The gut microbiota and inflammatory bowel disease:From exploration to clinical translation

Inflammatory bowel disease (IBD), including Crohn’s disease and ulcerative colitis, are chronic diseases of the gastrointestinal tract in which patients experience periods of inflammation alternated by periods of remission. In this thesis the role of the gut microbiota in IBD has been investigated. The gut microbiota – all micro-organisms such as bacteria, viruses and funghi which reside in the gastrointestinal tract – play an important role in our health. These micro-organisms have important functions in our body, including the digestion of food and by interacting with our immune system. In the first part of this thesis bacteria and metabolic functions were identified associated with IBD by using the sequencing technique “metagenomic sequencing”. The location of the inflammation has been identified to be associated with gut microbial changes and the gut microbiota can be used to distinguish between patients with IBD and patients with irritable bowel syndrome, a condition in which patients experience gastrointestinal complaints without the presence of inflammation. In the second part of the thesis the influence of medication on the gut microbiota composition has been investigated. In here, antibiotics, laxatives, proton-pump inhibitors and metformin have been identified to largely influence the gut microbiota. Additionally, the potential of the gut microbiota in predicting treatment response of the biological vedolizumab in patients with IBD have been shown. These findings contribute to unravel the role of the gut microbiota in IBD and also to translate these findings towards clinical practice by using the gut microbiota in the diagnostics, monitoring and treatment of IBD

Gut microbiota in inflammatory bowel diseases:moving from basic science to clinical applications

In recent years, large efforts have been made to unravel the role of the gut microbiota in inflammatory bowel disease (IBD), which is a chronic inflammatory disorder of the gastro-intestinal tract. Considering the heterogeneity patients with IBD display in their disease course and response to treatment, there is a big need in translating these findings towards clinical practise. In this perspective article, we discuss strategies to facilitate the transition from basic science on gut microbiota in IBD to clinical applications. We suggest that setting gold standards, improving and increasing the biobanking efforts, and studying other members of the gut microbiota are a necessary step to reveal the exact role of the gut microbiota in IBD. In addition, we discuss the potential of the gut microbiome as a clinical tool for the diagnoses, prediction and/or treatment of the disease. We believe that the growing interest in the gut microbiota will reveal its potential in the management of IBD in a not too distant future

Anti-inflammatory Gut Microbial Pathways Are Decreased During Crohn's Disease Exacerbations

BACKGROUND AND AIMS: Crohn's disease [CD] is a chronic inflammatory disorder of the gastrointestinal tract characterised by alternating periods of exacerbation and remission. We hypothesised that changes in the gut microbiome are associated with CD exacerbations, and therefore aimed to correlate multiple gut microbiome features to CD disease activity. METHODS: Faecal microbiome data generated using whole-genome metagenomic shotgun sequencing of 196 CD patients were of obtained from the 1000IBD cohort [one sample per patient]. Patient disease activity status at time of sampling was determined by re-assessing clinical records 3 years after faecal sample production. Faecal samples were designated as taken 'in an exacerbation' or 'in remission'. Samples taken 'in remission' were further categorised as 'before the next exacerbation' or 'after the last exacerbation', based on the exacerbation closest in time to the faecal production date. CD activity was correlated with gut microbial composition and predicted functional pathways via logistic regressions using MaAsLin software. RESULTS: In total, 105 bacterial pathways were decreased during CD exacerbation (false-discovery rate [FDR] <0.1) in comparison with the gut microbiome of patients both before and after an exacerbation. Most of these decreased pathways exert anti-inflammatory properties facilitating the biosynthesis and fermentation of various amino acids [tryptophan, methionine, and arginine], vitamins [riboflavin and thiamine], and short-chain fatty acids [SCFAs]. CONCLUSIONS: CD exacerbations are associated with a decrease in microbial genes involved in the biosynthesis of the anti-inflammatory mediators riboflavin, thiamine, and folate, and SCFAs, suggesting that increasing the intestinal abundances of these mediators might provide new treatment opportunities. These results were generated using bioinformatic analyses of cross-sectional data and need to be replicated using time-series and wet lab experiments

Long-term dietary patterns are associated with pro-inflammatory and anti-inflammatory features of the gut microbiome

Objective The microbiome directly affects the balance of pro-inflammatory and anti-inflammatory responses in the gut. As microbes thrive on dietary substrates, the question arises whether we can nourish an anti-inflammatory gut ecosystem. We aim to unravel interactions between diet, gut microbiota and their functional ability to induce intestinal inflammation. Design We investigated the relation between 173 dietary factors and the microbiome of 1425 individuals spanning four cohorts: Crohn's disease, ulcerative colitis, irritable bowel syndrome and the general population. Shotgun metagenomic sequencing was performed to profile gut microbial composition and function. Dietary intake was assessed through food frequency questionnaires. We performed unsupervised clustering to identify dietary patterns and microbial clusters. Associations between diet and microbial features were explored per cohort, followed by a meta-analysis and heterogeneity estimation. Results We identified 38 associations between dietary patterns and microbial clusters. Moreover, 61 individual foods and nutrients were associated with 61 species and 249 metabolic pathways in the meta-analysis across healthy individuals and patients with IBS, Crohn's disease and UC (false discovery rate Conclusion We identified dietary patterns that consistently correlate with groups of bacteria with shared functional roles in both, health and disease. Moreover, specific foods and nutrients were associated with species known to infer mucosal protection and anti-inflammatory effects. We propose microbial mechanisms through which the diet affects inflammatory responses in the gut as a rationale for future intervention studies

The composition and metabolic potential of the human small intestinal microbiota within the context of inflammatory bowel disease

BACKGROUND AND AIMS: The human gastrointestinal tract harbours distinct microbial communities essential for health. Little is known about small intestinal communities, despite the small intestine playing a fundamental role in nutrient absorption and host-microbe immune homeostasis. We aimed to explore the small intestine microbial composition and metabolic potential, in the context of inflammatory bowel disease (IBD). METHODS: Metagenomes derived from faecal samples and extensive phenotypes were collected from 57 individuals with an ileostomy or ileoanal pouch, and compared with 1178 general population and 478 IBD faecal metagenomes. Microbiome features were identified using MetaPhAn2 and HUMAnN2, and association analyses were performed using multivariate linear regression. RESULTS: Small intestinal samples had a significantly lower bacterial diversity, compared with the general population and, to a lesser extent, IBD samples. Comparing bacterial composition, small intestinal samples clustered furthest from general population samples and closest to IBD samples with intestinal resections. Veillonella atypica, Streptococcus salivarius and Actinomyces graevenitzii were among the species significantly enriched in the small intestine. Predicted metabolic pathways in the small intestine are predominantly involved in simple carbohydrate and energy metabolism, but also suggest a higher proinflammatory potential. CONCLUSION: We described the bacterial composition and metabolic potential of the small intestinal microbiota. The colonic microbiome of IBD patients, particularly with intestinal resections, showed resemblance to that of the small intestine. Moreover, several features characterising the small intestinal microbiome have been previously associated with IBD. These results highlight the importance of studying the small intestinal microbiota to gain new insight into disease pathogenesis

Impact of commonly used drugs on the composition and metabolic function of the gut microbiota

The human gut microbiota has now been associated with drug responses and efficacy, while chemical compounds present in these drugs can also impact the gut bacteria. However, drug–microbe interactions are still understudied in the clinical context, where polypharmacy and comorbidities co-occur. Here, we report relations between commonly used drugs and the gut microbiome. We performed metagenomics sequencing of faecal samples from a population cohort and two gastrointestinal disease cohorts. Differences between users and non-users were analysed per cohort, followed by a meta-analysis. While 19 of 41 drugs are found to be associated with microbial features, when controlling for the use of multiple medications, proton-pump inhibitors, metformin, antibiotics and laxatives show the strongest associations with the microbiome. We here provide evidence for extensive changes in taxonomy, metabolic potential and resistome in relation to commonly used drugs. This paves the way for future studies and has implications for current microbiome studies by demonstrating the need to correct for multiple drug use

Patient attitudes towards faecal sampling for gut microbiome studies and clinical care reveal positive engagement and room for improvement

Faecal sample collection is crucial for gut microbiome research and its clinical applications. However, while patients and healthy volunteers are routinely asked to provide stool samples, their attitudes towards sampling remain largely unknown. Here, we investigate the attitudes of 780 Dutch patients, including participants in a large Inflammatory Bowel Disease (IBD) gut microbiome cohort and population controls, in order to identify barriers to sample collection and provide recommendations for gut microbiome researchers and clinicians. We sent questionnaires to 660 IBD patients and 112 patients with other disorders who had previously been approached to participate in gut microbiome studies. We also conducted 478 brief interviews with participants in our general population cohort who had collected stool samples. Statistical analysis of the data was performed using R. 97.4% of respondents reported that they had willingly participated in stool sample collection for gut microbiome research, and most respondents (82.9%) and interviewees (95.6%) indicated willingness to participate again, with their motivations for participating being mainly altruistic (57.0%). Responses indicated that storing stool samples in the home freezer for a prolonged time was the main barrier to participation (52.6%), but clear explanations of the sampling procedures and their purpose increased participant willingness to collect and freeze samples (P = 0.046, P = 0.003). To account for participant concerns, gut microbiome researchers establishing cohorts and clinicians trying new faecal tests should provide clear instructions, explain the rationale behind their protocol, consider providing a small freezer and inform patients about study outcomes. By assessing the attitudes, motives and barriers surrounding participation in faecal sample collection, we provide important information that will contribute to the success of gut microbiome research and its near-future clinical applications

Faecal metabolome and its determinants in inflammatory bowel disease

OBJECTIVE: Inflammatory bowel disease (IBD) is a multifactorial immune-mediated inflammatory disease of the intestine, comprising Crohn's disease and ulcerative colitis. By characterising metabolites in faeces, combined with faecal metagenomics, host genetics and clinical characteristics, we aimed to unravel metabolic alterations in IBD.DESIGN: We measured 1684 different faecal metabolites and 8 short-chain and branched-chain fatty acids in stool samples of 424 patients with IBD and 255 non-IBD controls. Regression analyses were used to compare concentrations of metabolites between cases and controls and determine the relationship between metabolites and each participant's lifestyle, clinical characteristics and gut microbiota composition. Moreover, genome-wide association analysis was conducted on faecal metabolite levels.RESULTS: We identified over 300 molecules that were differentially abundant in the faeces of patients with IBD. The ratio between a sphingolipid and L-urobilin could discriminate between IBD and non-IBD samples (AUC=0.85). We found changes in the bile acid pool in patients with dysbiotic microbial communities and a strong association between faecal metabolome and gut microbiota. For example, the abundance of Ruminococcus gnavus was positively associated with tryptamine levels. In addition, we found 158 associations between metabolites and dietary patterns, and polymorphisms near NAT2 strongly associated with coffee metabolism.CONCLUSION: In this large-scale analysis, we identified alterations in the metabolome of patients with IBD that are independent of commonly overlooked confounders such as diet and surgical history. Considering the influence of the microbiome on faecal metabolites, our results pave the way for future interventions targeting intestinal inflammation.</p

Gut microbial co-abundance networks show specificity in inflammatory bowel disease and obesity

The gut microbiome is an ecosystem that involves complex interactions. Currently, our knowledge about the role of the gut microbiome in health and disease relies mainly on differential microbial abundance, and little is known about the role of microbial interactions in the context of human disease. Here, we construct and compare microbial co-abundance networks using 2,379 metagenomes from four human cohorts: an inflammatory bowel disease (IBD) cohort, an obese cohort and two population-based cohorts. We find that the strengths of 38.6% of species co-abundances and 64.3% of pathway co-abundances vary significantly between cohorts, with 113 species and 1,050 pathway co-abundances showing IBD-specific effects and 281 pathway co-abundances showing obesity-specific effects. We can also replicate these IBD microbial co-abundances in longitudinal data from the IBD cohort of the integrative human microbiome (iHMP-IBD) project. Our study identifies several key species and pathways in IBD and obesity and provides evidence that altered microbial abundances in disease can influence their co-abundance relationship, which expands our current knowledge regarding microbial dysbiosis in disease

SLC39A8 missense variant is associated with Crohn's disease but does not have a major impact on gut microbiome composition in healthy subjects

Background Gene-microbiome interactions are important in aetiology and pathogenesis of inflammatory bowel disease, a chronic inflammatory disorder of the gastrointestinal tract consisting of Crohn's disease and ulcerative colitis. Scarce studies on gene-microbiome interactions show very little overlap in their results. Therefore, it is of utmost importance that gene-microbiome studies are repeated. We aimed to replicate the association between the SLC39A8[Thr]391 risk allele and gut microbiome composition in patients with inflammatory bowel disease and healthy controls. Methods We collected faecal samples, peripheral blood and extensive phenotype data from 291 patients with inflammatory bowel disease and 476 healthy controls. Carrier status information was obtained from whole exome sequencing data, generated using the Illumina HiSeq. The gut microbiome composition was determined by tag-sequencing the 16S rRNA gene. Associations between carrier status and disease were tested using the Wilcoxon-Mann-Whitney test. Associations between carriers and gut microbiome composition were determined using principal coordinate analyses, variance explained, alpha diversity and additive general linear models in inflammatory bowel disease, healthy controls and all groups combined. Results Crohn's disease patients were more often carriers of the missense variant (21/171, 12.3%) than controls (30/476, 6.3%) (OR = 2.1, P = 0.01). We could not identify associations between carrier status and overall gut microbiome composition and microbial richness in all tested groups after correcting for potential confounding factors. We did identify 37 different operational taxonomical units to be associated with carrier status among the tested groups. Two of these 37 were identified before in the discovery study. Conclusions We could confirm the genetic association of the SLC39A8[Thr]391 risk allele with Crohn's disease but we could only limited replicate the association in gut microbiome composition. Independent replication of gene-microbiome studies is warranted to identify true biological mechanisms