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Modeling microbial systems as sparse and reproducible networks is a major challenge in network inference. Direct interactions between the microbial species of a given biome can help to understand how the microbial communities influence the system and through which mechanisms. Most state of the art methods reconstruct network from abundance data using Gaussian Graphical Models (GGM) for which statistically grounded and efficient inference approaches are available: Meinshausen-Bühlmann (MB) neighborhood selection, graphical Lasso (glasso), tree averaging approach, etc.In this article, we examine eight inference techniques and propose a two-step consensus method to combine them. All methods methods rely on stability selection, a resampling-based procedure to choose a regularization parameter by evaluating the selection frequency of selecting edges in the constructed networks. However, stability selection can result in very different precision - recall trade-offs for different methods, even when they have identical precision - recall curves. Rather than selecting an optimal regularization parameter and disregarding the selection frequencies, as is performed in stability selection, we suggest to combine edge frequencies directly to reconstruct the network. This approach ensures that only robust and reproducible edges are included in the consensus network.The effectiveness of the consensus method compared to individual methods is demonstrated on synthetic data derived from a set of healthy microbiota and then applied to a real study-case of microbiota from patients with liver cirrhosis. While the performances of all methods in terms of precision/recall are quite similar, the MB-based ones tend to outperform glasso-based ones and are in turn outperformed by the consensus method.

Supplementary Figure from A Natural Polyphenol Exerts Antitumor Activity and Circumvents Anti–PD-1 Resistance through Effects on the Gut Microbiota

Several approaches to manipulate the gut microbiome for improving the activity of cancer immune-checkpoint inhibitors (ICI) are currently under evaluation. Here, we show that oral supplementation with the polyphenol-rich berry camu-camu (CC; Myrciaria dubia) in mice shifted gut microbial composition, which translated into antitumor activity and a stronger anti–PD-1 response. We identified castalagin, an ellagitannin, as the active compound in CC. Oral administration of castalagin enriched for bacteria associated with efficient immunotherapeutic responses (Ruminococcaceae and Alistipes) and improved the CD8+/FOXP3+CD4+ ratio within the tumor microenvironment. Moreover, castalagin induced metabolic changes, resulting in an increase in taurine-conjugated bile acids. Oral supplementation of castalagin following fecal microbiota transplantation from ICI-refractory patients into mice supported anti–PD-1 activity. Finally, we found that castalagin binds to Ruminococcus bromii and promoted an anticancer response. Altogether, our results identify castalagin as a polyphenol that acts as a prebiotic to circumvent anti–PD-1 resistance.Significance:The polyphenol castalagin isolated from a berry has an antitumor effect through direct interactions with commensal bacteria, thus reprogramming the tumor microenvironment. In addition, in preclinical ICI-resistant models, castalagin reestablishes the efficacy of anti–PD-1. Together, these results provide a strong biological rationale to test castalagin as part of a clinical trial.This article is highlighted in the In This Issue feature, p. 873

International audience; Recent advances in the human microbiome characterization have revealed significant oral microbial detection in stools of dysbiotic patients. However, little is known about the potential interactions of these invasive oral microorganisms with commensal intestinal microbiota and the host. In this proof-of-concept study, we proposed a new model of oral-to-gut invasion by the combined use of an in vitro model simulating both the physicochemical and microbial (lumen-and mucus-associated microbes) parameters of the human colon (M-ARCOL), a salivary enrichment protocol, and whole-metagenome shotgun sequencing. Oral invasion of the intestinal microbiota was simulated by injection of enriched saliva in the in vitro colon model inoculated with a fecal sample from the same healthy adult donor. The mucosal compartment of M-ARCOL was able to retain the highest species richness levels over time, while species richness levels decreased in the luminal compartment. This study also showed that oral microorganisms preferably colonized the mucosal microenvironment, suggesting potential oral-to-intestinal mucosal competitions. This new model of oral-to-gut invasion can provide useful mechanistic insights into the role of oral microbiome in various disease processes. IMPORTANCE Here, we propose a new model of oral-to-gut invasion by the combined use of an in vitro model simulating both the physicochemical and microbial (lumen-and mucus-associated microbes) parameters of the human colon (M-ARCOL), a salivary enrichment protocol, and whole-metagenome shotgun sequencing. Our study revealed the importance of integrating the mucus compartment, which retained higher microbial richness during fermentation, showed the preference of oral microbial invaders for the mucosal resources, and indicated potential oral-to-intestinal mucosal competitions. It also underlined promising opportunities to further understand mechanisms of oral invasion into the human gut microbiome, define microbe-microbe and mucus-microbe interactions in a compartmentalized fashion, and help to better characterize the potential of oral microbial invasion and their persistence in the gut.

Background Antibiotics notoriously perturb the gut microbiota. We used untargeted and targeted phenotypic and genotypic approaches to study faecal samples collected up to 90 days following a 3-day course of intravenous β-lactam antibiotics in 22 healthy volunteers. We studied the changes in the bacterial, phage and fungal components of the microbiota as well as the metabolome and the β-lactamase activity of the stools. This allowed assessing their degrees of perturbation and resilience. Results While only two subjects had detectable concentrations of antibiotics in their faeces, suggesting important antibiotic degradation in the gut, the intravenous treatment perturbed very significantly the bacterial and phage microbiota, as well as the composition of the metabolome. In contrast, treatment impact was relatively low on the fungal microbiota. At the end of the surveillance period, we found evidence of resilience across the gut system since most components returned to a state like the initial one, even if the taxonomic composition of the bacterial microbiota changed and the dynamics of the different components over time were rarely correlated. The richness of the resistome was significantly reduced up to day 30, while a significant increase in the relative abundance of β-lactamase encoding genes was observed up to day 10, consistent with a concomitant increase in the β-lactamase activity of the microbiota. The level of β-lactamase activity at baseline was positively associated with the resilience of the metabolome content of the stools. Conclusions In healthy adults, antibiotics perturb all the components of the microbiota, which mostly return to its baseline state within 30 days. These data suggest an important role of endogenous β-lactamases producing anaerobes in protecting the functions of the microbiota by de-activating the antibiotics reaching the colon.

Recent advances in the human microbiome characterization have revealed significant oral microbial detection in stools of dysbiotic patients. However, little is known about the potential interactions of these invasive oral microorganisms with commensal intestinal microbiota and host. In this proof of concept study, we propose a new model of oral to gut invasion by the combined use of anin vitromodel simulating both the physicochemical and microbial (lumen and mucus-associated microbes) parameters of the human colon (M-ARCOL), a salivary enrichment protocol and whole metagenome shotgun sequencing. Oral invasion of the intestinal microbiota was simulated by injection of enriched saliva in thein vitrocolon model inoculated with faecal sample from the same healthy adult donor. The mucosal compartment of M-ARCOL was able to retain the highest species richness levels over time, whilst it decreased in the luminal compartment. This study also showed that oral microorganisms preferably colonized the mucosal microenvironment, suggesting potential oral-to-intestinal mucosal competitions. This new model of oral-to-gut invasion can provide useful mechanistic insights into the role of oral microbiome in various disease processes.

S10.3 The mycobiome characterization: future perspectives or just a trend?, September 24, 2022, 10:30 AM - 12:00 PM Objectives Candida albicans is both a harmless commensal intestinal yeast in healthy individuals and a harmful opportunistic pathogen in immunocompromised patients, causing life-threatening invasive candidiasis. C. albicans intestinal overgrowth is a prerequisite for intestinal translocation, which is at the root of invasive candidiasis. Therefore, to prevent invasive candidiasis in immunocompromised patients, it is necessary to curb intestinal C. albicans overgrowth. However, little is known of the role of bacterial species of the microbiota in dampening C. albicans intestinal colonization. We aimed to decipher the influence of the bacterial and fungal intestinal microbiota on C. albicans gut colonization in healthy individuals in whom their microbiota was modified or not by antibiotic treatment. Methods We studied two cohorts of healthy individuals: the first cohort included 22 volunteers for which fecal samples were collected before, during and after a 3-day regimen of third-generation cephalosporin antibiotics. The second cohort gathered 1000 healthy individuals for which a single fecal sample was collected. We quantified C. albicans carriage using a specific quantitative PCR approach. We monitored the antibiotic effect on the composition of the fungal microbiota—the so called mycobiota—in the gut of the individuals up to 180 days post-antibiotic exposure, using both the qPCR data and ITS1 targeted metagenomic. We also monitored the level of fecal β-lactamase activity, which is known to modulate the intensity of post-cephalosporin intestinal dysbiosis. Finally, to identify potential C. albicans inhibiting bacteria, we used MaAsLin2 to search for associations between C. albicans levels and bacterial species abundance, deduced from shotgun metagenomics data obtained from all individuals and annotated at the species level. Results A very high level of C. albicans carriage was observed in both cohorts, with a prevalence of 95.2% and 83.1% in the first and second cohort, respectively. Yet, the quantity of C. albicans DNA detected varied greatly between subjects. The mycobiota composition was significantly altered by antibiotics and the fungal load was increased both at short and long term. Particularly, C. albicans abundance was increased but with wide inter-subject variations. A part of these variations was explained by changes in the levels of endogenous fecal β-lactamase activity, with subjects characterized by a low increase of β-lactamase activity displaying a higher increase of C. albicans levels. Finally, using shotgun metagenomics data, we identified a first set of 50 bacterial and archaeal species whose abundance inversely correlated with C. albicans abundance. Conclusion These results (1) bring a new understanding of C. albicans overgrowth in healthy individuals, (2) lead to the identification of microbial signatures with a potential key role in controlling C. albicans gut colonization, and finally (3) show for the first time that changes in endogenous fecal β-lactamase activity is a key factor for C. albicans overgrowth in the gut after antibiotic exposure. Taken together, these results open the way for new intervention strategies to curb C. albicans intestinal overgrowth.

We gratefully acknowledge the kind contribution of Mrs. Nabila Yasmine Domingo-Saidji from the Clinical Research Unit of Paris Ile-de-France Ouest (URCPO), who supervised the administrative paperwork and helped us to manage ethical issues related to the study.; International audience; Objective: Gut microbiome dysbiosis has previously been reported in spondyloarthritis (SpA) patients and could be critically involved in the pathogenesis of this disorder. The objectives of this study were to further characterize the microbiota structure in SpA patients and to investigate the relationship between dysbiosis and disease activity in light of the putative influence of the genetic background. Methods: Shotgun sequencing was performed on fecal DNA isolated from stool samples from 2 groups of adult volunteers: SpA patients (n = 102) and healthy controls (n = 63). A subset of the healthy controls comprised the age-matched siblings of patients whose HLA–B27 status was known. Changes in gut microbiota composition were assessed based on species diversity, enterotypes, and taxonomic and functional differences. Results: Dysbiosis was confirmed in SpA patients as compared to healthy controls. The restriction of microbiota diversity was detected in patients with the most active disease, and the abundance of several bacterial species was correlated with Bath Ankylosing Spondylitis Disease Activity Index score. Among healthy controls, significant differences in microbiota composition were also detected between the HLA–B27–positive and the HLA–B27–negative siblings of SpA patients. We highlighted a decreased abundance of several species of bacteria in SpA patients, especially those bacteria belonging to the Clostridiales order. Among the few species of bacteria showing increased abundance, Ruminococcus gnavus was one of the top differentiating species. Conclusion: These findings reveal that genetic background and level of disease activity are likely to influence the composition of the gut microbiota of patients with SpA. It may be appropriate for further research on chronic arthritis to focus on these key parameters

An imbalance of the gut microbiota ("dysbiosis") is associated with numerous chronic diseases, and its modulation is a promising novel therapeutic approach. Dietary supplementation with soluble fiber is one of several proposed modulation strategies. This study aims at confirming the impact of the resistant dextrin NUTRIOSE (RD), a soluble fiber with demonstrated beneficial health effects, on the gut microbiota of healthy individuals.Fifty healthy women are enrolled and supplemented daily with either RD (n = 24) or a control product (n = 26) during 6 weeks. Characterization of the fecal metagenome with shotgun sequencing reveals that RD intake dramatically increases the abundance of the commensal bacterium Parabacteroides distasonis. Furthermore, presence in metagenomes of accessory genes from P. distasonis, coding for susCD (a starch-binding membrane protein complex) is associated with a greater increase of the species. This suggests that response to RD might be strain-dependent.Supplementation with RD can be used to specifically increase P. distasonis in gut microbiota of healthy women. The magnitude of the response may be associated with fiber-metabolizing capabilities of strains carried by subjects. Further research will seek to confirm that P. distasonis directly modulates the clinical effects observed in other studies.

Previous microbiome and metabolome analyses exploring non-communicable diseases have paid scant attention to major confounders of study outcomes, such as common, pre-morbid and co-morbid conditions, or polypharmacy. Here, in the context of ischemic heart disease (IHD), we used a study design that recapitulates disease initiation, escalation and response to treatment over time, mirroring a longitudinal study that would otherwise be difficult to perform given the protracted nature of IHD pathogenesis. We recruited 1,241 middle-aged Europeans, including healthy individuals, individuals with dysmetabolic morbidities (obesity and type 2 diabetes) but lacking overt IHD diagnosis and individuals with IHD at three distinct clinical stages-acute coronary syndrome, chronic IHD and IHD with heart failure-and characterized their phenome, gut metagenome and serum and urine metabolome. We found that about 75% of microbiome and metabolome features that distinguish individuals with IHD from healthy individuals after adjustment for effects of medication and lifestyle are present in individuals exhibiting dysmetabolism, suggesting that major alterations of the gut microbiome and metabolome might begin long before clinical onset of IHD. We further categorized microbiome and metabolome signatures related to prodromal dysmetabolism, specific to IHD in general or to each of its three subtypes or related to escalation or de-escalation of IHD. Discriminant analysis based on specific IHD microbiome and metabolome features could better differentiate individuals with IHD from healthy individuals or metabolically matched individuals as compared to the conventional risk markers, pointing to a pathophysiological relevance of these features. ispartof: NATURE MEDICINE vol:28 issue:2 pages:303-+ ispartof: location:United States status: published

Several approaches to manipulate the gut microbiome for improving the activity of cancer immune-checkpoint inhibitors (ICI) are currently under evaluation. Here, we show that oral supplementation with the polyphenol-rich berry camu-camu (CC; Myrciaria dubia) in mice shifted gut microbial composition, which translated into antitumor activity and a stronger anti–PD-1 response. We identified castalagin, an ellagitannin, as the active compound in CC. Oral administration of castalagin enriched for bacteria associated with efficient immunotherapeutic responses (Ruminococcaceae and Alistipes) and improved the CD8+/FOXP3+CD4+ ratio within the tumor microenvironment. Moreover, castalagin induced metabolic changes, resulting in an increase in taurine-conjugated bile acids. Oral supplementation of castalagin following fecal microbiota transplantation from ICI-refractory patients into mice supported anti–PD-1 activity. Finally, we found that castalagin binds to Ruminococcus bromii and promoted an anticancer response. Altogether, our results identify castalagin as a polyphenol that acts as a prebiotic to circumvent anti–PD-1 resistance. Significance: The polyphenol castalagin isolated from a berry has an antitumor effect through direct interactions with commensal bacteria, thus reprogramming the tumor microenvironment. In addition, in preclinical ICI-resistant models, castalagin reestablishes the efficacy of anti–PD-1. Together, these results provide a strong biological rationale to test castalagin as part of a clinical trial. This article is highlighted in the In This Issue feature, p. 873

ObjectiveGut microbial products are involved in regulation of host metabolism. In human and experimental studies, we explored the potential role of hippurate, a hepatic phase 2 conjugation product of microbial benzoate, as a marker and mediator of metabolic health.DesignIn 271 middle-aged non-diabetic Danish individuals, who were stratified on habitual dietary intake, we applied 1H-nuclear magnetic resonance (NMR) spectroscopy of urine samples and shotgun-sequencing-based metagenomics of the gut microbiome to explore links between the urine level of hippurate, measures of the gut microbiome, dietary fat and markers of metabolic health. In mechanistic experiments with chronic subcutaneous infusion of hippurate to high-fat-diet-fed obese mice, we tested for causality between hippurate and metabolic phenotypes.ResultsIn the human study, we showed that urine hippurate positively associates with microbial gene richness and functional modules for microbial benzoate biosynthetic pathways, one of which is less prevalent in the Bacteroides 2 enterotype compared with Ruminococcaceae or Prevotella enterotypes. Through dietary stratification, we identify a subset of study participants consuming a diet rich in saturated fat in which urine hippurate concentration, independently of gene richness, accounts for links with metabolic health. In the high-fat-fed mice experiments, we demonstrate causality through chronic infusion of hippurate (20 nmol/day) resulting in improved glucose tolerance and enhanced insulin secretion.ConclusionOur human and experimental studies show that a high urine hippurate concentration is a general marker of metabolic health, and in the context of obesity induced by high-fat diets, hippurate contributes to metabolic improvements, highlighting its potential as a mediator of metabolic health.

Florence Thirion,1,* Susie Guilly,1,* Sébastien Fromentin,1 Florian Plaza Oñate,1 Anne-Sophie Alvarez,1 Emmanuelle Le Chatelier,1 Nicolas Pons,1 Florence Levenez,1 Benoît Quinquis,1 Stanislav Ehrlich,1 Joel Doré,1,2 Richard Martin,3 Sophie Seité4 1University Paris-Saclay, INRAE, MGP, Jouy-en-Josas, 78350, France; 2University Paris-Saclay, INRAE, AgroParisTech, Micalis Institute, Jouy-en-Josas, France; 3Mercurialis Biotech, Rochecorbon, France; 4La Roche-Posay Dermatological Laboratories, Levallois-Perret, France*These authors contributed equally to this workCorrespondence: Sophie Seit&#x00E9La Roche-Posay Dermatological Laboratories, 62 quai Charles Pasqua, Levallois-Perret, 92300, France, Tel +33 1 49 64 33 40, Email sophie.seite@loreal.comIntroduction: Balneotherapy (BT) is the treatment of disease through the use of thermal spring water (TSW). It has been used for centuries and remains a popular form of treatment for dermatologic diseases such as atopic dermatitis (AD). Recent findings highlighted the role of the gut microbiota in AD and the possible crosstalk between gut and skin microbiomes in this pathology. Nevertheless, changes in the composition of the gut microbiota after balneotherapy remain to be elucidated.Methods: A total of 96 patients, with moderate to severe AD according to the SCORAD, were enrolled. Stool samples were collected prior and post a 3-week balneotherapy at the thermal care center of La Roche-Posay (France). Composition of the gut microbiota was assessed by shotgun metagenomic sequencing.Results: Species associated with high gut microbiota richness tended to correlate negatively with disease severity (SCORAD) and positively with SCORAD reduction, while species associated with low richness displayed the opposite pattern. Relative abundance of 23 species was significantly altered during BT, these changes being significantly associated with SCORAD reduction during BT, suggesting that gut microbiota composition and AD progression were associated through the treatment. Microbial functions related to gut-brain axis such as GABA and tryptophan metabolism were also altered by the treatment. Long-standing AD patients exhibited a better gut microbial profile than recently diagnosed patients, with higher MSP richness and species associated with SCORAD reduction.Conclusion: In patients with AD, clinical disease parameters such as SCORAD or disease duration are intricately linked to the gut microbiota composition. SCORAD reduction occurring during BT was also associated with gut microbiota. The gut-brain-skin axis via neurotransmitter such as GABA should be further studied in diseases such as AD.Keywords: atopic dermatitis, balneotherapy, gut microbiota, shotgun metagenomics, thermal spring water

Understanding the transmission of antibiotic resistance genes (ARGs) is critical for human health. For this, it is necessary to identify which type of mobile genetic elements is able to spread them from animal reservoirs into human pathogens. Previous research suggests that in pig feces, ARGs may be encoded by bacteriophages. However, convincing proof for phage-encoded ARGs in pig viromes is still lacking, because of bacterial DNA contaminating issues. We collected 14 pig fecal samples and performed deep sequencing on both highly purified viral fractions and total microbiota, in order to investigate phage and prophage-encoded ARGs. We show that ARGs are absent from the genomes of active, virion-forming phages (below 0.02% of viral contigs from viromes), but present in three prophages, representing 0.02% of the viral contigs identified in the microbial dataset. However, the corresponding phages were not detected in the viromes, and their genetic maps suggest they might be defective. Furthermore, our dataset allows for the first time a comprehensive view of the interplay between prophages and viral particles.

Background and Aims Cirrhosis is associated with changes in gut microbiome composition. Although acute-on-chronic liver failure (ACLF) is the most severe clinical stage of cirrhosis, there is lack of information about gut microbiome alterations in ACLF using quantitative metagenomics. We investigated the gut microbiome in patients with cirrhosis encompassing the whole spectrum of disease (compensated, acutely decompensated without ACLF, and ACLF). A group of healthy subjects was used as control subjects. Methods Stool samples were collected prospectively in 182 patients with cirrhosis. DNA library construction and sequencing were performed using the Ion Proton Sequencer (ThermoFisher Scientific, Waltham, MA). Microbial genes were grouped into clusters, denoted as metagenomic species. Results Cirrhosis was associated with a remarkable reduction in gene and metagenomic species richness compared with healthy subjects. This loss of richness correlated with disease stages and was particularly marked in patients with ACLF and persisted after adjustment for antibiotic therapy. ACLF was associated with a significant increase of Enterococcus and Peptostreptococcus sp and a reduction of some autochthonous bacteria. Gut microbiome alterations correlated with model for end-stage liver disease and Child-Pugh scores and organ failure and was associated with some complications, particularly hepatic encephalopathy and infections. Interestingly, gut microbiome predicted 3-month survival with good stable predictors. Functional analysis showed that patients with cirrhosis had enriched pathways related to ethanol production, γ-aminobutyric acid metabolism, and endotoxin biosynthesis, among others. Conclusions Cirrhosis is characterized by marked alterations in gut microbiome that parallel disease stages with maximal changes in ACLF. Altered gut microbiome was associated with complications of cirrhosis and survival. Gut microbiome may contribute to disease progression and poor prognosis. These results should be confirmed in future studies.

Aside from PD-L1 expression, biomarkers of response to immune checkpoint inhibitors (ICIs) in non-small-cell lung cancer (NSCLC) are needed. In a previous retrospective analysis, we documented that fecal Akkermansia muciniphila (Akk) was associated with clinical benefit of ICI in patients with NSCLC or kidney cancer. In the current study, we performed shotgun-metagenomics-based microbiome profiling in a large cohort of patients with advanced NSCLC (n = 338) treated with first- or second-line ICIs to prospectively validate the predictive value of fecal Akk. Baseline stool Akk was associated with increased objective response rates and overall survival in multivariate analyses, independent of PD-L1 expression, antibiotics, and performance status. Intestinal Akk was accompanied by a richer commensalism, including Eubacterium hallii and Bifidobacterium adolescentis, and a more inflamed tumor microenvironment in a subset of patients. However, antibiotic use (20% of cases) coincided with a relative dominance of Akk above 4.8% accompanied with the genus Clostridium, both associated with resistance to ICI. Our study shows significant differences in relative abundance of Akk that may represent potential biomarkers to refine patient stratification in future studies.

IntroductionIncreased adiposity, dyslipidemia and insulin resistance are associated with increased risk of developing cardiometabolic diseases (CM). Such deleterious phenotypes have been shown to be associated with a low gene-richness microbiota that can partly be restored by a short-term dietary intervention (energy-restricted high-protein diet, low glycemic index, enrichment with fibers) in parallel to an improvement of CM profile. In this study, we aimed at increasing fiber intake in quantity and diversity through a two-month consumption of bread enriched with a mix of selected fibers and evaluated the impact of this dietary intervention on gut microbiota gene richness and CM risk profile in subjects at risk of developing CM.Materials and methodsIn a randomized double blind cross-over design, thirty-nine subjects with CM risk profile (18–70 years old, BMI: 25–35 kg/m2, waist circumference > 80 cm for women and > 96 cm for men, fiber intake < 20g/day, low fiber diversity) consumed daily for 8 weeks 150 g of standard bread vs. 150 g of bread enriched with a 7-selected fibers mix (5.55 g vs. 16.35 g of fiber respectively; 4-week washout). Gut microbiota and CM risk factors’ analyzes were conducted before and after intervention. Stool samples were analyzed by shotgun metagenomics; microbial genes and metagenomics species (MSP) profiles were generated by mapping reads on a reference genes catalog (1529 MSP).ResultsThe included dyslipidemic subjects with CM risk profile presented a lower microbiota gene richness compared to reference healthy cohorts. The two-month consumption of fiber-rich bread did not alter microbiota gene richness but modified microbiota composition with a significant decrease of Bacteroides vulgatus (q = 1.7e-4) and a significant increase of Parabacteroides distasonis (q = 2.8e-6), Fusicatenibacter saccharivorans (q = 5e-5) and Clostridiales (q = 3.8e-2). We observed in parallel a significant decrease in total cholesterol (- 0.26 mmol/L; - 5%; p = 0.021), LDL-cholesterol (- 0.2 mmol/L; - 6%, p = 0.0061) and an improvement of insulin sensibility estimated by HOMA index (3.23–2.54 mUI/L; - 21%; p = 0.0079).These effects were even significantly more pronounced for subjects presenting the higher waist circumference. Anthropometric parameters were not altered.DiscussionThe enrichment of the diet with a mix of selected fibers for 2 months altered microbiota composition by modifying the relative abundance of specific gut bacterial species, in parallel to a significant improvement of cholesterol and insulin sensitivity parameters. Increasing the quantity and diversity of dietary fiber intake could be used as an efficient tool to favorably impact CM profile.

Objectifs Le microbiote intestinal humain est un écosystème complexe et dynamique. Il exerce de nombreuses fonctions biologiques et métaboliques. Le maintien de son équilibre (composition, activité) est important pour la santé et le bien-être de l’hôte et peut être influencé par de nombreux facteurs, comme l’alimentation. Les produits laitiers fermentés, qui contiennent une grande quantité de bactéries lactiques vivantes, peuvent contribuer à cet équilibre mais la compréhension de leur impact reste encore limitée. Ce travail a pour objectif de mieux comprendre, à partir de l’utilisation d’outils métaomiques, l’impact de la consommation de lait fermenté et de bactéries lactiques vivantes sur la composition et l’activité du microbiote intestinal. De plus, il vise à évaluer l’état physiologique et métabolique des bactéries lactiques en transit dans le tractus digestif. Matériels et Méthodes Trois groupes de 8 souris à flore humanisée (ie transplantation d’un microbiote fécal d’un individu sain à des souris axéniques) ont été alimentées avec (i) un régime contrôle sans lait (alimentation traditionnelle des souris + tampon PBS), (ii) un régime avec lait fermenté (alimentation traditionnelle des souris + un lait fermenté par les 2 bactéries du yaourt, Streptococcus thermophilus et Lactobacillus bulgaricus), ou (iii) un régime avec lait fermenté thermisé (alimentation traditionnelle des souris + un lait fermenté par les 2 bactéries du yaourt puis thermisé à 95°C pendant 25 min). La composition et l’activité du microbiote humain transplanté chez les souris ont été évaluées respectivement par métagénomique shotgun et métatranscriptomique à partir d’échantillons fécaux. Résultats principaux L’analyse quantitative et le suivi de l’activité du microbiote intestinal de souris à flore humanisée est réalisable grâce à des approches métaomiques. La consommation de lait fermenté (thermisé ou pas) ne modifie pas la composition du microbiote intestinal humain chez la souris mais a un impact significatif sur son activité. L’expression des gènes est modifiée pour la plupart des genres bactériens dominants, et modulant par conséquent leurs fonctions. Cet impact est d’autant plus important que la consommation de lait fermenté est régulière et prolongée dans le temps. En revanche, il n’a pas été possible de suivre l’activité des bactéries lactiques en transit qui sont minoritaires. Conclusion Ce travail met en évidence l’intérêt des méthodes métaomiques pour suivre l’impact de l’alimentation et ouvre des perspectives sur l’exploration des effets bénéfiques potentiels des laits fermentés sur le microbiote intestinal humain.

Motivation Analysis toolkits for shotgun metagenomic data achieve strain-level characterization of complex microbial communities by capturing intra-species gene content variation. Yet, these tools are hampered by the extent of reference genomes that are far from covering all microbial variability, as many species are still not sequenced or have only few strains available. Binning co-abundant genes obtained from de novo assembly is a powerful reference-free technique to discover and reconstitute gene repertoire of microbial species. While current methods accurately identify species core parts, they miss many accessory genes or split them into small gene groups that remain unassociated to core clusters. Results We introduce MSPminer, a computationally efficient software tool that reconstitutes Metagenomic Species Pan-genomes (MSPs) by binning co-abundant genes across metagenomic samples. MSPminer relies on a new robust measure of proportionality coupled with an empirical classifier to group and distinguish not only species core genes but accessory genes also. Applied to a large scale metagenomic dataset, MSPminer successfully delineates in a few hours the gene repertoires of 1661 microbial species with similar specificity and higher sensitivity than existing tools. The taxonomic annotation of MSPs reveals microorganisms hitherto unknown and brings coherence in the nomenclature of the species of the human gut microbiota. The provided MSPs can be readily used for taxonomic profiling and biomarkers discovery in human gut metagenomic samples. In addition, MSPminer can be applied on gene count tables from other ecosystems to perform similar analyses. Availability and implementation The binary is freely available for non-commercial users at www.enterome.com/downloads. Supplementary information Supplementary data are available at Bioinformatics online.

ObjectivesDecreased gut microbial gene richness (MGR) and compositional changes are associated with adverse metabolism in overweight or moderate obesity, but lack characterisation in severe obesity. Bariatric surgery (BS) improves metabolism and inflammation in severe obesity and is associated with gut microbiota modifications. Here, we characterised severe obesity-associated dysbiosis (ie, MGR, microbiota composition and functional characteristics) and assessed whether BS would rescue these changes.DesignSixty-one severely obese subjects, candidates for adjustable gastric banding (AGB, n=20) or Roux-en-Y-gastric bypass (RYGB, n=41), were enrolled. Twenty-four subjects were followed at 1, 3 and 12 months post-BS. Gut microbiota and serum metabolome were analysed using shotgun metagenomics and liquid chromatography mass spectrometry (LC-MS). Confirmation groups were included.ResultsLow gene richness (LGC) was present in 75% of patients and correlated with increased trunk-fat mass and comorbidities (type 2 diabetes, hypertension and severity). Seventy-eight metagenomic species were altered with LGC, among which 50% were associated with adverse body composition and metabolic phenotypes. Nine serum metabolites (includingglutarate,3-methoxyphenylacetic acidandL-histidine) and functional modules containing protein families involved in their metabolism were strongly associated with low MGR. BS increased MGR 1 year postsurgery, but most RYGB patients remained with low MGR 1 year post-BS, despite greater metabolic improvement than AGB patients.ConclusionsWe identified major gut microbiota alterations in severe obesity, which include decreased MGR and related functional pathways linked with metabolic deteriorations. The lack of full rescue post-BS calls for additional strategies to improve the gut microbiota ecosystem and microbiome–host interactions in severe obesity.Trial registration numberNCT01454232.

The gut bacterial species Akkermansia muciniphila is associated with a healthier clinical profile. The purpose of this study was to determine the association between A. muciniphila and glucose homeostasis in patients undergoing bariatric surgery (BS): gastric banding (GB) or Roux-en-Y gastric bypass (RYGB). This nonrandomized prospective study included 65 women with severe obesity. Longitudinal analysis included subjects for whom A. muciniphila data were available at follow-up [1, 3, and 12 mo; GB ( n = 10) or RYGB ( n = 11)]. Glucose homeostasis markers were measured under fasting conditions (glucose, insulin, and HbA1c) or during an oral glucose tolerance test. Fecal microbiota was analyzed using shotgun metagenomics, and A. muciniphila relative abundance was assessed with 16S rRNA quantitative PCR. A. muciniphila relative abundance was significantly lower in severe obesity [mean body mass index, 45.7 kg/m2 (SD 5.4)] than in moderate obesity [33.2 kg/m2 (SD 3.8)] but not associated with glucose homeostasis markers. A significant increase in A. muciniphila relative abundance after RYGB was not correlated with metabolic improvement. Baseline A. muciniphila abundance was correlated with bacterial gene richness and was highest in the high-richness Ruminococcaceae enterotype. A. muciniphila increased in relative abundance after BS in patients with low baseline A. muciniphila abundance, especially those with a Bacteroides type 2 enterotype classification. Although decreased in severe obesity, relative abundance of A. muciniphila was not associated with glucose homeostasis before or after BS. A certain level of A. muciniphila abundance might be required to observe a beneficial link to health. The severity of obesity and gut dysbiosis may partly explain the discrepancy with previous findings in less obese populations.

International audience; The Human Microbiome Project (HMP) identified the 16S rRNA gene sequences of 'most wanted' taxa-prevalent in the healthy human microbiota but distant from previously known sequences. Since 2012, few of the corresponding genomes have been isolated and sequenced, and only through advanced isolation techniques. We demonstrate that the genomes of the most wanted taxa can be identified computationally through their correlation in abundance across multiple public metagenomic data sets. We link over 200 most wanted sequences with nearly complete genome sequences, including half of the taxa identified as high-priority targets by the HMP. The genomes we identify have strong similarity to genomes reconstructed through expensive isolation techniques, and provide a more complete functional characterization of these organisms than can be extrapolated from their 16S rRNA gene. We also provide insights into the function of organisms for which 16S rRNA gene signatures were recently reported to be associated with health and host genetic factors.

MotivationAnalysis toolkits for shotgun metagenomic data achieve strain-level characterization of complex microbial communities by capturing intra-species gene content variation. Yet, these tools are hampered by the extent of reference genomes that are far from covering all microbial variability, as many species are still not sequenced or have only few strains available. Binning co-abundant genes obtained from de novo assembly is a powerful reference-free technique to discover and reconstitute gene repertoire of microbial species. While current methods accurately identify species core parts, they miss many accessory genes or split them into small gene groups that remain unassociated to core clusters.ResultsWe introduce MSPminer, a computationally efficient software tool that reconstitutes Metagenomic Species Pan-genomes (MSPs) by binning co-abundant genes across metagenomic samples. MSPminer relies on a new robust measure of proportionality coupled with an empirical classifier to group and distinguish not only species core genes but accessory genes also. Applied to a large scale metagenomic dataset, MSPminer successfully delineates in a few hours the gene repertoires of 1 661 microbial species with similar specificity and higher sensitivity than existing tools. The taxonomic annotation of MSPs reveals microorganisms hitherto unknown and brings coherence in the nomenclature of the species of the human gut microbiota. The provided MSPs can be readily used for taxonomic profiling and biomarkers discovery in human gut metagenomic samples. In addition, MSPminer can be applied on gene count tables from other ecosystems to perform similar analyses.AvailabilityThe binary is freely available for non-commercial users at enterome.fr/site/downloads/ Contact: florian.plaza-onate@inra.frSupplementary informationAvailable in the file named Supplementary Information.pdf

Liver cirrhosis occurs as a consequence of many chronic liver diseases that are prevalent worldwide. Here we characterize the gut microbiome in liver cirrhosis by comparing 98 patients and 83 healthy control individuals. We build a reference gene set for the cohort containing 2.69 million genes, 36.1% of which are novel. Quantitative metagenomics reveals 75,245 genes that differ in abundance between the patients and healthy individuals (false discovery rate

Nat. Biotechnol. 33, 1103–1108 (2015); published online 28 September 2015; corrected after print 18 December 2018 In the version of this article initially published, the y-axis numbering in Figure 1 was high by a factor of 10; the correct range is 0.5 to 2.5 million nonredundant genes. The error hasbeen corrected in the HTML and PDF versions of the article.

International audience; Background. During antibiotic treatments, antibiotics reach the colon and alter the microbiome, resulting in emergence and dissemination of resistant bacteria, antibiotic-associated diarrhea (AAD) and occurrence of Clostridium difficile (Cd) infections (CDI). We developed an oral adsorbent-based product, DAV132, to be co-administered with antibiotics in order to prevent these effects. We report here the results of protection of the microbiome in human volunteers treated with oral moxifloxacin (MOX), and protection against CDI in vitro in a human gut model (HGM) and in vivo in experimental hamsters.Methods. Four groups (MOX [400 mg OD for 5 days], MOX + DAV132 [7.5 g TID for 7 days], DAV132, or control) of healthy volunteers (n = 44) were included in a randomized clinical trial. Feces were collected at screening, daily from D1 to D9 and at D12, D16, D23, D30, D37 and plasma PK assayed at D1and5. Metagenomic analyses were done by shotgun sequencing at screening, D3, D6, D9, D16 and D37. The effect of DAV132 on MOX was also assessed in a HGM inoculated with Cd spores (ribotype 027, MOX MIC 32 mg/L); bacterial counts, toxin production and MOX concentrations were measured over time. MOX dosed hamsters were inoculated with 104 spores of a non-epidemic Cd (TcdA + , TcdB + , cdtB−) and treated ±DAV132. Mortality and Cd counts were assessed over time.Results. In humans, DAV132 was well tolerated and decreased free fecal MOX by > 99% compared to MOX alone: ratio of mean LogAUCD1-D16 = 0.0093 (95% CI 0.0061-0.0144), p = 0.03 × 10−16; MOX plasma PK was not changed significantly. Alterations of the fecal microbiome observed with MOX were prevented. In the HGM, DAV protected the microbiota and prevented Cd overgrowth and toxin production. Hamsters were fully protected by DAV against MOX-induced CDI (dose dependent), with decreased free fecal MOX and Cd counts.Conclusion. DAV132 protects humans against fecal microbiome disruption after oral MOX treatment, without altering its plasma PK. In vitro results in HGM and in vivo results in hamsters suggest that DAV132 should be protective against antibiotic-induced CDI. These results warrant further clinical development of DAV132 to protect the intestinal microbiota, and so prevent AAD and CDI, in patients receiving oral or parenteral antibiotics.

The gut microbiota has been implicated in obesity and its progression towards metabolic disease. Dietary interventions that target the gut microbiota have been suggested to improve metabolic health. The aim of the present study was to investigate the effect of interventions withLactobacillus paracaseiF19 or flaxseed mucilage on the gut microbiota and metabolic risk markers in obesity. A total of fifty-eight obese postmenopausal women were randomised to a single-blinded, parallel-group intervention of 6-week duration, with a daily intake of eitherL. paracaseiF19 (9·4 × 1010colony-forming units), flaxseed mucilage (10 g) or placebo. Quantitative metagenomic analysis of faecal DNA was performed to identify the changes in the gut microbiota. Diet-induced changes in metabolic markers were explored using adjusted linear regression models. The intake of flaxseed mucilage over 6 weeks led to a reduction in serum C-peptide and insulin release during an oral glucose tolerance test (PPPFaecalibacteriumspecies. These changes in the microbiota could not explain the effect of flaxseed mucilage on insulin sensitivity. The intake ofL. paracaseiF19 did not modulate metabolic markers compared with placebo. In conclusion, flaxseed mucilage improves insulin sensitivity and alters the gut microbiota; however, the improvement in insulin sensitivity was not mediated by the observed changes in relative abundance of bacterial species.