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Background: Microbial colonization of the gastrointestinal tract after birth is an essential event that influences infant health with life-long consequences. Therefore, it is important to investigate strategies to positively modulate colonization in early life., Objectives: This randomized, controlled intervention study included 540 infants to investigate the effects of a synbiotic intervention formula (IF) containing Limosilactobacillus fermentum CECT5716 and galacto-oligosaccharides on the fecal microbiome., Methods: The fecal microbiota from infants was analyzed by 16S rRNA amplicon sequencing at 4, 12, and 24 months of age. Metabolites (e.g., short-chain fatty acids) and other milieu parameters (e.g., pH, humidity, and IgA) were also measured in stool samples., Results: Microbiota profiles changed with age, with major differences in diversity and composition. Significant effects of the synbiotic IF compared with control formula (CF) were visible at month 4, including higher occurrence of Bifidobacterium spp. and Lactobacillaceae and lower occurrence of Blautia spp., as well as Ruminoccocus gnavus and relatives. This was accompanied by lower fecal pH and concentrations of butyrate. After de novo clustering at 4 months of age, overall phylogenetic profiles of the infants receiving IF were closer to reference profiles of those fed with human milk than infants fed CF. The changes owing to IF were associated with fecal microbiota states characterized by lower occurrence of Bacteroides compared with higher levels of Firmicutes (valid name Bacillota), Proteobacteria (valid name Pseudomonadota), and Bifidobacterium at 4 months of age. These microbiota states were linked to higher prevalence of infants born by Cesarean section., Conclusions: The synbiotic intervention influenced fecal microbiota and milieu parameters at an early age depending on the overall microbiota profiles of the infants, sharing a few similarities with breastfed infants. This trial was registered at clinicaltrials.gov as NCT02221687., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Microbiome research needs comprehensive repositories of cultured bacteria from the intestine of mammalian hosts. We expanded the mouse intestinal bacterial collection (www.dsmz.de/miBC) to 212 strains, all publicly available and taxonomically described. This includes strain-level diversity, small-sized bacteria, and previously undescribed taxa (one family, 10 genera, and 39 species). This collection enabled metagenome-educated prediction of synthetic communities (SYNs) that capture key functional differences between microbiomes, notably identifying communities associated with either resistance or susceptibility to DSS-induced colitis. Additionally, nine species were used to amend the Oligo-Mouse Microbiota (OMM)12 model, yielding the OMM19.1 model. The added strains compensated for phenotype differences between OMM12 and specific pathogen-free mice, including body composition and immune cells in the intestine and associated lymphoid tissues. Ready-to-use OMM stocks are available for future studies. In conclusion, this work improves our knowledge of gut microbiota diversity in mice and enables functional studies via the modular use of isolates., Competing Interests: Declaration of interests T.C. has ongoing scientific collaborations with Cytena GmbH, HiPP GmbH, and Savanna Ingredients GmbH, which are however unrelated to the present work., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

The genus Prevotella comprises 55 species with validly published, and correct, names (at June 2021) that are phenotypically, ecologically and functionally diverse. This study used a range of comparative genome approaches (marker gene-based genome phylogeny, core genome phylogeny, average amino acid identity, percentage of conserved proteins and clade-specific marker genes) to identify large differences between the 53 species for which genomes are available, as well as two effectively published yet not validly named species and four novel species. These differences were consistent between the various analysis methods and justify the separation of Prevotella into multiple genera. While the distribution across 19 ecosystem types was unique for each species and inconsistent within clades, the functional repertoire based on the presence/absence of both PFAMs and CAZy families revealed distinct clustering based on the proposed genera. Based on the integration of all results, we propose the reclassification of species previously assigned to the genus Prevotella into seven genera, including four novel genera for which the names Segatella, Hoylesella, Leyella and Palleniella are proposed. In addition to the reclassification of Prevotella, this work describes four novel species, Hallella faecis, Xylanibacter rodentium, Xylanibacter muris, and Palleniella intestinalis., (Copyright © 2022 Elsevier GmbH. All rights reserved.)

Bile acids are crucial for the uptake of dietary lipids and can shape the gut-microbiome composition. This latter function is associated with the toxicity of bile acids and can be modulated by bile acid modifying bacteria such as Eggerthella lenta , but the molecular details of the interaction of bacteria depending on bile acid modifications are not well understood. In order to unravel the molecular response to bile acids and their metabolites, we cultivated eight strains from a human intestinal microbiome model alone and in co-culture with Eggerthella lenta in the presence of cholic acid (CA) and deoxycholic acid (DCA). We observed growth inhibition of particularly gram-positive strains such as Clostridium ramosum and the gram-variable Anaerostipes cacae by CA and DCA stress. C. ramosum was alleviated through co-culturing with Eggerthella lenta . We approached effects on the membrane by zeta potential and genotoxic and metabolic effects by (meta)proteomic and metabolomic analyses. Co-culturing with Eggerthella lenta decreased both CA and DCA by the formation of oxidized and epimerized bile acids. Eggerthella lenta also produces microbial bile salt conjugates in a co-cultured species-specific manner. This study highlights how the interaction with other bacteria can influence the functionality of bacteria.

Cultivation via classical agar plate (CAP) approaches is widely used to study microbial communities, but they are time-consuming. An alternative approach is the application of single-cell dispensing (SCD), which allows high-throughput, label-free sorting of microscopic particles. We aimed to develop a new anaerobic SCD workflow to cultivate human gut bacteria and compared it with CAP using faecal communities on three rich culture media. We found that the SCD approach significantly decreased the experimental time to obtain pure cultures from 17 ± 4 to 5 ± 0 days, while the isolate diversity and relative abundance coverage were comparable for both approaches. We further tested the total captured fraction by sequencing the sorted bacteria directly after growth as bulk biomass from 2400 dispensed single cells without downstream identification of individual strains. In this approach, the cultured fraction increased from 35.2% to 52.2% for SCD, highlighting the potential for deeper cultivation projects from single samples. SCD-based cultivation also captured species not detected by sequencing (16 ± 5 per sample, including seven novel taxa). From this work, 82 human gut bacterial species across five phyla (Actinobacteriota, Bacteroidota, Desulfobacterota, Firmicutes and Proteobacteria) and 24 families were obtained, including the first cultured member of 11 novel genera and 10 novel species that were fully characterized taxonomically., (© 2022 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

D-Allulose, also referred to as psicose, is a C3-epimer of D-fructose used as a sugar substitute in low energy products. It can be formed naturally during processing of food and drinks containing sucrose and fructose or is prepared by chemical synthesis or via enzymatic treatment with epimerases from fructose. Estimated intakes via Western style diets including sweetened beverages are below 500 mg per d but, when used as a sugar replacement, intake may reach 10 to 30 g per d depending on the food consumed. Due to its structural similarity with fructose, allulose uses the same transport and distribution pathways. But in contrast to fructose, the human genome does not encode for enzymes that are able to metabolise allulose leading to an almost complete renal excretion of the absorbed dose and near-to-zero energetic yield. However, in vitro studies have shown that certain bacteria such as Klebsiella pneumonia are able to utilise allulose as a substrate. This finding has been a subject of concern, since Klebsiella pneumoniae represents an opportunistic human pathogen. It therefore raised the question of whether a high dietary intake of allulose may cause an undesirable growth advantage for potentially harmful bacteria at mucosal sites such as the intestine or at systemic sites following invasive infection. In this brief review, we discuss the current state of science on these issues and define the research needs to better understand the fate of allulose and its metabolic and microbiological effects when ingested as a sugar substitute.

Bile acids are a key mediator of the molecular microbiome-host interaction, and various mass spectrometry-based assays have been developed in the recent decade to quantify a wide range of bile acids. We compare existing methodologies to harmonize them. Methodology for absolute quantification of bile acids from six laboratories in Europe were compared for the quantification of the primary bile acids cholic acid (CA) and chenodeoxycholic acid (CDCA) and conjugated products glycocholic acid (GCA) and taurocholic acid (TCA). For the bacterially modified secondary bile acids, the quantification of deoxycholic acid (DCA) and lithocholic acid (LCA) was compared. For the murine bile acids, we used the primary muricholic acids (α-MCA and, β-MCA) and the intestinally produced secondary bile acid muricholic (ω-MCA). The standards were spiked into methanol:water (1:1) mix as well as in human and murine serum at either low concentration range (150-3000 nM) or high concentration range (1500-40,000 nM). The precision was better for higher concentrations. Measurements for the hydrophobic unconjugated bile acids LCA and ω-MCA were the most challenging. The quality assessments were generally very similar, and the comprehensive analyses demonstrated that data from chosen locations can be used for comparisons between studies.

The gut microbiome lies at the intersection between the environment and the host, with the ability to modify host responses to disease-relevant exposures and stimuli. This is evident in how enteric microbes interact with the immune system, e.g., supporting immune maturation in early life, affecting drug efficacy via modulation of immune responses, or influencing development of immune cell populations and their mediators. Many factors modulate gut ecosystem dynamics during daily life and we are just beginning to realise the therapeutic and prophylactic potential of microbiome-based interventions. These approaches vary in application, goal, and mechanisms of action. Some modify the entire community, such as nutritional approaches or faecal microbiota transplantation, while others, such as phage therapy, probiotics, and prebiotics, target specific taxa or strains. In this review, we assessed the experimental evidence for microbiome-based interventions, with a particular focus on their clinical relevance, ecological effects, and modulation of the immune system., (© 2022. The Author(s).)

Background: There is a growing interest in using gut commensal bacteria as "next generation" probiotics. However, this approach is still hampered by the fact that there are few or no strains available for specific species that are difficult to cultivate. Our objective was to adapt flow cytometry and cell sorting to be able to detect, separate, isolate, and cultivate new strains of commensal species from fecal material. We focused on the extremely oxygen sensitive (EOS) species Faecalibacterium prausnitzii and the under-represented, health-associated keystone species Christensenella minuta as proof-of-concept., Results: A BD Influx® cell sorter was equipped with a glovebox that covered the sorting area. This box was flushed with nitrogen to deplete oxygen in the enclosure. Anaerobic conditions were maintained during the whole process, resulting in only minor viability loss during sorting and culture of unstained F. prausnitzii strains ATCC 27766, ATCC 27768, and DSM 17677. We then generated polyclonal antibodies against target species by immunizing rabbits with heat-inactivated bacteria. Two polyclonal antibodies were directed against F. prausnitzii type strains that belong to different phylogroups, whereas one was directed against C. minuta strain DSM 22607. The specificity of the antibodies was demonstrated by sorting and sequencing the stained bacterial fractions from fecal material. In addition, staining solutions including LIVE/DEAD™ BacLight™ Bacterial Viability staining and polyclonal antibodies did not severely impact bacterial viability while allowing discrimination between groups of strains. Finally, we combined these staining strategies as well as additional criteria based on bacterial shape for C. minuta and were able to detect, isolate, and cultivate new F. prausnitzii and C. minuta strains from healthy volunteer's fecal samples., Conclusions: Targeted cell-sorting under anaerobic conditions is a promising tool for the study of fecal microbiota. It gives the opportunity to quickly analyze microbial populations, and can be used to sort EOS and/or under-represented strains of interest using specific antibodies, thus opening new avenues for culture experiments. Video abstract., (© 2022. The Author(s).)

A considerable percentage of the population is affected by alcoholic liver disease (ALD). It is characterized by inflammatory signals from the liver and other organs, such as the intestine. The NLR family pyrin domain containing 6 (NLRP6) inflammasome complex is one of the most important inflammatory mediators. The aim of this study was to evaluate a novel mouse model for ALD characterized by 8-week chronic-plus-binge ethanol administration and to investigate the role of NLRP6 inflammasome for intestinal homeostasis and ALD progression using Nlrp6 -/- mice. We showed that chronic-plus-binge ethanol administration triggers hepatic steatosis, injury, and neutrophil infiltration. Furthermore, we discovered significant changes of intestinal microbial communities, including increased relative abundances of bacteria within the phyla Bacteroidota and Campilobacterota , as well as reduced Firmicutes . In this ALD model, inhibiting NLRP6 signaling had no effect on liver steatosis or damage, but had a minor impact on intestinal homeostasis via affecting intestinal epithelium function and gut microbiota. Surprisingly, Nlrp6 loss resulted in significantly decreased hepatic immune cell infiltration. As a result, our novel mouse model encompasses several aspects of human ALD, such as intestinal dysbiosis. Interfering with NLRP6 inflammasome activity reduced hepatic immune cell recruitment, indicating a disease-aggravating role of NLRP6 during ALD.

Gut microbiome research has bloomed over the past 15 years. We have learnt a lot about the complex microbial communities that colonize our intestine. Promising avenues of research and microbiome-based applications are being implemented, with the goal of sustaining host health and applying personalized disease management strategies. Despite this exciting outlook, many fundamental questions about enteric microbial ecosystems remain to be answered. Organizational measures will also need to be taken to optimize the outcome of discoveries happening at an extremely rapid pace. This article highlights our own view of the field and perspectives for the next 15 years., (© 2021 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.)

(1) Background: Non-alcoholic fatty liver disease (NAFLD) is a growing global health problem. NAFLD progression involves a complex interplay of imbalanced inflammatory cell populations and inflammatory signals such as reactive oxygen species and cytokines. These signals can derive from the liver itself but also from adipose tissue or be mediated via changes in the gut microbiome. We analyzed the effects of a simultaneous migration blockade caused by L-selectin-deficiency and an enhancement of the anti-oxidative stress response triggered by hepatocytic Kelch-like ECH-associated protein 1 (Keap1) deletion on NAFLD progression. (2) Methods: L-selectin-deficient mice (Lsel -/- Keap1 flx/flx ) and littermates with selective hepatic Keap1 deletion (Lsel -/- Keap1 Δhepa ) were compared in a 24-week Western-style diet (WD) model. (3) Results: Lsel -/- Keap1 Δhepa mice exhibited increased expression of erythroid 2-related factor 2 (Nrf2) target genes in the liver, decreased body weight, reduced epidydimal white adipose tissue with decreased immune cell frequencies, and improved glucose response when compared to their Lsel -/- Keap1 flx/flx littermates. Although WD feeding caused drastic changes in fecal microbiota profiles with decreased microbial diversity, no genotype-dependent shifts were observed. (4) Conclusions: Upregulation of the anti-oxidative stress response improves metabolic changes in L-selectin-deficient mice but does not prevent NAFLD progression and shifts in the gut microbiota.

Environmental and host-associated microbial communities are complex ecosystems, of which many members are still unknown. Hence, it is challenging to study community dynamics and important to create model systems of reduced complexity that mimic major community functions. Therefore, we developed MiMiC, a computational approach for data-driven design of simplified communities from shotgun metagenomes. We first built a comprehensive database of species-level bacterial and archaeal genomes (n = 22 627) consisting of binary (presence/absence) vectors of protein families (Pfam = 17 929). MiMiC predicts the composition of minimal consortia using an iterative scoring system based on maximal match-to-mismatch ratios between this database and the Pfam binary vector of any input metagenome. Pfam vectorization retained enough resolution to distinguish metagenomic profiles between six environmental and host-derived microbial communities (n = 937). The calculated number of species per minimal community ranged between 5 and 11, with MiMiC selected communities better recapitulating the functional repertoire of the original samples than randomly selected species. The inferred minimal communities retained habitat-specific features and were substantially different from communities consisting of most abundant members. The use of a mixture of known microbes revealed the ability to select 23 of 25 target species from the entire genome database. MiMiC is open source and available at https://github.com/ClavelLab/MiMiC., (© 2021 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.)

16S rRNA gene amplicon sequencing is a popular approach for studying microbiomes. However, some basic concepts have still not been investigated comprehensively. We studied the occurrence of spurious sequences using defined microbial communities based on data either from the literature or generated in three sequencing facilities and analyzed via both operational taxonomic units (OTUs) and amplicon sequence variants (ASVs) approaches. OTU clustering and singleton removal, a commonly used approach, delivered approximately 50% (mock communities) to 80% (gnotobiotic mice) spurious taxa. The fraction of spurious taxa was generally lower based on ASV analysis, but varied depending on the gene region targeted and the barcoding system used. A relative abundance of 0.25% was found as an effective threshold below which the analysis of spurious taxa can be prevented to a large extent in both OTU- and ASV-based analysis approaches. Using this cutoff improved the reproducibility of analysis, i.e., variation in richness estimates was reduced by 38% compared with singleton filtering using six human fecal samples across seven sequencing runs. Beta-diversity analysis of human fecal communities was markedly affected by both the filtering strategy and the type of phylogenetic distances used for comparison, highlighting the importance of carefully analyzing data before drawing conclusions on microbiome changes. In summary, handling of artifact sequences during bioinformatic processing of 16S rRNA gene amplicon data requires careful attention to avoid the generation of misleading findings. We propose the concept of effective richness to facilitate the comparison of alpha-diversity across studies., (© 2021. The Author(s).)

The study of microbial communities is hampered by the large fraction of still unknown bacteria. However, many of these species have been isolated, yet lack a validly published name or description. The validation of names for novel bacteria requires that the uniqueness of those taxa is demonstrated and their properties are described. The accepted format for this is the protologue, which can be time-consuming to create. Hence, many research fields in microbiology and biotechnology will greatly benefit from new approaches that reduce the workload and harmonise the generation of protologues.We have developed Protologger, a bioinformatic tool that automatically generates all the necessary readouts for writing a detailed protologue. By producing multiple taxonomic outputs, functional features and ecological analysis using the 16S rRNA gene and genome sequences from a single species, the time needed to gather the information for describing novel taxa is substantially reduced. The usefulness of Protologger was demonstrated by using three published isolate collections to describe 34 novel taxa, encompassing 17 novel species and 17 novel genera, including the automatic generation of ecologically and functionally relevant names. We also highlight the need to utilise multiple taxonomic delineation methods, as while inconsistencies between each method occur, a combined approach provides robust placement. Protologger is open source; all scripts and datasets are available, along with a webserver at www.protologger.de., (© 2021. The Author(s).)

The gut microbiome is crucial for both maturation of the immune system and colonization resistance against enteric pathogens. Although chicken are important domesticated animals, the impact of their gut microbiome on the immune system is understudied. Therefore, we investigated the effect of microbiome-based interventions on host mucosal immune responses. Increased levels of IgA and IgY were observed in chickens exposed to maternal feces after hatching compared with strict hygienic conditions. This was accompanied by increased gut bacterial diversity as assessed by 16S rRNA gene amplicon sequencing. Cultivation work allowed the establishment of a collection of 43 bacterial species spanning 4 phyla and 19 families, including the first cultured members of 3 novel genera and 4 novel species that were taxonomically described. This resource is available at www.dsmz.de/chibac A synthetic community consisting of nine phylogenetically diverse and dominant species from this collection was designed and found to be moderately efficient in boosting immunoglobulin levels when provided to chickens early in life. IMPORTANCE The immune system plays a crucial role in sustaining animal health. Its development is markedly influenced by early microbial colonization of the gastrointestinal tract. As chicken are fully dependent on environmental microbes after hatching, extensive hygienic measures in production facilities are detrimental to the microbiota, resulting in low colonization resistance against pathogens. To combat enteric infections, antibiotics are frequently used, which aggravates the issue by altering gut microbiota colonization. Intervention strategies based on cultured gut bacteria are proposed to influence immune responses in chicken., (Copyright © 2021 Zenner et al.)

Gut microbes affect the physiology of their hosts. Studying their diversity and functions is thus of utmost importance as it will open new avenues towards the discovery of new biomolecules and the treatment of diseases. Gut microbiome research is currently boosted by the unification of metagenomics, which has dominated the field in the last two decades, and cultivation, which is experiencing a renaissance. Each of these approaches has advantages and drawbacks that can be overcome if used synergistically. In this brief article, we summarize recent literature and own studies on the cultivation of gut microbes, provide a succinct status quo of cultured fractions and collections of isolates, and give short opinions on challenges and next steps to take., (Copyright © 2021 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Background: Little is known about the complex interactions between the diet, the gut microbiota, and enteropathogens. Here, the impact of two specific diets on the composition of the mouse gut microbiota and on the transcriptional response of Salmonella Typhimurium (S. Typhimurium) was analyzed in an enteritis model., Results: Mice were fed for two weeks a fibre-rich, plant-based diet (PD), or a Westernized diet (WD) rich in animal fat and proteins and in simple sugars, and then infected with an invasin-negative S. Typhimurium strain ST4/74 following streptomycin-treatment. Seventy-two hours post infection, fecal pathogen loads were equal in both diet groups, suggesting that neither of the diets had negatively influenced the ability of this ST4/74 strain to colonize and proliferate in the gut at this time point. To define its diet-dependent gene expression pattern, S. Typhimurium was immunomagnetically isolated from the gut content, and its transcriptome was analyzed. A total of 66 genes were more strongly expressed in mice fed the plant-based diet. The majority of these genes was involved in metabolic functions degrading substrates of fruits and plants. Four of them are part of the gat gene cluster responsible for the uptake and metabolism of galactitol and D-tagatose. In line with this finding, 16S rRNA gene amplicon analysis revealed higher relative abundance of bacterial families able to degrade fiber and nutritive carbohydrates in PD-fed mice in comparison with those nourished with a WD. Competitive mice infection experiments performed with strain ST4/74 and ST4/74 ΔSTM3254 lacking tagatose-1,6-biphosphate aldolase, which is essential for galactitol and tagatose utilization, did not reveal a growth advantage of strain ST4/74 in the gastrointestinal tract of mice fed plant-based diet as compared to the deletion mutant., Conclusion: A Westernized diet and a plant-based diet evoke distinct transcriptional responses of S. Typhimurium during infection that allows the pathogen to adapt its metabolic activities to the diet-derived nutrients. This study therefore provides new insights into the dynamic interplay between nutrient availability, indigenous gut microbiota, and proliferation of S. Typhimurium.

Background: The gut microbiome impacts human health through various mechanisms and is involved in the development of a range of non-communicable diseases. Diet is a well-known factor influencing microbe-host interaction in health and disease. However, very few findings are based on large-scale analysis using population-based studies. Our aim was to investigate the cross-sectional relationship between habitual dietary intake and gut microbiota structure in the Cooperative Health Research in the Region of Augsburg (KORA) FF4 study., Results: Fecal microbiota was analyzed using 16S rRNA gene amplicon sequencing. Latent Dirichlet allocation (LDA) was applied to samples from 1992 participants to identify 20 microbial subgroups within the study population. Each participant's gut microbiota was subsequently described by a unique composition of these 20 subgroups. Associations between habitual dietary intake, assessed via repeated 24-h food lists and a Food Frequency Questionnaire, and the 20 subgroups, as well as between prevalence of metabolic diseases/risk factors and the subgroups, were assessed with multivariate-adjusted Dirichlet regression models. After adjustment for multiple testing, eight of 20 microbial subgroups were significantly associated with habitual diet, while nine of 20 microbial subgroups were associated with the prevalence of one or more metabolic diseases/risk factors. Subgroups 5 (Faecalibacterium, Lachnospiracea incertae sedis, Gemmiger, Roseburia) and 14 (Coprococcus, Bacteroides, Faecalibacterium, Ruminococcus) were particularly strongly associated with diet. For example, participants with a high probability for subgroup 5 were characterized by a higher Alternate Healthy Eating Index and Mediterranean Diet Score and a higher intake of food items such as fruits, vegetables, legumes, and whole grains, while participants with prevalent type 2 diabetes mellitus were characterized by a lower probability for subgroup 5., Conclusions: The associations between habitual diet, metabolic diseases, and microbial subgroups identified in this analysis not only expand upon current knowledge of diet-microbiota-disease relationships, but also indicate the possibility of certain microbial groups to be modulated by dietary intervention, with the potential of impacting human health. Additionally, LDA appears to be a powerful tool for interpreting latent structures of the human gut microbiota. However, the subgroups and associations observed in this analysis need to be replicated in further studies. Video abstract.

Anorexia nervosa (AN) is an eating disorder that leads to brain volume reduction and is difficult to treat since the underlying pathophysiology is poorly understood. The human gut microbiota is known to be involved in host metabolism, appetite- and bodyweight regulation, gut permeability, inflammation and gut-brain interactions. In this study, we used a translational activity-based anorexia (ABA) rat model including groups with food restriction, running-wheel access and a combination to disentangle the influences on the gut microbiota and associated changes in brain volume parameters. Our data demonstrated that chronic food restriction but not running-wheel activity had a major influence on the gut microbiota diversity and composition and reduced brain volume. Negative correlations were found between global brain weight and α-diversity, and astrocyte markers and relative abundances of the genera Odoribacter and Bifidobacterium. In contrast, the presence of lactobacilli was positively associated with white and grey brain matter volume. ABA and food-restricted rats are an interesting pre-clinical model to assess the causal influence of starvation on the gut microbiome and gut-brain interactions and can help to dissect the underlying pathophysiologic mechanisms relevant to AN., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Extibacter muris is a newly described mouse gut bacterium which metabolizes cholic acid (CA) to deoxycholic acid (DCA) via 7α-dehydroxylation. Although bile acids influence metabolic and inflammatory responses, few in vivo models exist for studying their metabolism and impact on the host. Mice were colonized from birth with the simplified community Oligo-MM 12 with or without E. muris . As the metabolism of bile acids is known to affect lipid homeostasis, mice were fed either a low- or high-fat diet for eight weeks before sampling and analyses targeting the gut and liver. Multiple Oligo-MM 12 strains were capable of deconjugating primary bile acids in vitro. E. muris produced DCA from CA either as pure compound or in mouse bile. This production was inducible by CA in vitro . Ursodeoxycholic, chenodeoxycholic, and β-muricholic acid were not metabolized under the conditions tested. All gnotobiotic mice were stably colonized with E. muris , which showed higher relative abundances after HF diet feeding. The presence of E. muris had minor, diet-dependent effects on Oligo-MM 12 communities. The secondary bile acids DCA and surprisingly LCA and their taurine conjugates were detected exclusively in E. muris -colonized mice. E. muris colonization did not influence body weight, white adipose tissue mass, liver histopathology, hepatic aspartate aminotransferase, or blood levels of cholesterol, insulin, and paralytic peptide (PP). However, proteomics revealed shifts in hepatic pathways involved in amino acid, glucose, lipid, energy, and drug metabolism in E. muris -colonized mice. Liver fatty acid composition was substantially altered by dietary fat but not by E. muris. In summary, E. muris stably colonized the gut of mice harboring a simplified community and produced secondary bile acids, which affected proteomes in the liver. This new gnotobiotic mouse model can now be used to study the pathophysiological role of secondary bile acids in vivo .

Living in a farm environment in proximity to animals is associated with reduced risk of developing allergies and asthma, and has been suggested to protect against other diseases, such as inflammatory bowel disease and cancer. Despite epidemiological evidence, experimental disease models that recapitulate such environments are needed to understand the underlying mechanisms. In this study, we show that feralizing conventional inbred mice by continuous exposure to a livestock farmyard-type environment conferred protection toward colorectal carcinogenesis. Two independent experimental approaches for colorectal cancer induction were used; spontaneous (Apc Min/+ mice on an A/J background) or chemical (AOM/DSS). In contrast to conventionally reared laboratory mice, the feralized mouse gut microbiota structure remained stable and resistant to mutagen- and colitis-induced neoplasia. Moreover, the feralized mice exhibited signs of a more mature immunophenotype, indicated by increased expression of NK and T-cell maturation markers, and a more potent IFN-γ response to stimuli. In our study, hygienically born and raised mice subsequently feralized post-weaning were protected to a similar level as life-long exposed mice, although the greatest effect was seen upon neonatal exposure. Collectively, we show protective implications of a farmyard-type environment on colorectal cancer development and demonstrate the utility of a novel animal modeling approach that recapitulates realistic disease responses in a naturalized mammal.

Our knowledge about the gut microbiota of pigs is still scarce, despite the importance of these animals for biomedical research and agriculture. Here, we present a collection of cultured bacteria from the pig gut, including 110 species across 40 families and nine phyla. We provide taxonomic descriptions for 22 novel species and 16 genera. Meta-analysis of 16S rRNA amplicon sequence data and metagenome-assembled genomes reveal prevalent and pig-specific species within Lactobacillus, Streptococcus, Clostridium, Desulfovibrio, Enterococcus, Fusobacterium, and several new genera described in this study. Potentially interesting functions discovered in these organisms include a fucosyltransferase encoded in the genome of the novel species Clostridium porci, and prevalent gene clusters for biosynthesis of sactipeptide-like peptides. Many strains deconjugate primary bile acids in in vitro assays, and a Clostridium scindens strain produces secondary bile acids via dehydroxylation. In addition, cells of the novel species Bullifex porci are coccoidal or spherical under the culture conditions tested, in contrast with the usual helical shape of other members of the family Spirochaetaceae. The strain collection, called 'Pig intestinal bacterial collection' (PiBAC), is publicly available at www.dsmz.de/pibac and opens new avenues for functional studies of the pig gut microbiota.

Bacillus cereus is a ubiquitous endospore-forming bacterium, which mainly affects humans as a food-borne pathogen. Bacillus cereus can contaminate groundwater used to irrigate food crops. Here, we examined the ability of the emetic strain B. cereus F4810/72 to survive abiotic conditions encountered in groundwater. Our results showed that vegetative B. cereus cells rapidly evolved in a mixed population composed of endospores and asporogenic variants bearing spo0A mutations. One asporogenic variant, VAR-F48, was isolated and characterized. VAR-F48 can survive in sterilized groundwater over a long period in a vegetative form and has a competitive advantage compared to its parental strain. Proteomics analysis allowed us to quantify changes to cellular and exoproteins after 24 and 72 h incubation in groundwater, for VAR-F48 compared to its parental strain. The results revealed a significant re-routing of the metabolism in the absence of Spo0A. We concluded that VAR-F48 maximizes its energy use to deal with oligotrophy, and the emergence of spo0A-mutated variants may contribute to the persistence of emetic B. cereus in natural oligotrophic environments., (© 2020 Society for Applied Microbiology and John Wiley & Sons Ltd.)

The gut is home to the body's largest population of plasma cells. In healthy individuals, IgA is the dominating isotype, whereas patients with inflammatory bowel disease also produce high concentrations of IgG. In the gut lumen, secretory IgA binds pathogens and toxins but also the microbiota. However, the antigen specificity of IgA and IgG for the microbiota and underlying mechanisms of antibody binding to bacteria are largely unknown. Here we show that microbiota binding is a defining property of human intestinal antibodies in both healthy and inflamed gut. Some bacterial taxa were commonly targeted by different monoclonal antibodies, whereas others selectively bound single antibodies. Interestingly, individual human monoclonal antibodies from both healthy and inflamed intestines bound phylogenetically unrelated bacterial species. This microbiota cross-species reactivity did not correlate with antibody polyreactivity but was crucially dependent on the accumulation of somatic mutations. Therefore, our data suggest that a system of affinity-matured, microbiota cross-species-reactive IgA is a common aspect of SIgA-microbiota interactions in the gut., Competing Interests: Disclosures: The authors declare no competing interests exist., (© 2020 Kabbert et al.)

Lifestyle, obesity, and the gut microbiome are important risk factors for metabolic disorders. We demonstrate in 1,976 subjects of a German population cohort (KORA) that specific microbiota members show 24-h oscillations in their relative abundance and identified 13 taxa with disrupted rhythmicity in type 2 diabetes (T2D). Cross-validated prediction models based on this signature similarly classified T2D. In an independent cohort (FoCus), disruption of microbial oscillation and the model for T2D classification was confirmed in 1,363 subjects. This arrhythmic risk signature was able to predict T2D in 699 KORA subjects 5 years after initial sampling, being most effective in combination with BMI. Shotgun metagenomic analysis functionally linked 26 metabolic pathways to the diurnal oscillation of gut bacteria. Thus, a cohort-specific risk pattern of arrhythmic taxa enables classification and prediction of T2D, suggesting a functional link between circadian rhythms and the microbiome in metabolic diseases., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Raw milk microbiota are complex communities with a significant impact on the hygienic, sensory and technological quality of milk products. However, there is a lack of knowledge on factors determining their composition. In the present study, four bulk tank milk samples of two farms at two different time points were analyzed in detail for their microbiota using cultivation and 16S rRNA amplicon sequencing. Diversity in samples from the first time point was assessed via cultivation of 500 aerobic mesophilic bacterial isolates in each sample. A high biodiversity of 70 and 110 species per sample was determined, of which 25-28% corresponded to yet unknown taxa. The isolates were dominated by Gram-positive members of the genera Staphylococcus, Corynebacterium, Streptococcus , or Janibacter , whilst Chryseobacterium and Acinetobacter were most abundant among the Gram-negative taxa. At the second time point, samples of the same farms were analyzed via both cultivation (1,500 individual colonies each) and high-throughput 16S rRNA gene amplicon sequencing. The latter revealed a threefold higher biodiversity at the genus level, as anaerobic or fastidious species were also detected. However, cultivation identified genera not captured by sequencing, indicating that both approaches are complementary. Using amplicon sequencing, the relative abundance of a few genera was distorted, which seems to be an artifact of sample preparation. Therefore, attention needs to be paid to the library preparation procedure with special emphasis on cell lysis and PCR., (Copyright © 2020 Breitenwieser, Doll, Clavel, Scherer and Wenning.)

Background: This study evaluated the efficacy, safety, and acceptability of a new ferrous sulfate oral solution (Tardyferon® 20 mg/mL) in young children with mild or moderate iron deficiency anemia (IDA)., Methods: This was a multicenter, national, single-arm, open-label study. Children aged 6-53 months presenting with mild or moderate IDA (i.e., blood hemoglobin (Hb) ranging from 7.0 to 10.9 g/dL and serum ferritin <12 ng/mL) were eligible for inclusion. The ferrous sulfate heptahydrate solution (2 mg/kg/day) was administered orally for 3 months. If normalization of either Hb or ferritin was not achieved at month 3 the treatment was continued for another 3 months., Results: Of the 100 children screened, 21 aged 6-17 months were included and received the study treatment, and 19 were analyzed for hematologic outcomes at month 3. Only one patient continued treatment for the additional 3 months. At month 3, mean ± SD Hb and ferritin levels were 12.0 ± 0.7 g/dL and 31.5 ± 19.4 ng/mL, respectively. Hemoglobin and ferritin levels were normalized in 95% (18/19) and 84% (16/19) of the patients, respectively. Treatment compliance and levels of satisfaction of both the parents and the investigators were high. Overall, 33.3% of patients (7/21) experienced at least one adverse event. Only one patient (4.8%) experienced a drug-related adverse event (upper abdominal pain)., Conclusions: A 2 mg/kg daily dose of the new oral ferrous sulfate heptahydrate solution provides substantial therapeutic benefit with high levels of tolerability in young children who have mild or moderate IDA., (© 2020 The Authors. Pediatrics International published by John Wiley & Sons Australia, Ltd on behalf of Japan Pediatric Society.)

The analysis of the gut microbiome with respect to health care prevention and diagnostic purposes is increasingly the focus of current research. We analyzed around 2000 stool samples from the KORA (Cooperative Health Research in the Region of Augsburg) cohort using high-throughput 16S rRNA gene amplicon sequencing representing a total microbial diversity of 2089 operational taxonomic units (OTUs). We evaluated the combination of three different components to assess the reflection of obesity related to microbiota profiles: (i) four prediction methods (i.e., partial least squares (PLS), support vector machine regression (SVMReg), random forest (RF), and M5Rules); (ii) five OTU data transformation approaches (i.e., no transformation, relative abundance without and with log-transformation, as well as centered and isometric log-ratio transformations); and (iii) predictions from nine measurements of obesity (i.e., body mass index, three measures of body shape, and five measures of body composition). Our results showed a substantial impact of all three components. The applications of SVMReg and PLS in combination with logarithmic data transformations resulted in considerably predictive models for waist circumference-related endpoints. These combinations were at best able to explain almost 40% of the variance in obesity measurements based on stool microbiota data (i.e., OTUs) only. A reduced loss in predictive performance was seen after sex-stratification in waist-height ratio compared to other waist-related measurements. Moreover, our analysis showed that the contribution of OTUs less prevalent and abundant is minor concerning the predictive power of our models.

The complexity of host-associated microbial ecosystems requires host-specific reference catalogs to survey the functions and diversity of these communities. We generate a comprehensive resource, the integrated mouse gut metagenome catalog (iMGMC), comprising 4.6 million unique genes and 660 metagenome-assembled genomes (MAGs), many (485 MAGs, 73%) of which are linked to reconstructed full-length 16S rRNA gene sequences. iMGMC enables unprecedented coverage and taxonomic resolution of the mouse gut microbiota; i.e., more than 92% of MAGs lack species-level representatives in public repositories (<95% ANI match). The integration of MAGs and 16S rRNA gene data allows more accurate prediction of functional profiles of communities than predictions based on 16S rRNA amplicons alone. Accompanying iMGMC, we provide a set of MAGs representing 1,296 gut bacteria obtained through complementary assembly strategies. We envision that integrated resources such as iMGMC, together with MAG collections, will enhance the resolution of numerous existing and future sequencing-based studies., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

The gastrointestinal tract is an ideal habitat for diverse bacterial species that reside in a homeostatic balance with local tissue and significantly contribute to host health. Negative shifts in gut microbiota profiles, also known as dysbiosis, may be implicated in the development of chronic disorders such as inflammatory bowel diseases (IBD). Adhesion molecule-dependent recruitment of immune cells to the gut is an important step in IBD pathogenesis. The adhesion molecule β7 integrin contributes to the development of the gut-associated lymphoid tissue (GALT), intestinal immune cell homing, and immune responses and is known to promote intestinal inflammation. Although many studies underlined the role of the gut microbiota in shaping the mucosal immune system, studies on the influence of the host immune system on the microbiota are rare, especially in homeostasis. We addressed this question via comparative 16S rRNA gene amplicon analysis of fecal microbial communities from wild-type and β7 integrin-deficient mice, the latter being characterized by a compromised GALT. Besides subtle changes in relative abundances of Muribaculaceae spp. and unknown members of the families Ruminococcaceae and Lachnospiraceae , there was altogether no major difference in microbiota profiles in β7 integrin-deficient mice vs. wild-type littermates. This indicates that, in conditions of homeostasis, there is only a minor influence of the host immune system on the fecal microbiota in our mouse model, stressing the potential importance of pathological factors for dysbiosis development., (Copyright © 2019 Babbar, Hitch, Pabst, Clavel, Hübel, Eswaran, Wagner and Schippers.)

The capacity of the Bacillus weihenstephanensis KBAB4 strain, a psychrotolerant species of the B. cereus sensu lato group, to multiply in carrot broth at 8 °C and 30 °C, in presence or absence of oxygen was determined. In aerobic carrot broth tyndallized in presence of oxygen, at both temperatures, the population of vegetative cells of B. weihenstephanensis inoculated at a level of 10 3 or 10 6  CFU/ml dropped immediately. After 16 h at 30 °C, B. weihenstephanensis reached around 10 3  CFU/ml, indicating that some vegetative cells had survived and multiplied, with lipid inclusions accumulated in cells, indicating possible stressing conditions. At 8 °C, no multiplication of B. weihenstephanensis was observed during 3 days to at least 12 days, depending of carrot broth batches. In anaerobic carrot broth tyndallized without oxygen, the vegetative cells of B. weihenstephanensis were not killed upon inoculation and multiplied in the broth at both 30 °C and 8 °C. Comparison with results from previous studies shows that B. weihenstephanensis behaves differently in carrot broth and in laboratory media at 8 °C with regards to presence or absence of oxygen., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Bacterial lipolytic enzymes represent an important class of proteins: they provide their host species with access to additional resources and have multiple applications within the biotechnology sector. Since the formalisation of lipolytic enzymes into families and subfamilies, advances in molecular biology have led to the discovery of lipolytic enzymes unable to be classified via the existing system. Utilising sequence-based comparison methods, we have integrated these novel families within the classification system so that it now consists of 35 families and 11 true lipase subfamilies. Representative sequences for each family and subfamily have been defined as well as methodology for accurate comparison of novel sequences against the reference proteins, facilitating the future assignment of novel proteins. Both the code and protein sequences required for integration of additional families are available at: https://github.com/thh32/Lipase&#95;reclassification., Competing Interests: The authors declare there are no competing interests.

A bacterial strain, designated WCA-9-b2 T , was isolated from the caecal content of an 18-week-old obese C57BL/6NTac male mouse. According to phenotypic analyses, the isolate was rod-shaped, strictly anaerobic, spore-forming, non-motile and Gram-stain-positive, under the conditions tested. Colonies were irregular and non-pigmented. Analysis of the 16S rRNA gene sequence indicated that the isolate belonged to the order Clostridiales with Dorea longicatena ATCC 27755 T (94.9 % sequence identity), Ruminococcus gnavus ATCC 29149 T (94.8%) and Clostridium scindens ATCC 35704 T (94.3%) being the closest relatives. Whole genome sequencing showed an average nucleotide identity <74.23 %, average amino acid identity <64.52-74.67 % and percentage of conserved proteins values <50 % against the nine closest relatives ( D. longicatena , Ruminococcus gnavus , C. scindens , Dorea formicigenerans , Ruminococcus lactaris , Clostridium hylemonae , Merdimonas faecis , Faecalicatena contorta and Faecalicatena fissicatena ). The genome-based G+C content of genomic DNA was 44.4 mol%. The major cellular fatty acids were C 16 : 0 (24.5%), C 18 : 1 cis 9 (19.8 %), C 16 : 0 DMA (11.7%), C 18 : 0 (8.4%) and C 14 : 0 (6.6%). Respiratory quinones were not detected. The predominant metabolic end products of glucose fermentation were acetate and succinate. Production of CO 2 and H 2 were detected. Based on these data, we propose that strain WCA-9-b2 T represents a novel species within a novel genus, for which the name Sporofaciens musculi gen. nov., sp. nov. is proposed. The type strain is WCA-9-b2 T (=DSM 106039 T =CECT 30156 T ).

Objective: The gut microbiota is an important influencing factor of metabolic health. Although dietary interventions with probiotics, prebiotics, and synbiotics can be effective means to regulate obesity and associated comorbidities, the underlying shifts in gut microbial communities, especially at the functional level, have not been characterized in great details. In this study, we sought to investigate the effects of synbiotics on the regulation of gut microbiota and the alleviation of high-fat diet (HFD)-induced metabolic disorders in mice., Methods: Specific pathogen-free (SPF) male C57BL/6J mice were fed diets with either 10% (normal diet, ND) or 60% (high-fat diet, HFD) of total calories from fat (lard). Dietary interventions in the HFD-fed mice included (i) probiotic (Bifidobacterium animalis subsp. lactis and Lactobacillus paracasei subsp. paracasei DSM 46331), (ii) prebiotic (oat β-glucan), and (iii) synbiotic (a mixture of i and ii) treatments for 12 weeks. Besides detailed characterization of host metabolic parameters, a multi-omics approach was used to systematically profile the microbial signatures at both the phylogenetic and functional levels using 16S rRNA gene sequencing, metaproteomics and targeted metabolomics analysis., Results: The synbiotic intervention significantly reduced body weight gain and alleviated features of metabolic complications. At the phylogenetic level, the synbiotic treatment significantly reversed HFD-induced changes in microbial populations, both in terms of richness and the relative abundance of specific taxa. Potentially important species such as Faecalibaculum rodentium and Alistipes putredinis that might mediate the beneficial effects of the synbiotic were identified. At the functional level, short-chain fatty acid and bile acid profiles revealed that all dietary interventions significantly restored cecal levels of acetate, propionate, and butyrate, while the synbiotic treatment reduced the bile acid pools most efficiently. Metaproteomics revealed that the effects of the synbiotic intervention might be mediated through metabolic pathways involved in carbohydrate, amino acid, and energy metabolisms., Conclusions: Our results suggested that dietary intervention using the novel synbiotic can alleviate HFD-induced weight gain and restore gut microbial ecosystem homeostasis phylogenetically and functionally., (Copyright © 2019 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Background: Bacteria within family S24-7 (phylum Bacteroidetes) are dominant in the mouse gut microbiota and detected in the intestine of other animals. Because they had not been cultured until recently and the family classification is still ambiguous, interaction with their host was difficult to study and confusion still exists regarding sequence data annotation., Methods: We investigated family S24-7 by combining data from large-scale 16S rRNA gene analysis and from functional and taxonomic studies of metagenomic and cultured species., Results: A total of 685 species was inferred by full-length 16S rRNA gene sequence clustering. While many species could not be assigned ecological habitats (93,045 samples analyzed), the mouse was the most commonly identified host (average of 20% relative abundance and nine co-occurring species). Shotgun metagenomics allowed reconstruction of 59 molecular species, of which 34 were representative of the 16S rRNA gene-derived species clusters. In addition, cultivation efforts allowed isolating five strains representing three species, including two novel taxa. Genome analysis revealed that S24-7 spp. are functionally distinct from neighboring families and versatile with respect to complex carbohydrate degradation., Conclusions: We provide novel data on the diversity, ecology, and description of bacterial family S24-7, for which the name Muribaculaceae is proposed.

Background/aims: The gut microbiota is altered in irritable bowel syndrome (IBS), and microbiota manipulations by diet or antibiotics can reduce its symptoms. As fecal microbiota transfer (FMT) in IBS is still controversial, we investigated the clinical and side effects of FMT in a cohort of IBS patients with recurrent, treatment refractory symptoms, and studied gut microbiota signatures., Methods: Using an observational, prospective study design, we applied FMTs from one unrelated, healthy donor to 13 IBS patients. Fecal samples of patients and the donor were analyzed by 16S ribosomal RNA amplicon sequencing., Results: On a symptom level, primarily abdominal pain symptoms were reduced after FMT, and no adverse effects were observed. Studying the microbiome, we found an increase in alpha diversity and changes in the composition of the gut microbiota after FMT. Beta diversity changes after FMT were prominent in a subset of 7 patients with microbiota profiles coming very close to the donor. These patients also showed most pronounced visceral pain reduction. The relative abundance of Akkermansia muciniphila was inversely correlated with pain reduction in our cohort., Conclusion: Although exploratory in nature and with a pilot character, this study highlights the potential role of microbiota manipulations in IBS and describes a novel association of intestinal Akkermansia and pain modulation., (© 2018 S. Karger AG, Basel.)

Background & Aims: Activating transcription factor 6 (ATF6) regulates endoplasmic reticulum stress. We studied whether ATF6 contributes to the development of colorectal cancer (CRC) using tissue from patients and transgenic mice., Methods: We analyzed data from 541 patients with CRC in The Cancer Genome Atlas database for genetic variants and aberrant expression levels of unfolded protein response genes. Findings were validated in a cohort of 83 patients with CRC in Germany. We generated mice with intestinal epithelial cell-specific expression of the active form of Atf6 (nATF6IEC) from 2 alleles (homozygous), mice with expression of nATF6IEC from 1 allele (heterozygous), and nATF6IECfl/fl mice (controls). All nATF6IEC mice were housed under either specific-pathogen-free or germ-free conditions. Cecal microbiota from homozygous nATF6IEC mice or control mice was transferred into homozygous nATF6IEC mice or control mice. nATF6IEC mice were crossed with mice with disruptions in the myeloid differentiation primary response gene 88 and toll-like receptor adaptor molecule 1 gene (Myd88/Trif-knockout mice). Intestinal tissues were collected from mice and analyzed by histology, immunohistochemistry, immunoblots, gene expression profiling of unfolded protein response and inflammatory genes, array-based comparative genome hybridization, and 16S ribosomal RNA gene sequencing., Results: Increased expression of ATF6 was associated with reduced disease-free survival times of patients with CRC. Homozygous nATF6IEC mice developed spontaneous colon adenomas at 12 weeks of age. Compared with controls, homozygous nATF6IEC mice had changes in the profile of their cecal microbiota, increased proliferation of intestinal epithelial cells, and loss of the mucus barrier-all preceding tumor formation. These mice had increased penetration of bacteria into the inner mucus layer and activation of signal transducer and activator of transcription 3, yet inflammation was not observed at the pretumor or tumor stages. Administration of antibiotics to homozygous nATF6IEC mice greatly reduced tumor incidence, and germ-free housing completely prevented tumorigenesis. Analysis of nATF6IEC MyD88/TRIF-knockout mice showed that tumor initiation and growth required MyD88/TRIF-dependent activation of signal transducer and activator of transcription 3. Transplantation of cecal microbiota from nATF6IEC mice and control mice, collected before tumor formation, caused tumor formation in ex-germ-free nATF6IEC mice., Conclusions: In patients with CRC, ATF6 was associated with reduced time of disease-free survival. In studies of nATF6IEC mice, we found sustained intestinal activation of ATF6 in the colon to promote dysbiosis and microbiota-dependent tumorigenesis., (Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Interactions between the gut microbial ecosystem and host lipid homeostasis are highly relevant to host physiology and metabolic diseases. We present a comprehensive multi-omics view of the effect of intestinal microbial colonization on hepatic lipid metabolism, integrating transcriptomic, proteomic, phosphoproteomic, and lipidomic analyses of liver and plasma samples from germfree and specific pathogen-free mice. Microbes induce monounsaturated fatty acid generation by stearoyl-CoA desaturase 1 and polyunsaturated fatty acid elongation by fatty acid elongase 5, leading to significant alterations in glycerophospholipid acyl-chain profiles. A composite classification score calculated from the observed alterations in fatty acid profiles in germfree mice clearly differentiates antibiotic-treated mice from untreated controls with high sensitivity. Mechanistic investigations reveal that acetate originating from gut microbial degradation of dietary fiber serves as precursor for hepatic synthesis of C16 and C18 fatty acids and their related glycerophospholipid species that are also released into the circulation.

Background: As the gut microbiota contributes to metabolic health, it is important to determine specific diet-microbiota interactions that influence host metabolism. Bile acids and dietary fat source can alter phenotypes of diet-induced obesity, but the interplay with intestinal microorganisms is unclear. Here, we investigated metabolic consequences of diets enriched in primary bile acids with or without addition of lard or palm oil, and studied gut microbiota structure and functions in mice., Results: In combination with bile acids, dietary lard fed to male C57BL/6N mice for a period of 8 weeks enhanced fat mass accumulation in colonized, but not in germ-free mice when compared to palm oil. This was associated with impaired glucose tolerance, lower fasting insulin levels, lower counts of enteroendocrine cells, fatty liver, and elevated amounts of hepatic triglycerides, cholesteryl esters, and monounsaturated fatty acids. Lard- and bile acid-fed mice were characterized by shifts in dominant gut bacterial communities, including decreased relative abundances of Lachnospiraceae and increased occurrence of Desulfovibrionaceae and the species Clostridium lactatifermentans and Flintibacter butyricus. Metatranscriptomic analysis revealed shifts in microbial functions, including lipid and amino acid metabolism., Conclusions: Caution is required when interpreting data from diet-induced obesity models due to varying effects of dietary fat source. Detrimental metabolic consequences of a diet enriched with lard and primary bile acids were dependent on microbial colonization of the host and were linked to hepatic lipid rearrangements and to alterations of dominant bacterial communities in the cecum.

Background & Aims: Antibiotic (ABx) therapy is associated with increased risk for Crohn's disease but underlying mechanisms are unknown. We observed high fecal serine protease activity (PA) to be a frequent side effect of ABx therapy. The aim of the present study was to unravel whether this rise in large intestinal PA may promote colitis development via detrimental effects on the large intestinal barrier., Methods: Transwell experiments were used to assess the impact of high PA in ABx-treated patients or vancomycin/metronidazole-treated mice on the epithelial barrier. Serine protease profiling was performed using liquid chromatography-mass spectrometry/mass spectrometry analysis. The impact of high large intestinal PA on the intestinal barrier in wild-type and interleukin (IL)10 -/- mice and on colitis development in IL10 -/- mice was investigated using vancomycin/metronidazole with or without oral serine protease inhibitor (AEBSF) treatment., Results: The ABx-induced, high large intestinal PA was caused by significantly increased levels of pancreatic proteases and impaired epithelial barrier integrity. In wild-type mice, the rise in PA caused a transient increase in intestinal permeability but did not affect susceptibility to chemically induced acute colitis. In IL10 -/- mice, increased PA caused a consistent impairment of the intestinal barrier associated with inflammatory activation in the large intestinal tissue. In the long term, the vancomycin/metronidazole-induced lasting increase in PA aggravated colitis development in IL10 -/- mice., Conclusions: High large intestinal PA is a frequent adverse effect of ABx therapy, which is detrimental to the large intestinal barrier and may contribute to the development of chronic intestinal inflammation in susceptible individuals.

Inflammatory bowel disease (IBD) is associated with enhanced levels of the IL-1 family cytokines IL-1β and IL-18, which are activated by the Nlrp3 inflammasome. Here, we investigated the role of inflammasome-driven cytokine release on T cell polarization and DC differentiation in steady state and T cell transfer colitis. In vitro and in vivo data showed that IL-1β induces Th17 polarization and increases GM‑CSF production by T cells. Reduced IL-1β levels in Nlrp3-/- mice correlated with enhanced FLT3L levels and increased frequency of tolerogenic CD103+ DC. In the T cell transfer colitis model, Nlrp3 deficiency resulted in lower IL‑1β levels, reduced Th17 immunity, and less severe colitis. Unaltered IL-18 levels in both mouse strains pointed toward Nlrp3-independent processing. Importantly, cohousing revealed that the gut microbiome had no impact on the observed Nlrp3-/- phenotype. This study demonstrates that NLRP3 acts as a molecular switch of intestinal homeostasis by shifting local immune cells toward an inflammatory phenotype via IL-1β.

Background: High-fat diets (HFDs) have been linked to low-grade inflammation and insulin resistance., Objective: The main purpose of the present study was to assess whether acute overfeeding with an HFD affects insulin sensitivity, gut barrier function, and fecal microbiota in humans., Methods: In a prospective intervention study, 24 healthy men [mean ± SD: age 23.0 ± 2.8 y, body mass index (in kg/m2) 23.0 ± 2.1] received an HFD (48% of energy from fat) with an additional 1000 kcal/d (as whipping cream) above their calculated energy expenditure for 7 d. Insulin sensitivity (hyperinsulinemic euglycemic clamp), gut permeability (sugar and polyethylene glycol absorption tests, plasma zonulin), and gut microbiota profiles (high-throughput 16S rRNA gene sequencing) were assessed before and after overfeeding, and 14 d after intervention. Additionally, inflammation markers such as high-sensitivity C-reactive protein, lipopolysaccharide-binding protein, leptin, high-molecular-weight adiponectin, calprotectin, regulated on activation normal, T cell expressed and secreted (RANTES), and monocyte chemoattractant protein-1 were measured in plasma by ELISA. Finally, lipid parameters were analyzed in serum by a laboratory service., Results: Although participants gained 0.9 ± 0.6 kg (P < 0.001) body weight, overnutrition was not associated with a significant change in insulin sensitivity (M value and glucose disposal). Overfeeding for 7 d resulted in elevated serum total (10.2%), LDL (14.6%) and HDL (14.8%) cholesterol concentrations (P < 0.01). In contrast, fasting plasma triglyceride significantly declined (29.3%) during overfeeding (P < 0.001). In addition, there were no significant changes in inflammatory markers. Urine excretion of 4 sugars and polyethylene glycol, used as a proxy for gut permeability, and plasma concentration of zonulin, a marker of paracellular gut permeability, were unchanged. Moreover, overfeeding was not associated with consistent changes in gut microbiota profiles, but marked alterations were observed in a subgroup of 6 individuals., Conclusions: Our findings suggest that short-term overfeeding with an HFD does not significantly impair insulin sensitivity and gut permeability in normal-weight healthy men, and that changes in dominant communities of fecal bacteria occur only in certain individuals. The study was registered in the German Clinical Trial Register as DRKS00006211., (© 2018 American Society for Nutrition. All rights reserved.)

Background: Early-life colonization of the intestinal tract is a dynamic process influenced by numerous factors. The impact of probiotic-supplemented infant formula on the composition and function of the infant gut microbiota is not well defined. Objective: We sought to determine the effects of a bifidobacteria-containing formula on the healthy human intestinal microbiome during the first year of life. Design: A double-blind, randomized, placebo-controlled study of newborn infants assigned to a standard whey-based formula containing a total of 10 7 colony-forming units (CFU)/g of Bifidobacterium bifidum , Bifidobacterium breve , Bifidobacterium longum , B. longum subspecies infantis (intervention), or to a control formula without bifidobacteria (placebo). Breastfed controls were included. Diversity and composition of fecal microbiota were determined by 16S ribosomal RNA gene amplicon sequencing, and metabolite profiles were analyzed by ultrahigh-performance liquid chromatography-mass spectrometry over a period of 2 y. Results: Infants ( n = 106) were randomly assigned to either the interventional ( n = 48) or placebo ( n = 49) group; 9 infants were exclusively breastfed throughout the entire intervention period of 12 mo. Infants exposed to bifidobacteria-supplemented formula showed decreased occurrence of Bacteroides and Blautia spp. associated with changes in lipids and unknown metabolites at month 1. Microbiota and metabolite profiles of intervention and placebo groups converged during the study period, and long-term colonization (24 mo) of the supplemented Bifidobacterium strains was not detected. Significant differences in microbiota and metabolites were detected between infants fed breast milk and those fed formula ( P < 0.005) and between infants birthed vaginally and those birthed by cesarean delivery ( P < 0.005). No significant differences were observed between infant feeding groups regarding growth, antibiotic uptake, or other health variables ( P > 0.05). Conclusion: The supplementation of bifidobacteria to infant diet can modulate the occurrence of specific bacteria and metabolites during early life with no detectable long-term effects. This trial was registered at germanctr.de as DRKS00003660., (© 2017 American Society for Nutrition.)

Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Numerous iron-containing preparations are available on the market; these vary in dosage, salt, chemical state of iron (ferric or ferrous) and in the iron delivery process (immediate or prolonged-release). Tardyferon® is a prolonged-release tablet containing 80 mg ferrous sulphate. The formulation has recently been modified; changes to the excipients which constitute the inert formulation matrix have allowed a decrease in tablet size for easier swallowing. The aim of this multicenter open-label study was to characterize the serum pharmacokinetics of iron in non-pregnant women aged 23-45 years with iron deficiency anaemia (IDA) following single oral administration of 160 mg Tardyferon® under fasting conditions. Blood samples were collected from the 29 participants before dosing and until 24 h post-dosing. Serum iron concentrations were determined using a routine colorimetric analytical method; pharmacokinetic parameters were derived using a non-compartmental approach. In these patients, median time to maximum serum concentrations (Tmax) was 4 h. Serum profiles were consistent with prolonged release; iron levels were elevated up to 12 h after dosing, with mean C 12h still more than 7 times higher than baseline (C T0 ), and mean C 2h and C 8h representing 69.7% and 81.9% of the C max , respectively. In vitro dissolution testing performed on the clinical batch also demonstrated prolonged release of iron from this formulation. A single oral dose of 160 mg Tardyferon® administered under fasting conditions to this target population resulted in a long-lasting release of iron in the gastrointestinal tract, leading to optimal iron absorption. Moreover, Tardyferon® was well tolerated., Competing Interests: Conflict of Interest: Dr A. LEARY works for a Regulatory Affairs Consultancy which had provided paid consultancy services to Pierre Fabre Medicament. L. BARTHE, T. CLAVEL, C. SANCHEZ,, Z. ISSIAKHEM, B. PAILLARD, J.M. EDMOND, are employees of Pierre Fabre Medicament, which provided financial and material support for the design and concept of the study, data collection, data management, data analysis and the publication of this study., (© Georg Thieme Verlag KG Stuttgart · New York.)

Bile acids (BA) are signaling molecules with a wide range of biological effects, also identified among the most responsive plasma metabolites in the postprandial state. We here describe this response to different dietary challenges and report on key determinants linked to its interindividual variability. Healthy men and women ( n = 72, 62 ± 8 yr, mean ± SE) were enrolled into a 12-wk weight loss intervention. All subjects underwent an oral glucose tolerance test and a mixed-meal tolerance test before and after the intervention. BA were quantified in plasma by liquid chromatography-tandem mass spectrometry combined with whole genome exome sequencing and fecal microbiota profiling. Considering the average response of all 72 subjects, no effect of the successful weight loss intervention was found on plasma BA profiles. Fasting and postprandial BA profiles revealed high interindividual variability, and three main patterns in postprandial BA response were identified using multivariate analysis. Although the women enrolled were postmenopausal, effects of sex difference in BA response were evident. Exome data revealed the contribution of preselected genes to the observed interindividual variability. In particular, a variant in the SLCO1A2 gene, encoding the small intestinal BA transporter organic anion-transporting polypeptide-1A2 (OATP1A2), was associated with delayed postprandial BA increases. Fecal microbiota analysis did not reveal evidence for a significant influence of bacterial diversity and/or composition on plasma BA profiles. The analysis of plasma BA profiles in response to two different dietary challenges revealed a high interindividual variability, which was mainly determined by genetics and sex difference of host with minimal effects of the microbiota. NEW & NOTEWORTHY Considering the average response of all 72 subjects, no effect of the successful weight loss intervention was found on plasma bile acid (BA) profiles. Despite high interindividual variability, three main patterns in postprandial BA response were identified using multivariate analysis. A variant in the SLCO1A2 gene, encoding the small intestinal BA transporter organic anion-transporting polypeptide-1A2 (OATP1A2), was associated with delayed postprandial BA increases in response to both the oral glucose tolerance test and the mixed-meal tolerance test., (Copyright © 2017 the American Physiological Society.)

Recent findings suggest an association between obesity, loss of gut barrier function and changes in microbiota profiles. Our primary objective was to examine the effect of caloric restriction and subsequent weight reduction on gut permeability in obese women. The impact on inflammatory markers and fecal microbiota was also investigated. The 4-week very-low calorie diet (VLCD, 800 kcal/day) induced a mean weight loss of 6.9 ± 1.9 kg accompanied by a reduction in HOMA-IR (Homeostasis model assessment-insulin resistance), fasting plasma glucose and insulin, plasma leptin, and leptin gene expression in subcutaneous adipose tissue. Plasma high-molecular weight adiponectin (HMW adiponectin) was significantly increased after VLCD. Plasma levels of high-sensitivity C-reactive protein (hsCRP) and lipopolysaccharide-binding protein (LBP) were significantly decreased after 28 days of VLCD. Using three different methods, gut paracellular permeability was decreased after VLCD. These changes in clinical parameters were not associated with major consistent changes in dominant bacterial communities in feces. In summary, a 4-week caloric restriction resulted in significant weight loss, improved gut barrier integrity and reduced systemic inflammation in obese women.

During the last 15 years, molecular techniques have been preferred over culture-based approaches for the study of mammalian gut microbiota, i.e. the communities of microorganisms dwelling in the intestine of mammals. The main reason is the belief that the majority of gut bacteria, especially strict anaerobes, escape cultivation. Despite numerous such statements in publications, the literature does not provide a clear overview on the subject. In the present manuscript, we highlight recent work on the cultivation of bacteria from the intestine of mammals, review the literature and provide novel data pertaining to cultured fractions of mammalian gut microbiota. These data show that, despite marked inter-sample variations, 35 to 65% of molecular species detected by sequencing have representative strains in culture.