Composition of the human small intestine microbiome in the context of inflammatory bowel disease

Overview

The human gastrointestinal tract harbors a unique microbiota that is crucial for health. Although the small intestine plays an important role in nutrient absorption and host-microbe immune homeostasis, little is known about the small intestinal communities. We aim to explore the composition of small intestinal microbiota in the context of inflammatory bowel disease (IBD).

The human gut microbiome refers to the trillions of bacteria, viruses, fungi, and archaea residing in the gastrointestinal (GI) tract, playing a significant role in maintaining health. Changes in the composition of the gut microbiome have been linked to various diseases, ranging from gastrointestinal disorders, including inflammatory bowel disease (IBD), to neurological diseases. However, the use of fecal samples in most studies means that the majority of the content primarily targets the colon; that is, the fecal microbiome does not capture all the microbial communities present in other parts of the gastrointestinal tract (e.g., the small intestine), which remain relatively under-studied.

The small intestine is responsible for approximately 90% of the total nutrients absorbed by the body from the diet and plays a central role in maintaining host-microbe immune homeostasis. Dysbiosis of the duodenal microbiota has been associated with certain gastrointestinal-related diseases, such as functional dyspepsia, bloating, and diarrhea. Additionally, ectopic colonization of typical oral microbes is believed to play a role in the pathogenesis of several diseases: this phenomenon is referred to as 'oralization.' For example, specific strains of Klebsiella pneumoniae isolated from the salivary microbiome of IBD patients can cause abnormal activation of the immune system after colonizing the colons of mice susceptible to colitis.

Studying small intestinal contents, particularly in healthy environments, is challenging due to its poor accessibility. To date, most studies have relied on mucosal samples collected during routine endoscopies, after bowel resections, or from deceased patients. However, this sampling method is prone to contamination and may be hindered by lavage procedures performed prior to some of these procedures. Furthermore, they do not represent the microbial content within the lumen of the small intestine and are limited by the lower taxonomic and functional resolution of 16s rRNA sequencing. This article aims to describe the composition of the small intestinal microbiota, with a particular focus on its potential impact on IBD.

1.1   The small intestinal microbiota is characterized by lower microbial richness and different bacterial composition.

On average, samples from the IBD-SI (patients with IBD who underwent ileostomy) group exhibited lower microbial richness [mean Shannon index IBD-SI = 1.71; mean Shannon index IBD-Res (patients with resected intestines) = 2.44, p = 5.10× 10^-14; mean Shannon index IBD-NoRes (patients with IBD who did not undergo resection) = 2.77, p = 2.22× 10^-16; mean Shannon index general population = 2.84, p = 2.22× 10^-16] [Figure 1a]. To gain a rough understanding of the bacterial composition between groups, we measured beta diversity using Bray-Curtis dissimilarity [Figures 1b and c]. Samples from the IBD-SI group clustered the farthest from the general population samples. IBD-Res and IBD-NoRes samples formed a gradient between IBD-SI and general population samples, with IBD-Res samples closer to SI samples. After correcting for potential confounding factors, the differences in microbial richness and overall bacterial composition between groups remained significant FDR < 0.05.

Compared to colonic samples, bacterial diversity in the small intestine is lower and composition is different. [a] represents the violin plot of Shannon index values distribution for each study group. Small intestinal samples on average exhibit lower bacterial diversity compared to general population samples, IBD non-resected samples, and IBD resected samples. The box plot shows the median and interquartile range [25th and 75th]. The scatter plot shows the Bray-Curtis differences between samples as a measure of overall bacterial composition differences. Samples are colored according to the group classifications used throughout the study (gray, general population samples; purple, samples from IBD patients without bowel resection; yellow, samples from IBD patients with bowel resection; red, representing small intestinal samples). The percentages on the x-axis and y-axis indicate the total variance explained by each coordinate. Triangles represent the average coordinate values for each group. Figure [b] highlights the differences among all samples used in this study. Small intestinal samples form a distinct cluster with almost no overlap with general population samples. Samples from IBD patients [purple and yellow] form a gradient between the small intestine and general population. Figure [c] highlights the heterogeneity among IBD samples.

1.2   Overall genus composition in the small intestine.

To characterize the differences observed in the β diversity analysis, we compared the top 12 most abundant genera in general population and IBD-SI samples. The most abundant genera in the IBD-SI group were Streptococcus, Escherichia, Blautia, noname genus of the family Ruminococcaceae, Clostridium, Lactobacillus, and Veillonella [mean relative abundances = 26%, 10%, 8.1%, 6.7%, 5.2%, and 5.5%, 4.8%]. Except for Blautia, all abundances were significantly higher compared to other groups [Streptococcus: IBD-SI vs. general population, FDR = 2.73 × 10^–23; IBD-SI vs. IBD-NoRes, FDR = 6.39 × 10^–17; IBD-SI vs. IBD-Res, FDR = 3.73 × 10^–14. Notably, the total average abundance of genera in the IBD-Res group was the second highest, while the general population had the lowest. The opposite trend was also observed for the most abundant genera in the general population, which included Bifidobacterium, Ruminococcus, Faecalibacterium, Subdoligranulum, and Faecalibacterium [mean relative abundances = 15%, 15%, 14%, 5.8%, 5.6%, and 5.2%; the order of total relative abundance increase was: IBD-SI, IBD-Res, IBD-NoRes, and general population.

Compared to samples representing the colon, the bacterial genus profiles in samples representing the small intestine microbiome are significantly different. [A,B] The bar charts represent the relative abundance of the top 10 most abundant genera in each study group for the general population samples and small intestine samples, respectively. Bifidobacterium and Lactobacillus are the most abundant bacterial genera in the general population samples and IBD samples from patients who have not undergone intestinal resection. In contrast, Streptococcus and Escherichia are the most abundant in samples representing small intestinal contents. [C,D] The box plots show the average relative abundance of the top 12 abundant bacteria in each test group for the general population samples [C] and small intestine samples [D]. Comparisons marked as 'ns' are not significant. A two-sided Wilcoxon test was used to assess significance. The box plots display the median and interquartile range [25th and 75th].

Conclusion

We found that samples belonging to the small intestine group had significantly lower bacterial diversity compared to other groups. Small intestine samples were also markedly different from samples representing the general population in terms of overall bacterial composition, as indicated by Bray-Curtis differences. These findings highlight the selective pressures that can drive bacterial selection between the small intestine and colon. For example, the small intestine is considered a harsh environment for microbial survival due to its acidic environment, higher oxygen concentration, shorter transit time, and regular influx of digestive enzymes and bile.

In the small intestine, compared to the fecal microbiome of the general population, the bacterial species that are relatively abundant include Veillonella atypica, Streptococcus pneumoniae, Streptococcus salivarius, Bifidobacterium dentium, Haemophilus parainfluenzae, and Actinomyces graevenitzii. Additionally, species belonging to genera such as Clostridium, Lactobacillus, Klebsiella, Gemella, and Enterococcus, which are rarely observed in fecal samples from the general population, have significantly higher prevalence in small intestine samples, ranging between 15% and 75%.

Overall, the experiments demonstrate that the colonic contents of a subset of IBD patients resemble the small intestine microbiome, suggesting the translocation of small intestinal pathogens to the colon. However, further experimental evidence is needed to support this notion and to elucidate the mechanisms of small intestine microbiome and its interactions with the host.