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. 2024 Jun 18;25(12):6697.
doi: 10.3390/ijms25126697.

Integrated Analysis of Microbiome and Metabolome Reveals Disease-Specific Profiles in Inflammatory Bowel Diseases and Intestinal Behçet's Disease

Yehyun Park  1   2 , Jae Bum Ahn  1 , Da Hye Kim  1 , I Seul Park  1   3 , Mijeong Son  1   3 , Ji Hyung Kim  1   3 , Hyun Woo Ma  1   3 , Seung Won Kim  1   3 , Jae Hee Cheon  1   3
Affiliations

Integrated Analysis of Microbiome and Metabolome Reveals Disease-Specific Profiles in Inflammatory Bowel Diseases and Intestinal Behçet's Disease

Yehyun Park et al. Int J Mol Sci. .

Abstract

The gut microbial and metabolic characteristics of intestinal Behçet's disease (BD), a condition sharing many clinical similarities with ulcerative colitis (UC) and Crohn's disease (CD), are largely unexplored. This study investigated the gut microbial and metabolic characteristics of intestinal BD as well as potential biomarkers, comparing them with those in UC, CD, and healthy controls. Colon tissue and stool samples from 100 patients (35 UC, 30 CD, and 35 intestinal BD) and 41 healthy volunteers were analyzed using 16S ribosomal RNA sequencing to assess microbial diversity, taxonomic composition, and functional profiling. Plasma metabolomic analyses were performed using gas chromatography and ultra-performance liquid chromatography-mass spectrometry. Results indicated reduced microbial diversity in CD but not in intestinal BD, with intestinal BD showing fewer changes compared to controls yet distinct taxonomic features from UC, CD, and controls. Common alterations across all diseases included a reduction in beneficial bacteria producing short-chain fatty acids. Intestinal BD-specific changes featured a decreased abundance of Bacteroides fragilis. Metabolomic profiles in intestinal BD were similar to those in CD but distinct from those in UC, displaying significant changes in energy metabolism and genetic information processing. This integrative analysis revealed both shared and unique profiles in intestinal BD compared with UC, CD, and controls, advancing our understanding of the distinctive features of these diseases.

Keywords: Crohn’s disease; intestinal Behçet’s disease; metabolome; microbiome; multi-omics; ulcerative colitis.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Flow diagram of the study. In total, 192 samples, comprising tissue, stool, and blood, were collected from 141 subjects (35 UC, 30 CD, 35 intestinal BD, and 41 healthy volunteers) and analyzed. IBD: inflammatory bowel disease, BD: Behçet’s disease, rRNA: ribosomal ribonucleic acid, GC-TOF-MS: gas chromatography time-of-flight mass spectrometry, UPLC-Q-TOF-MS: ultra-performance liquid chromatography-quadrupole/time-of-flight mass spectrometry.
Figure 2
Figure 2
Venn diagram of samples collected from 141 patients or controls according to sample type. UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 3
Figure 3
Stacked bar chart of the microbial composition in colon tissue. The microbial analyses of fecal samples from the control and IBD participants showed different patterns. At the phylum level, IBD showed decreased Firmicutes and Actinobacteria and increased Proteobacteria and Bacteroidetes compared with the control (Supplementary Figure S1). The asterisk (*) indicates microbiota that show significant differences when compared to the control.
Figure 4
Figure 4
Microbial α-diversity index from tissue and fecal samples. Microbial richness and evenness were evaluated by the Shannon index. Each small shape represents an individual patient. In the tissue samples, CD had a significant decrease in α-diversity compared with the control, UC, and intestinal BD. HC: healthy control, CD: Crohn’s disease, UC: ulcerative colitis, BD: Behçet’s disease, ns: not significant.
Figure 5
Figure 5
Principal coordinate analysis (PCoA) plots of tissue samples (A) and stool samples (B). Beta-diversity was analyzed by the Bray-Curtis method. (A) PCoA plot for tissue samples shows clustering according to groups and significantly different microbial compositions between the control and CD, the UC and CD, the UC and intestinal BD, and the CD and intestinal BD samples (all p < 0.05). (B) PCoA plot for fecal samples shows greater separation between IBD and the control and significantly different microbial composition between the control and UC and the control and CD samples (all p < 0.05). HC: healthy control, UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 5
Figure 5
Principal coordinate analysis (PCoA) plots of tissue samples (A) and stool samples (B). Beta-diversity was analyzed by the Bray-Curtis method. (A) PCoA plot for tissue samples shows clustering according to groups and significantly different microbial compositions between the control and CD, the UC and CD, the UC and intestinal BD, and the CD and intestinal BD samples (all p < 0.05). (B) PCoA plot for fecal samples shows greater separation between IBD and the control and significantly different microbial composition between the control and UC and the control and CD samples (all p < 0.05). HC: healthy control, UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 6
Figure 6
Taxonomic biomarkers analyzed by LEfSe of tissue samples (A,B) and fecal samples (C). Taxa with an LDA effect size > 3 and p < 0.05 were visualized. (A) LEfSe analysis of tissue samples demonstrated significantly different abundances of specific taxa among the control, UC, CD, and intestinal BD samples. (B) LEfSe analysis performed separately for intestinal BD vs. the control showed a decreased abundance of butyrate-producing bacteria in intestinal BD. (C) LEfSe analysis of fecal samples also demonstrated significantly different abundances of specific taxa among the control, UC, and CD samples. LDA: linear discriminant analysis, LEfSe: LDA effect size, UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 7
Figure 7
Microbial taxonomic biomarkers of IBD and intestinal BD. The downward arrow indicates a decrease in the respective microbiota, while the upward arrow indicates an increase. The genus Fusicatenibacter and species Fusicatenibacter saccharivorans, Coprococcus comes, Blautia obeum, Dorea formicigenerans, and Roseburia cecicola consistently exhibited decreased abundance, indicating their ‘protective’ role in UC, CD, and intestinal BD. Intestinal BD displayed fewer significant changes, mainly characterized by a decrease in the abundance of several taxa. IBD: inflammatory bowel disease, UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 8
Figure 8
Predictive functional profiling based on PICRUSt analysis of tissue samples. (A) KEGG orthology. (B) KEGG module. The PICRUSt analysis revealed multiple enhanced gene allocations for each group. Intestinal BD exhibited pronounced enhancements in functions related to drug resistance, signaling and cellular processes, and metabolic pathways. PICRUSt: phylogenetic investigation of communities by reconstruction of unobserved states, KEGG: Kyoto Encyclopedia of Genes and Genomes, UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 9
Figure 9
Partial least squares discriminant analysis (PLS-DA) of plasma metabolites by GC-TOF-MS analysis. (A) Using five components, we can identify combinations of two components that best explain the differences between groups. Components 1 and 2 have high explanatory power and effectively illustrate the differences between groups. (B) PLS score 2D plot generated using components 1 and 2. The plot demonstrates the segregation of three groups: control, UC, and a combined group of CD and intestinal BD. (C) PLS score 3D plot using components 1, 2, and 3. GC-TOF-MS: gas chromatography time-of-flight mass spectrometry, UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 10
Figure 10
Heatmap of whole plasma metabolite profiles in individual samples (A) and by group (B). Overall, the metabolite profiles showed similarity between the control and UC and between CD and intestinal BD. UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 10
Figure 10
Heatmap of whole plasma metabolite profiles in individual samples (A) and by group (B). Overall, the metabolite profiles showed similarity between the control and UC and between CD and intestinal BD. UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 11
Figure 11
Plasma partial least squares discriminant analysis (PLS-DA) variable importance in a projection (VIP) plot. The key metabolites that contributed most to the separation among controls, UC, CD, and intestinal BD samples are shown in a PLS-DA VIP plot ranked by importance. UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
Figure 12
Figure 12
Potential metabolomic biomarkers of IBD and intestinal BD. In total, 26 metabolites with a VIP > 1.0, ANOVA p < 0.05, and FDR-adjusted p < 0.1 (20 different from control, 6 different among UC, CD, and intestinal BD) were noted. IBD: inflammatory bowel disease, UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease, VIP: variable importance in projection, ANOVA: analysis of variance, FDR: false discovery rate. The downward arrow indicates a decrease in the respective microbiota, while the upward arrow indicates an increase.
Figure 13
Figure 13
Quantitative enrichment analysis (QEA) in IBD and intestinal BD (A), UC (B), CD (C), and intestinal BD (D) compared with the control and mapped to KEGG pathways (FDR-adjusted p < 0.1). IBD: inflammatory bowel disease, UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease, KEGG: Kyoto Encyclopedia of Genes and Genomes, FDR: false discovery rate.
Figure 14
Figure 14
Correlation heatmap of potential microbial and metabolomic biomarkers in UC, CD, and intestinal BD. The Spearman rank correlation analysis between potential microbial and metabolomic biomarkers is shown in the correlation heatmap. The microbial taxa that mainly decreased in UC correlate positively with most of the metabolomic biomarkers, whereas the microbial taxa that mainly decreased in CD and intestinal BD correlate negatively with most of the metabolomic biomarkers. The cells marked with asterisks (*) indicate p < 0.05 in the Spearman correlation analysis. UC: ulcerative colitis, CD: Crohn’s disease, BD: Behçet’s disease.
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