Dirk Gevers

Minimum information about a marker gene sequence (MIMARKS) and minimum information about any (x) sequence (MIxS) specifications

Here we present a standard developed by the Genomic Standards Consortium (GSC) for reporting marker gene sequences—the minimum information about a marker gene sequence (MIMARKS). We also introduce a system for describing the environment from which a biological sample originates. The environmental packages apply to any genome sequence of known origin and can be used in combination with MIMARKS and other GSC checklists. Finally, to establish a unified standard for describing sequence data and to provide a single point of entry for the scientific community to access and learn about GSC checklists, we present the minimum information about any (x) sequence (MIxS). Adoption of MIxS will enhance our ability to analyze natural genetic diversity documented by massive DNA sequencing efforts from myriad ecosystems in our ever-changing biosphere.

Host genetic variation impacts microbiome composition across human body sites

BackgroundThe composition of bacteria in and on the human body varies widely across human individuals, and has been associated with multiple health conditions. While microbial communities are influenced by environmental factors, some degree of genetic influence of the host on the microbiome is also expected. This study is part of an expanding effort to comprehensively profile the interactions between human genetic variation and the composition of this microbial ecosystem on a genome- and microbiome-wide scale.ResultsHere, we jointly analyze the composition of the human microbiome and host genetic variation. By mining the shotgun metagenomic data from the Human Microbiome Project for host DNA reads, we gathered information on host genetic variation for 93 individuals for whom bacterial abundance data are also available. Using this dataset, we identify significant associations between host genetic variation and microbiome composition in 10 of the 15 body sites tested. These associations are driven by host genetic variation in immunity-related pathways, and are especially enriched in host genes that have been previously associated with microbiome-related complex diseases, such as inflammatory bowel disease and obesity-related disorders. Lastly, we show that host genomic regions associated with the microbiome have high levels of genetic differentiation among human populations, possibly indicating host genomic adaptation to environment-specific microbiomes.ConclusionsOur results highlight the role of host genetic variation in shaping the composition of the human microbiome, and provide a starting point toward understanding the complex interaction between human genetics and the microbiome in the context of human evolution and disease.

Sub-clinical detection of gut microbial biomarkers of obesity and type 2 diabetes

BackgroundObesity and type 2 diabetes (T2D) are linked both with host genetics and with environmental factors, including dysbioses of the gut microbiota. However, it is unclear whether these microbial changes precede disease onset. Twin cohorts present a unique genetically-controlled opportunity to study the relationships between lifestyle factors and the microbiome. In particular, we hypothesized that family-independent changes in microbial composition and metabolic function during the sub-clinical state of T2D could be either causal or early biomarkers of progression.MethodsWe collected fecal samples and clinical metadata from 20 monozygotic Korean twins at up to two time points, resulting in 36 stool shotgun metagenomes. While the participants were neither obese nor diabetic, they spanned the entire range of healthy to near-clinical values and thus enabled the study of microbial associations during sub-clinical disease while accounting for genetic background.ResultsWe found changes both in composition and in function of the sub-clinical gut microbiome, including a decrease in Akkermansia muciniphila suggesting a role prior to the onset of disease, and functional changes reflecting a response to oxidative stress comparable to that previously observed in chronic T2D and inflammatory bowel diseases. Finally, our unique study design allowed us to examine the strain similarity between twins, and we found that twins demonstrate strain-level differences in composition despite species-level similarities.ConclusionsThese changes in the microbiome might be used for the early diagnosis of an inflamed gut and T2D prior to clinical onset of the disease and will help to advance toward microbial interventions.

Increased Intestinal Microbial Diversity Following Fecal Microbiota Transplant for Active Crohn's Disease

Background:The microbiota in the lumen of patients with Crohns disease (CD) is characterized by reduced diversity, particularly Firmicutes and Bacteroidetes. It is unknown whether the introduction of the intestinal microbiota from healthy individuals could correct this dysbiosis and reverse mucosal inflammation. We investigated the response to fecal microbial transplantation (FMT) from healthy individuals to subjects with active CD. Methods:We performed a prospective open-label study (uncontrolled) of FMT from healthy donors to subjects with active CD. A single FMT was performed by colonoscopy. Recipients microbial diversity, mucosal T-cell phenotypes, and clinical and inflammatory parameters were measured over 12 weeks, and safety over 26 weeks. Results:Nineteen subjects were treated with FMT and completed the study follow-up. Fifty-eight percent (11/19) demonstrated a clinical response (Harvey–Bradshaw Index decrease >3) following FMT. Fifteen subjects had sufficient pre/postfecal samples for analysis. A significant increase in microbial diversity occurred after FMT (P = 0.02). This was greater in clinical responders than nonresponders. Patients who experienced a clinical response demonstrated a significant shift in fecal microbial composition toward their donors profile as assessed by the Bray–Curtis index at 4 weeks (P = 0.003). An increase in regulatory T cells (CD4+CD25+CD127lo) was also noted in recipients lamina propria following FMT. No serious adverse events were noted over the 26-week study period. Conclusions:In this open-label study, FMT led to an expansion in microbial bacterial diversity in patients with active CD. FMT was overall safe, although the clinical response was variable. Determining donor microbial factors that influence clinical response is needed before randomized clinical trials of FMT in CD.

MetaRef: a pan-genomic database for comparative and community microbial genomics

Microbial genome sequencing is one of the longest-standing areas of biological database development, but high-throughput, low-cost technologies have increased its throughput to an unprecedented number of new genomes per year. Several thousand microbial genomes are now available, necessitating new approaches to organizing information on gene function, phylogeny and microbial taxonomy to facilitate downstream biological interpretation. MetaRef, available at http://metaref.org, is a novel online resource systematically cataloguing a comprehensive pan-genome of all microbial clades with sequenced isolates. It organizes currently available draft and finished bacterial and archaeal genomes into quality-controlled clades, reports all core and pan gene families at multiple levels in the resulting taxonomy, and it annotates families’ conservation, phylogeny and consensus functional information. MetaRef also provides a comprehensive non-redundant reference gene catalogue for metagenomic studies, including the abundance and prevalence of all gene families in the >700 shotgun metagenomic samples of the Human Microbiome Project. This constitutes a systematic mapping of clade-specific microbial functions within the healthy human microbiome across multiple body sites and can be used as reference for identifying potential functional biomarkers in disease-associate microbiomes. MetaRef provides all information both as an online browsable resource and as downloadable sequences and tabular data files that can be used for subsequent offline studies.

Advancing the Microbiome Research Community

The human microbiome has become a recognized factor in promoting and maintaining health. We outline opportunities in interdisciplinary research, analytical rigor, standardization, and policy development for this relatively new and rapidly developing field. Advances in these aspects of the research community may in turn advance our understanding of human microbiome biology.

A Microbiome Foundation for the Study of Crohn's Disease

Our 2014 study published in Cell Host & Microbe, The Treatment-Naive Microbiome in New-Onset Crohns Disease, was designed to improve our understanding of the microbiomes role in Crohns disease by studying a unique, well-suited cohort and sample set. This commentary provides a hindsight perspective of this original study as well as future outlook.

A novel Ruminococcus gnavus clade enriched in inflammatory bowel disease patients

BackgroundInflammatory bowel disease (IBD) is characterized by chronic inflammation of the gastrointestinal tract that is associated with changes in the gut microbiome. Here, we sought to identify strain-specific functional correlates with IBD outcomes.MethodsWe performed metagenomic sequencing of monthly stool samples from 20 IBD patients and 12 controls (266 total samples). These were taxonomically profiled with MetaPhlAn2 and functionally profiled using HUMAnN2. Differentially abundant species were identified using MaAsLin and strain-specific pangenome haplotypes were analyzed using PanPhlAn.ResultsWe found a significantly higher abundance in patients of facultative anaerobes that can tolerate the increased oxidative stress of the IBD gut. We also detected dramatic, yet transient, blooms of Ruminococcus gnavus in IBD patients, often co-occurring with increased disease activity. We identified two distinct clades of R. gnavus strains, one of which is enriched in IBD patients. To study functional differences between these two clades, we augmented the R. gnavus pangenome by sequencing nine isolates from IBD patients. We identified 199 IBD-specific, strain-specific genes involved in oxidative stress responses, adhesion, iron-acquisition, and mucus utilization, potentially conferring an adaptive advantage for this R. gnavus clade in the IBD gut.ConclusionsThis study adds further evidence to the hypothesis that increased oxidative stress may be a major factor shaping the dysbiosis of the microbiome observed in IBD and suggests that R. gnavus may be an important member of the altered gut community in IBD.

The human gut microbiome in early-onset type 1 diabetes from the TEDDY study

Type 1 diabetes (T1D) is an autoimmune disease that targets pancreatic islet beta cells and incorporates genetic and environmental factors1, including complex genetic elements2, patient exposures3 and the gut microbiome4. Viral infections5 and broader gut dysbioses6 have been identified as potential causes or contributing factors; however, human studies have not yet identified microbial compositional or functional triggers that are predictive of islet autoimmunity or T1D. Here we analyse 10,913 metagenomes in stool samples from 783 mostly white, non-Hispanic children. The samples were collected monthly from three months of age until the clinical end point (islet autoimmunity or T1D) in the The Environmental Determinants of Diabetes in the Young (TEDDY) study, to characterize the natural history of the early gut microbiome in connection to islet autoimmunity, T1D diagnosis, and other common early life events such as antibiotic treatments and probiotics. The microbiomes of control children contained more genes that were related to fermentation and the biosynthesis of short-chain fatty acids, but these were not consistently associated with particular taxa across geographically diverse clinical centres, suggesting that microbial factors associated with T1D are taxonomically diffuse but functionally more coherent. When we investigated the broader establishment and development of the infant microbiome, both taxonomic and functional profiles were dynamic and highly individualized, and dominated in the first year of life by one of three largely exclusive Bifidobacterium species (B. bifidum, B. breve or B. longum) or by the phylum Proteobacteria. In particular, the strain-specific carriage of genes for the utilization of human milk oligosaccharide within a subset of B. longum was present specifically in breast-fed infants. These analyses of TEDDY gut metagenomes provide, to our knowledge, the largest and most detailed longitudinal functional profile of the developing gut microbiome in relation to islet autoimmunity, T1D and other early childhood events. Together with existing evidence from human cohorts7,8 and a T1D mouse model9, these data support the protective effects of short-chain fatty acids in early-onset human T1D.An analysis of more than 10,000 metagenomes from the TEDDY study provides a detailed functional profile of the gut microbiome in relation to islet autoimmunity, and supports the protective effects of short-chain fatty acids in early-onset type 1 diabetes.

Microbial community function and biomarker discovery in the human microbiome

Microbial communities carry out the majority of the biochemical activity on the planet, and they play integral roles in processes such as metabolism and immune homeostasis in the human microbiome. Whole genome shotgun sequencing of such communities’ metagenomes is becoming an increasingly feasible complement to obtaining organismal information from taxonomic markers. However, the resultant dataset typically comprises short reads from hundreds of different organisms, making it challenging to assemble and functionally annotate these sequences in the standard manner for single-organism genomes. n nWe describe an alternative to this approach to infer the functional and metabolic potential of a microbial community metagenome by determining whether gene families and pathways are present or absent, as well as their relative abundances, directly from short sequence reads. We validated this methodology using synthetic metagenomes, recovering the presence and abundance of large pathways and of small functional modules with high accuracy. We subsequently applied this approach to the microbial communities of 649 metagenomes drawn from 7 primary body sites on 102 individuals as part of the Human Microbiome Project (HMP), demonstrating the scalability of our methodology and the critical importance of microbial metabolism in the human microbiota. This provided a framework in which to define functional diversity in comparison to organismal ecology, including an example of microbial metabolism linked to specific organisms and to host phenotype (vaginal pH) in the posterior fornix. We provide profiles of 168 functional modules and 196 metabolic pathways that were determined to be specific to one or more niches within the human microbiome, including details of glycosaminoglycan degradation in the gut. n nUnderstanding how and why these biomolecular activities differ among environmental conditions or disease phenotypes is, more broadly, one of the central questions addressed by high-throughput biology. We have thus developed the linear discriminant analysis (LDA) effect size algorithm (LEfSe) to discover and explain microbial and functional biomarkers in the human microbiota and other microbiomes. We demonstrate this method to be effective for mining human microbiomes for metagenomic biomarkers associated with mucosal tissues and with different levels of oxygen availability. Similarly, when applied to 16S rRNA gene data from a murine ulcerative colitis gut community, LEfSe confirms the key role played by Bifidobacterium in this disease and suggests the involvement of additional clades, including the Clostridia and Metascardovia. A quantitative validation of LEfSe highlights a lower false positive rate, consistent ranking of biomarker relevance, and concise representations of taxonomic and functional shifts in microbial communities associated with environmental conditions or disease phenotypes. n nImplementations of both methodologies are available at the Huttenhower laboratory’s website [1,2]. Together, they provide a way to accurately and efficiently characterize microbial metabolic pathways and functional modules directly from high-throughput sequencing reads and, subsequently, to identify organisms, genes or pathways that consistently explain the differences between two or more microbial communities. This has allowed the determination of community roles in the HMP cohort, as well as their niche and population specificity, which we anticipate will be applicable to future metagenomic studies.