Janneke P. Ouwerkerk

Cross-talk between Akkermansia muciniphila and intestinal epithelium controls diet-induced obesity

Obesity and type 2 diabetes are characterized by altered gut microbiota, inflammation, and gut barrier disruption. Microbial composition and the mechanisms of interaction with the host that affect gut barrier function during obesity and type 2 diabetes have not been elucidated. We recently isolated Akkermansia muciniphila, which is a mucin-degrading bacterium that resides in the mucus layer. The presence of this bacterium inversely correlates with body weight in rodents and humans. However, the precise physiological roles played by this bacterium during obesity and metabolic disorders are unknown. This study demonstrated that the abundance of A. muciniphila decreased in obese and type 2 diabetic mice. We also observed that prebiotic feeding normalized A. muciniphila abundance, which correlated with an improved metabolic profile. In addition, we demonstrated that A. muciniphila treatment reversed high-fat diet-induced metabolic disorders, including fat-mass gain, metabolic endotoxemia, adipose tissue inflammation, and insulin resistance. A. muciniphila administration increased the intestinal levels of endocannabinoids that control inflammation, the gut barrier, and gut peptide secretion. Finally, we demonstrated that all these effects required viable A. muciniphila because treatment with heat-killed cells did not improve the metabolic profile or the mucus layer thickness. In summary, this study provides substantial insight into the intricate mechanisms of bacterial (i.e., A. muciniphila) regulation of the cross-talk between the host and gut microbiota. These results also provide a rationale for the development of a treatment that uses this human mucus colonizer for the prevention or treatment of obesity and its associated metabolic disorders.

Glycobiome: Bacteria and mucus at the epithelial interface

The human gastrointestinal tract is colonised with a dense and diverse microbial community, that is an important player in human health and physiology. Close to the epithelial cells the mucosal microbiota is separated from the host with a thin lining of host derived glycans, including the cell surface glycocalyx and the extracellular secreted mucus. The mucosa-associated microbial composition differs from the luminal content and could be particularly important for nutrient exchange, communication with the host, development of the immune system, and resistance against invading pathogens. The mucosa-associated microbiota has adapted to the glycan rich environment by the production of mucus-degrading enzymes and mucus-binding extracellular proteins, and include mucus-degrading specialists such as Akkermansia muciniphila and Bacteroides thetaiotaomicron. This review is focussed on the host-microbe interactions within the glycan landscape at the epithelial interface and considers the spatial organisation and composition of the mucosa-associated microbiota in health and disease.

Akkermansia muciniphila induces gut microbiota remodelling and controls islet autoimmunity in NOD mice

Objective Intestinal microbiota is implicated in the pathogenesis of autoimmune type 1 diabetes in humans and in non-obese diabetic (NOD) mice, but evidence on its causality and on the role of individual microbiota members is limited. We investigated if different diabetes incidence in two NOD colonies was due to microbiota differences and aimed to identify individual microbiota members with potential significance. Design We profiled intestinal microbiota between two NOD mouse colonies showing high or low diabetes incidence by 16S ribosomal RNA gene sequencing and colonised the high-incidence colony with the microbiota of the low-incidence colony. Based on unaltered incidence, we identified a few taxa which were not effectively transferred and thereafter, transferred experimentally one of these to test its potential significance. Results Although the high-incidence colony adopted most microbial taxa present in the low-incidence colony, diabetes incidence remained unaltered. Among the few taxa which were not transferred, Akkermansia muciniphila was identified. As A. muciniphila abundancy is inversely correlated to the risk of developing type 1 diabetes-related autoantibodies, we transferred A. muciniphila experimentally to the high-incidence colony. A. muciniphila transfer promoted mucus production and increased expression of antimicrobial peptide Reg3γ, outcompeted Ruminococcus torques from the microbiota, lowered serum endotoxin levels and islet toll-like receptor expression, promoted regulatory immunity and delayed diabetes development. Conclusion Transfer of the whole microbiota may not reduce diabetes incidence despite a major change in gut microbiota, but single symbionts such as A. muciniphila with beneficial metabolic and immune signalling effects may reduce diabetes incidence when administered as a probiotic.

Adaptation of Akkermansia muciniphila to the oxic-anoxic interface of the mucus layer

ABSTRACT Akkermansia muciniphila colonizes the mucus layer of the gastrointestinal tract, where the organism can be exposed to the oxygen that diffuses from epithelial cells. To understand how A. muciniphila is able to survive and grow at this oxic-anoxic interface, its oxygen tolerance and response and reduction capacities were studied. A. muciniphila was found to be oxygen tolerant. On top of this, under aerated conditions, A. muciniphila showed significant oxygen reduction capacities and its growth rate and yield were increased compared to those seen under strict anaerobic conditions. Transcriptome analysis revealed an initial oxygen stress response upon exposure to oxygen. Thereafter, genes related to respiration were expressed, including those coding for the cytochrome bd complex, which can function as a terminal oxidase. The functionality of A. muciniphila cytochrome bd genes was proven by successfully complementing cytochrome-deficient Escherichia coli strain ECOM4. We conclude that A. muciniphila can use oxygen when it is present at nanomolar concentrations. IMPORTANCE This article explains how Akkermansia muciniphila, previously described as a strictly anaerobic bacterium, is able to tolerate and even benefit from low levels of oxygen. Interestingly, we measured growth enhancement of A. muciniphila and changes in metabolism as a result of the oxygen exposure. In this article, we discuss similarities and differences of this oxygen-responsive mechanism with respect to those of other intestinal anaerobic isolates. Taken together, we think that these are valuable data that indicate how anaerobic intestinal colonizing bacteria can exploit low levels of oxygen present in the mucus layer and that our results have direct relevance for applicability, as addition of low oxygen concentrations could benefit the in vitro growth of certain anaerobic organisms.

Preparation and preservation of viable Akkermansia muciniphila cells for therapeutic interventions

The anaerobic gut bacterium Akkermansia muciniphila is a well-characterised member of the mucosal microbiota and has shown to be a gut symbiont in human. A. muciniphila has been negatively associated with obesity and its associated metabolic disorders in various human cohorts while treatment with A. muciniphila cells reversed highfat diet-induced obesity and its associated metabolic disorders in mouse models. Therefore, administration of A. muciniphila has been suggested as a possible new therapeutic treatment for these omnipresent diseases. Here we describe a potentially scalable workflow for the preparation and preservation of high numbers of viable cells of A. muciniphila obtained from 1 l laboratory scale growth under strict anaerobic conditions for therapeutic interventions. This resulted in viable A. muciniphila cells with high yields and very high stability, with up to 97.9±4.5% survival for a time period of 1 year at -80 °C in glycerol-amended medium. Moreover, various quality assessment and control procedures were developed to ensure the use of viable cells of A. muciniphila. Several microscopic, culturing, and molecular approaches were applied to monitor the presence, abundance and recovery of A. muciniphila before, during, and after its administration to high-fat treated mice. We show that viable A. muciniphila cells can be recovered from caecal and colon content (up to 1×1010 cells/g), testifying for the efficiency of the described workflow.

Complete genome sequence of Akkermansia glycaniphila strain Pyt T , a mucin-degrading specialist of the reticulated python gut

ABSTRACT Akkermansia glycaniphila is a novel Akkermansia species that was isolated from the intestine of the reticulated python and shares the capacity to degrade mucin with the human strain Akkermansia muciniphila MucT. Here, we report the complete genome sequence of strain PytT of 3,074,121 bp. The genomic analysis reveals genes for mucin degradation and aerobic respiration.

Akkermansia glycaniphila sp. nov., an anaerobic mucin-degrading bacterium isolated from reticulated python faeces

A Gram-stain-negative, non-motile, strictly anaerobic, oval-shaped, non-spore-forming bacterium (strain PytT) was isolated from reticulated python faeces. Strain PytT was capable of using mucin as sole carbon, energy and nitrogen source. Cells could grow singly, in pairs, and were also found to aggregate. Scanning electron microscopy revealed the presence of filamentous structures connecting individual bacterial cells. Strain PytT could grow on a limited number of single sugars, including N-acetylglucosamine, N-acetylgalactosamine, glucose, lactose and galactose, but only when a plentiful protein source was provided. Phylogenetic analysis based on 16S rRNA gene sequencing showed strain PytT to belong to the Verrucomicrobiae class I, family Akkermansiaceae, genus Akkermansia, with Akkermansia muciniphila MucT as the closest relative (94.4 % sequence similarity). DNA-DNA hybridization revealed low relatedness of 28.3 % with A. muciniphila MucT. The G+C content of DNA from strain PytT was 58.2 mol%. The average nucleotide identity (ANI) of the genome of strain PytT compared to the genome of strain MucT was 79.7 %. Chemotaxonomic data supported the affiliation of strain PytT to the genus Akkermansia. Based on phenotypic, phylogenetic and genetic characteristics, strain PytT represents a novel species of the genus Akkermansia, for which the name Akkermansia glycaniphila sp. nov. is proposed. The type strain is PytT (=DSM 100705T=CIP 110913T).

O80 Akkermansia muciniphila communique avec l’épithélium intestinal pour contrôler le développement de l’obésité et du diabète de type 2

Introduction : Le microbiote intestinal est implique dans le developpement de l’obesite, de l’insulino-resistance et du diabete de type 2. Nos travaux precedents ont mis en evidence qu’une bacterie decouverte recemment et vivant dans le mucus intestinal, Akkermansia muciniphila, etait diminuee (100 a 1000 fois) dans des modeles nutritionnel ou genetique d’obesite et de diabete de type 2. Par contre sa presence dans le contenu caecal est normalisee lors d’un traitement a l’aide de prebiotiques. Cette modulation du microbiote intestinal est associee a une diminution de l’inflammation de bas grade et a une amelioration de la sensibilite a l’insuline et a la leptine. Cependant l’impact direct d’Akkermansia muciniphila sur le metabolisme n’a encore jamais ete investigue. Materiel et methode : Akkermansia muciniphila a ete administree quotidiennement par gavage oral chez des souris nourries avec un regime hyperlipidique pendant 4 semaines. Des marqueurs de l’homeostasie glucidique, du metabolisme lipidique et de la fonction barriere de l’intestin ont ete investigues. Resultats : Le traitement avec Akkermansia muciniphila ameliore les marqueurs d’insulino-sensibilite et corrige l’hyperglycemie a jeun induite par le regime hyperlipidique. Ces effets benefiques sont notamment associes a une diminution de la neoglucogenese hepatique (diminution de l’expression de l’ARNm de la glucose-6-phosphatase). L’administration d’Akkermansia muciniphila empeche la prise de masse grasse et le developpement de l’inflammation du tissu adipeux sous regime hyperlipidique, sans affecter la quantite d’energie ingeree. Akkermansia muciniphila restaure la fonction barriere de l’intestin (disparition de l’endotoxemie metabolique, restauration de la couche de mucus, modification de la production de peptides antimicrobiens). Conclusions : Ces resultats mettent en evidence de nouveaux mecanismes de regulation du metabolisme par le microbiote intestinal (Akkermansia.muciniphila) et permettent egalement de mettre en avant l’interet du developpement d’un traitement utilisant Akkermansia muciniphila pour la prevention ou le traitement de l’obesite et du diabete de type 2.