compounds.

This work was supported by CONICYT-Chile through the FONDECYT [n° 3160525] (DM) and by Universidad San Sebastián, and by grants to AM from the Spanish Ministry of Economy and Competitiveness (projects SAF2012-31187, SAF2013-49788-EXP, SAF2015-65878-R), Carlos III Health Institute (Projects PIE14/00045 and AC15/00022), Generalitat Valenciana (Project PrometeoII/2014/065 and Prometeo/2018/A/133), Asociación Española Contra el Cancer (Project AECC 2017-1485) and co-financed by the European Regional Development Fund (ERDF).

Fructoselysine is formed upon heating during processing of food products, and being a key intermediate in advanced glycation end product formation considered to be potentially hazardous to human health. Human gut microbes can degrade fructoselysine to yield the short chain fatty acid butyrate. However, quantitative information on these biochemical reactions is lacking, and interindividual differences therein are not well established. Anaerobic incubations with pooled and individual human fecal slurries were optimized and applied to derive quantitative kinetic information for these biochemical reactions. Of 16 individuals tested, 11 were fructoselysine metabolizers, with Vmax, Km and kcat-values varying up to 14.6-fold, 9.5-fold, and 4.4-fold, respectively. Following fructoselysine exposure, 10 of these 11 metabolizers produced significantly increased butyrate concentrations, varying up to 8.6-fold. Bacterial taxonomic profiling of the fecal samples revealed differential abundant taxa for these reactions (e.g. families Ruminococcaceae, Christenellaceae), and Ruminococcus_1 showed the strongest correlation with fructoselysine degradation and butyrate production (ρ ≥ 0.8). This study highlights substantial interindividual differences in gut microbial degradation of fructoselysine. The presented method allows for quantification of gut microbial degradation kinetics for foodborne xenobiotics, and interindividual differences therein, which can be used to refine prediction of internal exposure.

Wheat bran is a cereal rich in dietary fibers that have high levels of ferulic acid, which has prebiotic effects on the intestinal microbiota and the host. Herein we explored the effect of xylooligosaccharide, xylan, and whole wheat bran on the human gut bacteria and screened for potential ferulic acid esterase genes. Using in vitro fermentation, we analyzed the air pressure, pH-value, and short-chain fatty acid levels. We also performed 16S rRNA gene and metagenomic sequencing. A Venn diagram analysis revealed that 80% of the core operational taxonomic units (OTUs) were shared among the samples, and most of the xylooligosaccharide treatment core OTUs (319/333 OTUs) were shared with the other two treatments’ core OTUs. A significant difference analysis revealed that the relative abundance of Dorea, Bilophila, and Sulfurovum in wheat bran treatment was higher than that in xylan and xylooligosaccharide treatments. The clusters of orthologous groups of proteins functional composition of all samples was similar to the microbiota composition of the control. Using metagenomic sequencing, we revealed seven genes containing the conserved residues, Gly-X-Ser-X-Gly, and the catalytic triad, Ser-His-Asp, which are thus potential ferulic acid esterase genes. All the results indicate that xylan and/or xylooligosaccharide, the main dietary fibers in wheat bran, plays a major role in in vitro fermentation by the human gut microbiota.

The use of additives in food products has become an important public health concern. In recent reports, dietary emulsifiers have been shown to affect the gut microbiota, contributing to a pro-inflammatory phenotype and metabolic syndrome. So far, it is not yet known whether similar microbiome shifts are observable for a more diverse set of emulsifier types and to what extent these effects vary with the unique features of an individual’s microbiome.To bridge this gap, we investigated the effect of five dietary emulsifiers on the fecal microbiota from 10 human individuals upon a 48 hour exposure. Community structure was assessed with quantative microbial profiling, functionality was evaluated by measuring fermentation metabolites and pro-inflammatory properties were assessed with the phylogenetic prediction algorythm PICRUSt, together with a TLR5 reporter cell assay for flagellin. A comparison was made between two mainstream chemical emulsifiers (carboxymethylcellulose and P80), a natural extract (soy lecithin) and biotechnological emulsifiers (sophorolipids and rhamnolipids).While fecal microbiota responded in a donor-dependent manner to the different emulsifiers, profound differences between emulsifier were observed. Rhamnolipids, sophorolipids and soy lecithin eliminated 91% ± 0%, 89% ± 1% and 87% ± 1% of the viable bacterial population after 48 hours, yet they all selectively increased the proportional abundance of putative pathogens. Moreover, profound shifts in butyrate (−96% ± 6 %, −73% ± 24% and −34 ± 25% respectively) and propionate (+13% ± 24 %, +88% ± 50% and +29% ± 16% respectively) production were observed for these emulsifiers. Phylogenetic prediction indicated higher motility, which was, however, not confirmed by increased flagellin levels using the TLR5 reporter cell assay.We conclude that dietary emulsifiers can severely impact the gut microbiota and this seems to be proportional to their emulsifying strength, rather than emulsifier type or origin. As biotechnological emulsifiers were especially more impactful than chemical emulsifiers, caution is warranted when considering them as more natural alternatives for clean label strategies.

The link between the gut microbiota and metabolic syndrome (MetS) has attracted widespread attention. Christensenellaceae was recently described as an important player in human health, while its distribution and relationship with MetS in Chinese population is still unknown. This study sought to observe the association between Christensenellaceae and metabolic indexes in a large sample of residents in South China. A total of 4,781 people from the GGMP project were included, and the fecal microbiota composition of these individuals was characterized by 16S rRNA sequencing and analyzed the relation between Christensenellaceae and metabolism using QIIME (Quantitative Insight Into Microbial Ecology, Version 1.9.1). The results demonstrated that microbial richness and diversity were increased in the group with a high abundance of Christensenellaceae, who showed a greater complexity of the co-occurrence network with other bacteria than residents who lacked Christensenellaceae. The enriched bacterial taxa were predominantly represented by Oscillospira, Ruminococcaceae, RF39, Rikenellaceae and Akkermansia as the Christensenellaceae abundance increased, while the abundances of Veillonella, Fusobacterium and Klebsiella were significantly reduced. Furthermore, Christensenellaceae was negatively correlated with the pathological features of MetS, such as obesity, hypertriglyceridemia and body mass index (BMI). We found reduced levels of lipid biosynthesis and energy metabolism pathways in people with a high abundance of Christensenellaceae, which may explain the negative relationship between body weight and Christensenellaceae. In conclusion, we found a negative correlation between Christensenellaceae and MetS in a large Chinese population and reported the geographical distribution of Christensenellaceae in the GGMP study. The association data from this population-level research support the investigation of strains within Christensenellaceae as potentially beneficial gut microbes.

Microbes in the intestinal tract have a strong influence on human health. Their fermentation of dietary nondigestible carbohydrates leads to the formation of health-promoting short-chain fatty acids, including butyrate, which is the main fuel for the colonic wall and has anticarcinogenic and anti-inflammatory properties. A good understanding of the growth requirements of butyrate-producing bacteria is important for the development of efficient strategies to promote these microbes in the gut, especially in cases where their abundance is altered. The demonstration of the inability of several dominant butyrate producers to grow in the absence of certain vitamins confirms the results of previous in silico analyses. Furthermore, establishing that strains prototrophic for thiamine or folate (butyrate producers and non-butyrate producers) were able to stimulate growth and affect the composition of auxotrophic synthetic communities suggests that the provision of prototrophic bacteria that are efficient cross feeders may stimulate butyrate-producing bacteria under certain in vivo conditions., We investigated the requirement of 15 human butyrate-producing gut bacterial strains for eight B vitamins and the proteinogenic amino acids by a combination of genome sequence analysis and in vitro growth experiments. The Ruminococcaceae species Faecalibacterium prausnitzii and Subdoligranulum variabile were auxotrophic for most of the vitamins and the amino acid tryptophan. Within the Lachnospiraceae, most species were prototrophic for all amino acids and several vitamins, but biotin auxotrophy was widespread. In addition, most of the strains belonging to Eubacterium rectale and Roseburia spp., but few of the other Lachnospiraceae strains, were auxotrophic for thiamine and folate. Synthetic coculture experiments of five thiamine or folate auxotrophic strains with different prototrophic bacteria in the absence and presence of different vitamin concentrations were carried out. This demonstrated that cross-feeding between bacteria does take place and revealed differences in cross-feeding efficiency between prototrophic strains. Vitamin-independent growth stimulation in coculture compared to monococulture was also observed, in particular for F. prausnitzii A2-165, suggesting that it benefits from the provision of other growth factors from community members. The presence of multiple vitamin auxotrophies in the most abundant butyrate-producing Firmicutes species found in the healthy human colon indicates that these bacteria depend upon vitamins supplied from the diet or via cross-feeding from other members of the microbial community.

Our study aim is to investigate the role of Intestinimonas, Nε-fructosyllysine (FL)-degrading bacterium, in infants and adults. We used lysine and subsequently FL in anaerobic serial dilutions of stools of infants and adults to enrich lysine and FL-degrading species. The fecal microbiota of adults were able to ferment lysine and FL to butyrate. Different groups of Intestinimonas spp. were detected from all lysine enrichments whereas the FL enrichments consisted of broader taxonomic groups with a reduced abundance of Intestinimonas-related species. Remarkably, the capability to degrade FL was only observed in formula-fed but not in breast-fed infants, which may relate to high contents of FL in formulas after thermal treatment. This possibility was supported by analyzing metagenomic datasets of 3-month and 4-month infants. Our data indicate the key role of Intestinimonas-like bacteria in FL degradation in formula-fed infants and adults as a profound example of adaptation of intestinal bacteria to dietary components.

Ginger, a widely used functional food and food additive, little is known about the effect of ginger juice, which is rich in many healthful agents, on healthy humans or on its relationship with gut microbiota composition variation. The aim of this study was to investigate the changes in the gut microbial communities that occur following the supplementation of fresh ginger-derived juice in healthy adults and its potential associations with function. A crossover intervention study in which 123 healthy subjects (63 men and 60 women) consumed fresh ginger juice from Zingiber officinale Rosc. or sterile 0.9% sodium chloride was conducted. 16S rRNA sequencing analyses were applied to characterize gut microbiota variation. We found that ginger juice intervention increased the species number of intestinal flora. A decreased relative abundance of the Prevotella-to-Bacteroides ratio and pro-inflammatory Ruminococcus_1 and Ruminococcus_2 while a tendency toward an increased Firmicutes-to-Bacteroidetes ratio, Proteobacteria and anti-inflammatory Faecalibacterium were found. When we did not consider gender, we found differences in bacterial diversity both in community evenness and in richness caused by ginger intervention. In fact, there were different changes in bacterial α-diversity induced by the ginger juice in men and women. We identified 19 bacterial genera with significant differences between the control group (women) and ginger group (women) and 15 significant differences between the control group (men) and ginger group (men) at the genus level. Our results showed that short-term intake of ginger juice had substantial effects on the composition and function of gut microbiota in healthy people. Moreover, our findings underscored the importance of analyzing both male and female individuals to investigate the effects of ginger on gut microbiota. Additional studies are necessary to confirm these findings.

Zoonotic Campylobacter, including C. jejuni and C. coli, are among the most prevalent agents of food-borne enteritis worldwide. The immunopathological sequelae of campylobacteriosis are caused by Toll-like Receptor-4 (TLR4)-dependent host immune responses, induced by bacterial lipooligosaccharide (LOS). In order to investigate C. coli-host interactions, including the roles of the human gut microbiota and TLR4, upon infection, we applied a clinical acute campylobacteriosis model, and subjected secondary abiotic, TLR4-deficient IL10-/- mice and IL10-/- controls to fecal microbiota transplantation derived from human donors by gavage, before peroral C. coli challenge. Until day 21 post-infection, C. coli could stably colonize the gastrointestinal tract of human microbiota-associated (hma) mice of either genotype. TLR4-deficient IL10-/- mice, however, displayed less severe clinical signs of infection, that were accompanied by less distinct apoptotic epithelial cell and innate as well as adaptive immune cell responses in the colon, as compared to IL10-/- counterparts. Furthermore, C. coli infected IL10-/-, as opposed to TLR4-deficient IL10-/-, mice displayed increased pro-inflammatory cytokine concentrations in intestinal and, strikingly, systemic compartments. We conclude that pathogenic LOS might play an important role in inducing TLR4-dependent host immune responses upon C. coli infection, which needs to be further addressed in more detail.

In this work, we investigate differences in gut microbial diversity driven by drug use or by the widely used methods for drug cessation: methadone maintenance treatment (MMT) and compulsory detention (CD). Methods: 99 participants (28 CD participants, 16 MMT patients, 27 drug users, and 28 healthy controls) were selected using strict inclusion criteria. Nutritional intake and gut microbial diversity were analyzed with bioinformatics tools and SPSS 20.0. Results: Alpha diversity was not significantly different among groups, whereas beta diversity of gut microbiota and nutrient intake were significantly higher among MMT patients. Taxa were unevenly distributed between groups, with drug users having the highest proportion of Ruminococcus and MMT patients having the highest abundance of Bifidobacterium and Lactobacillus. Conclusion: Drug use, cessation method, and diet contribute to shaping human gut communities. High beta diversity among MMT patients is likely driven by methadone use and high nutrient intake, leading to increased orexin A and enrichment for beneficial bacteria, while diversity in CD participants is largely influenced by diet.

Strains Marseille-P4001 and Marseille-P3668 are new species from the order Bacteroidales isolated from healthy French volunteers. They are anaerobic Gram-negative rod-shaped bacteria. They exhibited 92.68% and 96.68% 16S rRNA sequence identities with Parabacteroides gordonii strain MS-1 and Parabacteroides chinchillae JCM 17104, respectively, the phylogenetically closest species. Their respective draft genomes measured 5.23 Mb and 3.73 Mb with 39.2 mol% and 40.8 mol% of G + C content. Using a taxonogenomics method, we propose here a brief description of Parabacteroides pacaensis sp. nov., strain Marseille-P4001T and Parabacteroides provencensis sp. nov., strain Marseille-P3668T as new bacterial species. Keywords: Culturomics, human gut microbiota, new bacteria, Parabacteroides pacaensis sp. nov., Parabacteroides provencensis sp. nov

International audience; A Gram-negative and facultative anaerobic bacterium, designated strain SN4T, was isolated from the stool sample of an obese Amazonian patient. The new isolate was characterized by the taxonogenomics approach. The strain SN4T was beige-colored, circular and not haemolytic. Cells are rod shaped and motile with several flagella. Strain SN4T grows optimally at pH 7 and can survive in the presence of a saline concentration of up to 75 g/l NaCl. The 16S ribosomal RNA gene sequence analysis of the novel strain SN4T showed 95.28% similarity in nucleotide sequence with Gorillibacterium massiliense G5T, the phylogenetically closest neighbor and the type species of this genus. Anteiso-C15:0, iso-C15:0 and C16:0 were found as the major components in the cellular fatty acid analysis of this isolate. The genomic draft of strain SN4T is 5,263,742 bp long with 53.33% of G+C content. The differences in physiological, biochemical characteristics and phylogenetic and genomic data make it possible to clearly distinguish the strain SN4T from G. massiliense G5T. Based on the taxonogenomic description and the phenotypic and biochemical characteristics of this bacterium presented in this article, we propose the SN4T strain (= CSUR P2011 = DSM 100,698) as a new species, Gorillibacterium timonense sp. nov.

Environmental health is increasingly compromised by persistent toxic substances, which may have serious implications in food safety and, thus, in human health. Polybrominated diphenyl ethers (PBDEs) are anthropogenic contaminants with endocrine disruption abilities and are commonly found in seafood, the main route of human exposure. Growing evidence points out that the human gut microbiota interacts with xenobiotics, which may lead to impairment of host homeostasis if functions of microbiota become compromised. The aim of this study was to ascertain if the physiological balance of human gut microbiome is affected by the presence and degree of exposure to PBDEs. Fermentation was performed in a batch closed-system using an inoculum made from fresh human stool. The volatolomic profile was analysed by solid-phase microextraction coupled to gas chromatography-mass spectrometry. Mesophilic, Gram-negative bacteria and coliforms were quantified by classic plating methods. Changes in the gut microbiome were evaluated after DNA extraction followed by deep sequencing of the 16S rDNA region. The exposure to PBDEs resulted in an imbalance in sulfur, short-chain fatty acids and aromatic organic compounds, changing the microbial volatolome in a dose- and time-dependent manner. Slight deviations in the microbial structure of human gut occurred in the presence of PBDEs, especially for high doses of exposure. For the first time, the impact of PBDEs on the microbial homeostasis of human gut microbiota was taken into consideration, revealing noteworthy modifications with serious health implications even at oral exposure doses considered as safe by worldwide regulatory entities. The research leading to these results received financial support from FEDER (Programa Operacional Competitividade e Internacionalização - COMPETE 2020), from PIDDAC through FCT/MCTES project POCI-01-0145-FEDER-028708 and from UID/QUI/50006/2019 with funding from FCT/MCTES. Rebeca Cruz thanks to FCT for the PhD grant SFRH/BD/101945/2014. António Marques thanks to FCT IF program. Miguel Faria acknowledges FCT the researcher contract. Sara C. Cunha also acknowledges FCT for the IF/01616/2015 contract.

An epidemiological survey on Blastocystis was carried out enrolling a total of 2524 subjects referred to the Umberto I Academic Hospital in Rome, for the routine parasitological exams, during 2017–2018. The studied population included a sample of immunocompromised individuals (N = 130) followed at the same hospital. DNA sequencing of the small subunit rRNA gene (SSU rDNA) locus was performed on samples positive to the coproparasitological analysis to molecular characterize the Blastocystis-subtypes. Microscopical analysis detected Blastocystis in 192/2524 (7.6%) of the enrolled subjects. It was the organism most frequently identified in the analysed faecal samples diagnosed in single infection (5.6%) or in co-infection with other enteric protozoa (2%). Furthermore, it was found mainly in immunocompromised patients (22.3%) compared to immunocompetent ones (6.8%). As expected, ST3 was the most occurring subtype identified in 40% of the subjects, followed by ST1 (29%), ST2 (16%), ST4 (12%), and ST7 (3%). Next-generation sequencing (NGS) of the 16S rDNA was performed on a sub-sample of Blastocystis-ST3-carriers, homogenous by age and gender, as well as on Blastocystis-free subjects, to profile and compare their gut bacterial composition. A higher bacterial diversity was found in ST3-Blastocystis-carriers, which exhibited a high abundance of Prevotella, Methanobrevibacter and Ruminococcus while, a high percentage of Bacteroides was found in Blastocystis-free subjects. This study evidenced the presence of Blastocystis in 7.6% of faecal samples in Italy and a high circulation of the protist among immunocompromised patients (22.3%). Molecular characterization of positive samples evidenced the occurrence of five different subtypes, including zoonotic ST such as the ST7, highlighting the risk of transmission from animals. Study of the gut microbiota composition confirms previous evidences according to which, the colonisation by Blastocystis would be linked with an eubiotic gut characterized by potentially beneficial species such as Prevotella and Ruminococcus, rather than with a dysbiotic state, with a high abundance of Enterobacteriaceae, and corroborated the role of the protist as “an old friend” of the human gut., Highlights • Microscopical analysis detected Blastocystis in 7.6% subjects in Italy. • Molecular methods allowed the identification of 5 STs (ST1, ST2, ST3, ST4, ST7). • ST3 resulted the most frequent subtype. • NGS of the 16S rDNA gene was performed in Blastocystis-ST3-carriers. • Blastocystis-ST3 resulted correlated to an eubiotic gut characterized by potentially beneficial species.

peer-reviewed Background Faecalibacterium prausnitzii is a ubiquitous member of the human gut microbiome, constituting up to 15% of the total bacteria in the human gut. Substantial evidence connects decreased levels of F. prausnitzii with the onset and progression of certain forms of inflammatory bowel disease, which has been attributed to its anti-inflammatory potential. Two phylogroups of F. prausnitzii have been identified, with a decrease in phylogroup I being a more sensitive marker of intestinal inflammation. Much of the genomic and physiological data available to date was collected using phylogroup II strains. Little analysis of F. prausnitzii genomes has been performed so far and genetic differences between phylogroups I and II are poorly understood. Results In this study we sequenced 11 additional F. prausnitzii genomes and performed comparative genomics to investigate intraspecies diversity, functional gene complement and the mobilome of 31 high-quality draft and complete genomes. We reveal a very low level of average nucleotide identity among F. prausnitzii genomes and a high level of genome plasticity. Two genomogroups can be separated based on differences in functional gene complement, albeit that this division does not fully agree with separation based on conserved gene phylogeny, highlighting the importance of horizontal gene transfer in shaping F. prausnitzii genomes. The difference between the two genomogroups is mainly in the complement of genes associated with catabolism of carbohydrates (such as a predicted sialidase gene in genomogroup I) and amino acids, as well as defense mechanisms. Conclusions Based on the combination of ANI of genomic sequences, phylogenetic analysis of core proteomes and functional differences we propose to separate the species F. prausnitzii into two new species level taxa: F. prausnitzii sensu stricto (neotype strain A2–165T = DSM 17677T = JCM 31915T) and F. moorei sp. nov. (type strain ATCC 27768T = NCIMB 13872T). This research was conducted with the financial support of Science Foundation Ireland (SFI) under Grant Number SFI/12/RC/2273, a Science Foundation Ireland’s Spokes Programme which is co-funded under the European Regional Development Fund under Grant Number SFI/14/SP APC/B3032, and a research grant from Janssen Biotech, Inc.

Recent metagenomic analysis has revealed that our gut microbiota plays an important role in not only the maintenance of our health but also various diseases such as obesity, diabetes, inflammatory bowel disease, and allergy. However, most intestinal bacteria are considered ‘unculturable’ bacteria, and their functions remain unknown. Although culture-independent genomic approaches have enabled us to gain insight into their potential roles, culture-based approaches are still required to understand their characteristic features and phenotypes. To date, various culturing methods have been attempted to obtain these ‘unculturable’ bacteria, but most such methods require advanced techniques. Here, we have tried to isolate possible unculturable bacteria from a healthy Japanese individual by using commercially available media. A 16S rRNA (ribosomal RNA) gene metagenomic analysis revealed that each culture medium showed bacterial growth depending on its selective features and a possibility of the presence of novel bacterial species. Whole genome sequencing of these candidate strains suggested the isolation of 8 novel bacterial species classified in the Actinobacteria and Firmicutes phyla. Our approach indicates that a number of intestinal bacteria hitherto considered unculturable are potentially culturable and can be cultured on commercially available media. We have obtained novel gut bacteria from a healthy Japanese individual using a combination of comprehensive genomics and conventional culturing methods. We would expect that the discovery of such novel bacteria could illuminate pivotal roles for the gut microbiota in association with human health.

The human gut microbiota is highly dynamic, and host physiology and diet exert major influences on its composition. In our recent study, we integrated new quantitative measurements on bacterial growth physiology with a reanalysis of published data on human physiology to build a comprehensive modeling framework. This can generate predictions of how changes in different host factors influence microbiota composition. For instance, hydrodynamic forces in the colon, along with colonic water absorption that manifests as transit time, exert a major impact on microbiota density and composition. This can be mechanistically explained by their effect on colonic pH which directly affects microbiota competition for food. In this addendum, we describe the underlying analysis in more detail. In particular, we discuss the mixing dynamics of luminal content by wall contractions and its implications for bacterial growth and density, as well as the broader implications of our insights for the field of gut microbiota research.

‘Cellulomonas timonensis’ sp. nov. strain sn7T is a new species within the Cellulomonas genus. We present the main phenotypic characteristics and provide a complete annotation of its genome sequence. This facultative anaerobic bacterium, isolated from the stool of 38-year-old obese Frenchman, is Gram-positive, has motile rods and is sporulating. The genome is 4 057 828 bp long with 72.42% G + C content. Of the 3732 predicted genes, 3667 were protein-coding genes and 65 were RNAs. Keywords: Cellulomonas timonensis, culturomics, genome, human gut microbiota, taxonogenomics

Interest in gut microbiota analyses is at an all-time high. Gut microbiota is thought to relate to an increasing range of diseases of interest to physicians and nutritionists. Overweight, obesity, response to diet, metabolic syndrome, low grade inflammation, diabetes and colon neoplasms could maybe be observed in microbiota if affordable markers were available. Possible biomarkers like the Firmicutes/Bacteroidetes ratio, the Gram-positive/Gram-negative ratio, the Prevotella/Bacteroides ratio, and the Fusobacterium nucleatum/Faecalibacterium prausnitzii ratio are here reviewed in a narrative way in the attempt to highlight their possible future role in routine practice and clinically relevant diagnostics.

Among the various parameters obtainable through the analysis of the human gut microbiota, the enterotype is one of the first classifications of the bacterial consortia, which tried to obtain, at the same time, as much information as possible to be applied in clinical medicine. Although some authors observed the existence not of clusters, but only of a real continuous gradient, enterotypes are commonly described according to various models. The first model predicted either clustering into enterotypes 1 and 2 based on two specific dominances, Bacteroides and Prevotella, respectively, with the Ruminococcus dominance blurred within the Bacteroides dominance, or it predicted a threedominant condition, in which the Ruminococcus driver constituted enterotype 3, separated from enterotype 1. A second model envisaged three possible ways to cluster gut microbiota, respectively centred on two, three, and four dominances. In the first case, enterotypes 1 and 2 coincided with the two original enterotypes, with the dominance of Bacteroides and Prevotella, respectively. In the second case, the existence of enterotype 3 was evident and whose dominance was not centred on Ruminococcus but extended more towards the entire Firmicutes phylum. In the third case, the presence of the phylum Firmicutes was split into two different enterotypes generating the clusters defined and named as Mixtures 1 and 2. Subsequently, the analysis of the water content (hydration) in the stool allowed the splitting of the Bacteroides enterotype into two sub-enterotype, respectively known as B1 and B2. All these models have allowed us to highlight some correlations between a specific enterotype, or cluster, and some characteristics, such as the greater predisposition of the respective hosts towards certain pathologies. These observations, coupled with the attempt to derive the different microbiota on an evolutionary basis, can help to shed new light on this topic and demonstrate the possible utility that the different ways of clustering the gut microbiota can have in a clinical application perspective and in preventive medicine.

The major nutrients available to human colonic Bacteroides species are glycans, exemplified by pectins, a network of covalently linked plant cell wall polysaccharides containing galacturonic acid (GalA). Metabolism of complex carbohydrates by the Bacteroides genus is orchestrated by polysaccharide utilization loci (PULs). In Bacteroides thetaiotaomicron, a human colonic bacterium, the PULs activated by different pectin domains have been identified; however, the mechanism by which these loci contribute to the degradation of these GalA-containing polysaccharides is poorly understood. Here we show that each PUL orchestrates the metabolism of specific pectin molecules, recruiting enzymes from two previously unknown glycoside hydrolase families. The apparatus that depolymerizes the backbone of rhamnogalacturonan-I is particularly complex. This system contains several glycoside hydrolases that trim the remnants of other pectin domains attached to rhamnogalacturonan-I, and nine enzymes that contribute to the degradation of the backbone that makes up a rhamnose-GalA repeating unit. The catalytic properties of the pectin-degrading enzymes are optimized to protect the glycan cues that activate the specific PULs ensuring a continuous supply of inducing molecules throughout growth. The contribution of Bacteroides spp. to metabolism of the pectic network is illustrated by cross-feeding between organisms., This work was supported in part by an Advanced Grant from the European Research Council (Grant No. 322820) awarded to H.J.G. and B.H. supporting A.S.L., D.N., A.C. and N.T., a Wellcome Trust Senior Investigator Award to H.J.G. (grant No. WT097907MA) that supported J.B. and E.C.L. a European Union Seventh Framework Initial Training Network Programme entitled the “WallTraC project” (Grant Agreement number 263916) awarded to M-C.R. and H.J.G, which supported X.Z. and J.S. The Biotechnology and Biological Research Council project ‘Ricefuel’ (grant numbers BB/K020358/1) awarded to H.J.G. supported A.L.

The shotgun sequencing data presented in this report are related to the research article named “Gut microbiome shotgun sequencing in assessment of microbial community changes associated with H. pylori eradication therapy” (Khusnutdinova et al., 2016) [1] . Typically, the H. pylori eradication protocol includes a prolonged two-week use of the broad-spectrum antibiotics. The presented data on the whole-genome sequencing of the total DNA from stool samples of patients before the start of the eradication, immediately after eradication and several weeks after the end of treatment could help to profile the gut microbiota both taxonomically and functionally. The presented data together with those described in Glushchenko et al. (2017) [2] allow researchers to characterize the metagenomic profiles in which the use of antibiotics could result in dramatic changes in the intestinal microbiota composition. We perform 15 gut metagenomes from 5 patients with H. pylori infection, obtained through the shotgun sequencing on the SOLiD 5500 W platform. Raw reads are deposited in the ENA under project ID PRJEB21338.

Clostridium (C.) difficile-infection (CDI), a nosocomial gastrointestinal disorder, is of growing concern due to its rapid rise in recent years. Antibiotic therapy of CDI is associated with disrupted metabolic function and altered gut microbiota. The use of probiotics as an adjunct is being studied extensively due to their potential to modulate metabolic functions and the gut microbiota. In the present study, we assessed the ability of several single strain probiotics and a probiotic mixture to change the metabolic functions of normal and C. difficile-infected fecal samples. The production of short-chain fatty acids (SCFAs), hydrogen sulfide (H2S), and ammonia was measured, and changes in microbial composition were assessed by 16S rRNA gene amplicon sequencing. The C. difficile-infection in fecal samples resulted in a significant decrease (p &lt, 0.05) in SCFA and H2S production, with a lower microbial alpha diversity. All probiotic treatments were associated with significantly increased (p &lt, 0.05) levels of SCFAs and restored H2S levels. Probiotics showed no effect on microbial composition of either normal or C. difficile-infected fecal samples. These findings indicate that probiotics may be useful to improve the metabolic dysregulation associated with C. difficile infection.

Genome sequencing has revealed substantial variation in the predicted abilities of individual species within animal gut microbiota to metabolize the complex carbohydrates comprising dietary fiber. At the same time, a currently limited body of functional studies precludes a richer understanding of how dietary glycan structures affect the gut microbiota composition and community dynamics. Here, using biochemical and biophysical techniques, we identified and characterized differences among recombinant proteins from syntenic xyloglucan utilization loci (XyGUL) of three Bacteroides and one Dysgonomonas species from the human gut, which drive substrate specificity and access to distinct polysaccharide side chains. Enzymology of four syntenic glycoside hydrolase family 5 subfamily 4 (GH5_4) endo-xyloglucanases revealed surprising differences in xyloglucan (XyG) backbone cleavage specificity, including the ability of some homologs to hydrolyze congested branched positions. Further, differences in the complement of GH43 alpha-l-arabinofuranosidases and GH95 alpha-l-fucosidases among syntenic XyGUL confer distinct abilities to fully saccharify plant species-specific arabinogalactoxyloglucan and/or fucogalactoxyloglucan. Finally, characterization of highly sequence-divergent cell surface glycan-binding proteins (SGBPs) across syntenic XyGUL revealed a novel group of XyG oligosaccharide-specific SGBPs encoded within select Bacteroides. IMPORTANCE The catabolism of complex carbohydrates that otherwise escape the endogenous digestive enzymes of humans and other animals drives the composition and function of the gut microbiota. Thus, detailed molecular characterization of dietary glycan utilization systems is essential both to understand the ecology of these complex communities and to manipulate their compositions, e.g., to benefit human health. Our research reveals new insight into how ubiquitous members of the human gut microbiota have evolved a set of microheterogeneous gene clusters to efficiently respond to the structural variations of plant xyloglucans. The data here will enable refined functional prediction of xyloglucan utilization among diverse environmental taxa in animal guts and beyond.

Background Human gut microbiota are important for human health and have been regarded as a “forgotten organ”, whose variation is closely linked with various factors, such as host genetics, diet, pathological conditions and external environment. The diversity of human gut microbiota has been correlated with aging, which was characterized by different abundance of bacteria in various age groups. In the literature, most of the previous studies of age-related gut microbiota changes focused on individual species in the gut community with supervised methods. Here, we aimed to examine the underlying aging progression of the human gut microbial community from an unsupervised perspective. Results We obtained raw 16S rRNA sequencing data of subjects ranging from newborns to centenarians from a previous study, and summarized the data into a relative abundance matrix of genera in all the samples. Without using the age information of samples, we applied an unsupervised algorithm to recapitulate the underlying aging progression of microbial community from hosts in different age groups and identify genera associated to this progression. Literature review of these identified genera indicated that for individuals with advanced ages, some beneficial genera are lost while some genera related with inflammation and cancer increase. Conclusions The multivariate unsupervised analysis here revealed the existence of a continuous aging progression of human gut microbiota along with the host aging process. The identified genera associated to this aging process are meaningful for designing probiotics to maintain the gut microbiota to resemble a young age, which hopefully will lead to positive impact on human health, especially for individuals in advanced age groups.

Gut microbiota play an important role in metabolism of intake saponins, and parallelly, the polysaccharides deriving from herbal products possess effects on gut microbiota. Ophiopogonis Radix is a common Chinese herb that is popularly used as functional food in China. Polysaccharide and steroidal saponin, e.g., ophiopogonin, mainly ophiopogonin D (Oph-D) and ophiopogonin D&rsquo, (Oph-D&rsquo, ), are the major constituents in this herb. In order to reveal the role of gut microbiota in metabolizing ophiopogonin, an in vitro metabolism of Oph-D and Oph-D&rsquo, by human gut microbiota, in combination with or without Ophiopogon polysaccharide, was conducted. A sensitive and reliable UPLC-MS/MS method was developed to simultaneously quantify Oph-D, Oph-D&rsquo, and their final metabolites, i.e., ruscogenin and diosgenin in the broth of microbiota. An elimination of Oph-D and Oph-D&rsquo, was revealed in a time-dependent manner, as well as the recognition of a parallel increase of ruscogenin and diosgenin. Ophiopogon polysaccharide was shown to stimulate the gut microbiota-induced metabolism of ophiopogonins. This promoting effect was further verified by increased activities of &beta, D-glucosidase, &beta, D-xylosidase, &alpha, L-rhamnosidase and &beta, D-fucosidase in the broth. This study can be extended to investigate the metabolism of steroidal saponins by gut microbiota when combined with other herbal products, especially those herbs enriched with polysaccharides.

Made available in DSpace on 2019-10-06T16:05:40Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-06-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Strategiske Forskningsråd Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) The gut microbiota composition and its metabolites have high impact on human health. Exploitation of prebiotics and probiotics for modulation of gut microbiota can lead to promising outcomes. This study aimed to evaluate the effects of the probiotic strain Bifidobacterium longum BB-46 alone and in combination with a citric pectin from lemon on the gut microbiota from healthy adults using the Simulator of Human Intestinal Microbial Ecosystem (SHIME ® ). Changes in microbiota composition and in metabolic activity were assessed by the 16S rRNA gene sequencing and by analyses of short-chain fatty acids (SCFAs) and ammonium ions (NH 4 + ). An increase in the relative abundances of Firmicutes (especially the members of Lachnospiraceae and Lactobacillaceae families) and Bacteroidetes was observed during treatment with B. longum BB-46 alone in all compartments of the colon. Treatment with B. longum BB-46 and pectin stimulated an increase in the proportions of genera Faecalibacterium, Eubacterium and Lactobacillus, as well as in the Ruminococcaceae family in the transverse and descending colons. Concurrently, the butyrate levels increased in these two compartments. Additionally, the combination of B. longum BB-46 and pectin reduced the abundance of proteolytic bacteria Bacteroides, Clostridium, Peptoniphilus, and Streptococcus, along with decreased NH 4 + production. No significant changes could be observed on NH 4 + production by treatment with B. longum BB-46, nor did it increase the amount of SCFAs. In this study, we observed that although each treatment was able to modulate the microbiota, the combination of B. longum BB-46 and pectin was more efficient in decreasing the intestinal NH 4 + levels and in increasing butyric acid-producing bacteria. These findings indicate that B. longum BB-46, especially when combined with the specific citric pectin, might have beneficial impact on human health. Department of Food and Nutrition School of Pharmaceutical Sciences State University of São Paulo (UNESP) Department of Food Science Faculty of Science University of Copenhagen Department of Hydraulics and Sanitation School of Engineering of São Carlos University of São Paulo (USP) Department of Biochemical and Pharmaceutical Technology University of São Paulo (USP) Food Research Center University of São Paulo (USP) Department of Food and Nutrition School of Pharmaceutical Sciences State University of São Paulo (UNESP)

Cross talk occurs between the human gut and the lung through a gut-lung axis involving the gut microbiota. However, the signatures of the human gut microbiota after active Mycobacterium tuberculosis infection have not been fully understood. Here, we investigated changes in the gut microbiota in tuberculosis (TB) patients by shotgun sequencing the gut microbiomes of 31 healthy controls and 46 patients. We observed a dramatic changes in gut microbiota in tuberculosis patients as reflected by significant decreases in species number and microbial diversity. The gut microbiota of TB patients were mostly featured by the striking decrease of short-chain fatty acids (SCFAs)-producingbacteria as well as associated metabolic pathways. A classification model based on the abundance of three species, Haemophilus parainfluenzae, Roseburia inulinivorans, and Roseburia hominis, performed well for discriminating between healthy and diseased patients. Additionally, the healthy and diseased states can be distinguished by SNPs in the species of B. vulgatus. We present a comprehensive profile of changes in the microbiota in clinical TB patients. Our findings will shed light on the design of future diagnoses and treatments for M. tuberculosis infections.

Dietary phenolics or polyphenols are mostly metabolized by the human gut microbiota. These metabolites appear to confer the beneficial health effects attributed to phenolics. Microbial composition affects the type of metabolites produced. Reciprocally, phenolics modulate microbial composition. Understanding this relationship could be used to positively impact health by phenolic supplementation and thus create favorable colonic conditions. This study explored the effect of six stilbenoids (batatasin III, oxyresveratrol, piceatannol, pinostilbene, resveratrol, thunalbene) on the gut microbiota composition. Stilbenoids were anaerobically fermented with fecal bacteria from four donors, samples were collected at 0 and 24 h, and effects on the microbiota were assessed by 16S rRNA gene sequencing. Statistical tests identified affected microbes at three taxonomic levels. Observed microbial composition modulation by stilbenoids included a decrease in the Firmicutes to Bacteroidetes ratio, a decrease in the relative abundance of strains from the genus Clostridium, and effects on the family Lachnospiraceae. A frequently observed effect was a further decrease of the relative abundance when compared to the control. An opposite effect to the control was observed for Faecalibacterium prausnitzii, whose relative abundance increased. Observed effects were more frequently attributed to resveratrol and piceatannol, followed by thunalbene and batatasin III.

Background Intestinal microbiota have been suggested to play an important role in the pathogenesis of obesity and type 2 diabetes. Bariatric surgery improves both conditions and has been associated with changes in intestinal microbiota composition. We investigated the effect of a nonsurgical bariatric technique on intestinal microbiota composition in relation to metabolic improvement.Methods Seventeen patients with obesity and type 2 diabetes were treated with the nonsurgical duodenal-jejunal bypass liner, which excludes the proximal 60 cm small intestine from food. Fecal samples as well as metabolic parameters reflecting obesity and type 2 diabetes were obtained from the patients at baseline, after 6 months with the device in situ, and 6 months after explantation.Results After 6 months of treatment, both obesity and type 2 diabetes had improved with a decrease in weight from 106.1 [99.4-123.5] to 97.4 [89.4-114.0] kg and a decrease in HbA(1c) from 8.5% [7.6-9.2] to 7.2% [6.3-8.1] (both p Conclusion Improvement of obesity and type 2 diabetes after exclusion of the proximal 60 cm small intestine by treatment with a nonsurgical duodenal-jejunal bypass liner may be promoted by changes in fecal microbiota composition.

Made available in DSpace on 2018-12-11T17:22:38Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-01-01 In this study, we first investigated the survival of three probiotic strains, individually and combined with acerola by-product during simulated gastrointestinal conditions. Next, we investigated the effects of acerola by-product combined with Bifidobacterium longum BB-46 on a gut microbiota model (SHIME®). Chemical composition, total phenolic compounds, antioxidant activity of the acerola by-product and microbial counts, denaturing gradient gel electrophoresis (DGGE), ammonium ions (NH+ 4) and short-chain fatty acids (SCFAs) analysis of the SHIME® samples were performed. Acerola by-product revealed high protein and fibre, reduced lipid contents, and showed to be an excellent source of total phenolic compounds with high in vitro antioxidant activity. A decreased amount of NH+ 4 in the ascending colon and an increase (p

Bifidobacteria are exposed to substantial amounts of dietary β-galactosides. Distinctive preferences for growth on different β-galactosides are observed within Bifidobacterium members, but the basis of these preferences remains unclear. We previously described the first β-(1,6)/(1,3)-galactosidase from Bifidobacterium animalis subsp. lactis Bl-04. This enzyme is relatively promiscuous, exhibiting only 5-fold higher efficiency on the preferred β-(1,6)-galactobiose than the β-(1,4) isomer. Here, we characterize the solute-binding protein (Bal6GBP) that governs the specificity of the ABC transporter encoded by the same β-galactoside-utilization locus. We observed that although Bal6GBP recognizes both β-(1,6)- and β-(1,4)-galactobiose, Bal6GBP has a 1630-fold higher selectivity for the former, reflected in dramatic differences in growth, with several hours lag on less preferred β-(1,4)- and β-(1,3)-galactobiose. Experiments performed in the presence of varying proportions of β-(1,4)/ β-(1,6)-galactobioses indicated that the preferred substrate was preferentially depleted from the culture supernatant. This established that the poor growth on the non-preferred β-(1,4) was due to inefficient uptake. We solved the structure of Bal6GBP in complex with β-(1,6)-galactobiose at 1.39 Å resolution, revealing the structural basis of this strict selectivity. Moreover, we observed a close evolutionary relationship with the human milk disaccharide lacto-N-biose-binding protein from Bifidobacterium longum, indicating that the recognition of the non-reducing galactosyl is essentially conserved, whereas the adjacent position is diversified to fit different glycosidic linkages and monosaccharide residues. These findings indicate that oligosaccharide uptake has a pivotal role in governing selectivity for distinct growth substrates and have uncovered evolutionary trajectories that shape the diversification of sugar-uptake proteins within Bifidobacterium.

Undigestible, insoluble food particles, such as wheat bran, are important dietary constituents that serve as a fermentation substrate for the human gut microbiota. The first step in wheat bran fermentation involves the poorly studied solubilization of fibers from the complex insoluble wheat bran structure. Attachment of bacteria has been suggested to promote the efficient hydrolysis of insoluble substrates, but the mechanisms and drivers of this microbial attachment and colonization, as well as subsequent fermentation remain to be elucidated. We have previously shown that an individually dependent subset of gut bacteria is able to colonize the wheat bran residue. Here, we isolated these bran-attached microorganisms, which can then be used to gain mechanistic insights in future pure culture experiments. Four healthy fecal donors were screened to account for inter-individual differences in gut microbiota composition. A combination of a direct plating and enrichment method resulted in the isolation of a phylogenetically diverse set of species, belonging to theBacteroidetes,Firmicutes,ProteobacteriaandActinobacteriaphyla. A comparison with 16S rRNA gene sequences that were found enriched on wheat bran particles in previous studies, however, showed that the isolates do not yet cover the entire diversity of wheat-bran colonizing species, comprising among others a broad range ofPrevotella,BacteroidesandClostridiumcluster XIVa species. We, therefore, suggest several modifications to the experiment set-up to further expand the array of isolated species.

Type 2 diabetes mellitus (T2DM) is an important public health problem. The knowledge of bacterial communities in the gut of Vietnamese patients with T2DM and non diabetic controls is still insufficient. We report in this article the 16S rDNA amplicon data of the gut microbiomes of Vietnamese patients with T2DM and nondiabetic controls carried out using the Illumina sequencing. This work included 7 patients and 7 controls. A total of 1,627,646 reads were obtained and a total of 13 phyla, 25 classes, 94 genera were revealed. The top three dominant bacterial phyla in all subjects were Firmicutes, Bacteroidetes and Proteobacteria. Significant differences in the relative abundances of the phylum Firmicutes and class Clostridia between patients and controls were observed, suggesting that the reducing of phylum Firmicutes and class Clostridia in the gut may be linked to obesity and T2DM. All sequencing libraries were deposited in the NCBI SRA as BioProject PRJNA668251. The datasets are needed to determine the association between the bacterial composition of the gut and the pathogenesis of T2DM in Vietnamese patients.

We investigated associations of habitual dietary intake with the taxonomic composition and diversity of the human gut microbiota in 222 Koreans aged 18&ndash, 58 years in a cross-sectional study. Gut microbiota data were obtained by 16S rRNA gene sequencing on DNA extracted from fecal samples. The habitual diet for the previous year was assessed by a food frequency questionnaire. After multivariable adjustment, intake of several food groups including vegetables, fermented legumes, legumes, dairy products, processed meat, and non-alcoholic beverages were associated with major phyla of the gut microbiota. A dietary pattern related to higher &alpha, diversity (Hi&alpha, DP) derived by reduced rank regression was characterized by higher intakes of fermented legumes, vegetables, seaweeds, and nuts/seeds and lower intakes of non-alcoholic beverages. The Hi&alpha, DP was positively associated with several genera of Firmicutes such as Lactobacillus, Ruminococcus, and Eubacterium (all p &lt, 0.05). Among enterotypes identified by principal coordinate analysis based on the &beta, diversity, the Ruminococcus enterotype had higher Hi&alpha, DP scores and was strongly positively associated with intakes of vegetables, seaweeds, and nuts/seeds, compared to the two other enterotypes. We conclude that a plant- and fermented food-based diet was positively associated with some genera of Firmicutes (e.g., Lactobacillus, Ruminococcus, and Eubacterium) reflecting better gut microbial health.

Complex glycans that evade our digestive system are major nutrients that feed the human gut microbiota (HGM). The prevalence of Bacteroidetes in the HGM of populations worldwide is engendered by the evolution of polysaccharide utilization loci (PULs), which encode concerted protein systems to utilize the myriad complex glycans in our diets. Despite their crucial roles in glycan recognition and transport, cell-surface glycan-binding proteins (SGBPs) remained understudied cogs in the PUL machinery. Here, we report the structural and biochemical characterization of a suite of SGBP-A and SGBP-B structures from three syntenic β(1,3)-glucan utilization loci (1,3GULs) from Bacteroides thetaiotaomicron (Bt), Bacteroides uniformis (Bu), and B. fluxus (Bf), which have varying specificities for distinct β-glucans. Ligand complexes provide definitive insight into β(1,3)-glucan selectivity in the HGM, including structural features enabling dual β(1,3)-glucan/mixed-linkage β(1,3)/β(1,4)-glucan-binding capability in some orthologs. The tertiary structural conservation of SusD-like SGBPs-A is juxtaposed with the diverse architectures and binding modes of the SGBPs-B. Specifically, the structures of the trimodular BtSGBP-B and BuSGBP-B revealed a tandem repeat of carbohydrate-binding module-like domains connected by long linkers. In contrast, BfSGBP-B comprises a bimodular architecture with a distinct β-barrel domain at the C terminus that bears a shallow binding canyon. The molecular insights obtained here contribute to our fundamental understanding of HGM function, which in turn may inform tailored microbial intervention therapies.