Characteristics of mucosa-associated gut microbiota during treatment in Crohn's disease

Abstract

Background: The dysbiosis of the gut microbiome is evident in Crohn's disease (CD) compared with healthy controls (HC), although the alterations from active CD to remission after treatment are unclear.

Aim: To characterize the mucosa-associated gut microbiota in patients with CD before and after the induction therapy.

Methods: The basic information was collected from the subjects and the CD activity index (CDAI) was calculated in patients. A 16S rRNA sequencing approach was applied to determine the structures of microbial communities in mucosal samples including the terminal ileal, ascending colon, descending colon and rectum. The composition and function of mucosa-associated gut microbiota were compared between samples from the same cohort of patients before and after treatment. Differential taxa were identified to calculate the microbial dysbiosis index (MDI) and the correlation between MDI and CDAI was analyzed using Pearson correlation test. Predictive functional profiling of microbial communities was obtained with PICRUSt.

Results: There were no significant differences in microbial richness among the four anatomical sites in individuals. Compared to active disease, the alpha diversity of CD in remission was increased towards the level of HC compared to the active stage. The principal coordinate analysis revealed that samples of active CD were clearly separated from those in remission, which clustered close to HC. Sixty-five genera were identified as differentially abundant between active and quiescent CD, with a loss of Fusobacterium and a gain of potential beneficial bacteria including Lactobacillus, Akkermansia, Roseburia, Ruminococcus and Lachnospira after the induction of remission. The combination of these taxa into a MDI showed a positive correlation with clinical disease severity and a negative correlation with species richness. The increased capacity for the inferred pathways including Lipopolysaccharide biosynthesis and Lipopolysaccharide biosynthesis proteins in patients before treatment negatively correlated with the abundance of Roseburia, Ruminococcus and Lachnospira.

Conclusion: The dysbiosis of mucosa-associated microbiota was associated with the disease phenotype and may become a potential diagnostic tool for the recurrence of disease.

Keywords: 16S rRNA sequence; Active; Crohn’s disease; Mucosa-associated gut microbiota; Remission.

Figure 1

Richness and diversity in the mucosa-associated microbiota of the patients with Crohn’s disease and healthy controls before and after induction of remission. A: The number of observed species was 388.26 ± 94.22 in the Before group, 475.03 ± 96.01 in the After group and 547.85 ± 52.85 in the healthy controls (HC) group. The Chao1 index was 617.78 ± 161.04 in the Before group, 771.40 ± 146.62 in the After group and 864.08 ± 91.59 in the HC group; B: Plots shown were generated using the weighted version of the UniFrac-based Principal coordinate analysis. Samples from After group (green triangle) clustered separately from Before group (blue circle) while got close to the HC group (red square). HC: Healthy controls.

Figure 2

The microbial dysbiosis index characterizes the activity of Crohn’s disease. A: The composition of mucosa-associated microbiota at phylum level; B: The heatmap of 65 differentially abundant genera between patients before and after treatment. Each column represented a mucosal sample from patients before and after treatment as well as healthy controls. The first letter means the site of the colon (L: Ascending colon; R: Descending colon; i: Terminal ileum; Z: Rectum); C: The identified 65 genera were used to calculate the microbial dysbiosis index (MDI). The Pearson correlation analysis was constructed between MDI and the Crohn’s disease activity index as well as MDI and Chao index. MDI: Microbial dysbiosis index; HC: Healthy controls; CDAI: Crohn’s disease activity index.

Figure 3

Spearman correlations among the 65 Crohn’s disease-associated genera in gut mucosal samples. The green circle represented taxa enriched in patients after treatment while the red circle represented taxa enriched in patients before treatment. Lines between nodes denoted Pearson correlation (r > 0.2 and P < 0.05). The red and blue lines represented the positive and negative correlation, respectively.

Figure 4

The predicted functional module involving pro-inflammatory pathways altered in Crohn’s disease compared to healthy controls. A: Pathways including Lipopolysaccharide biosynthesis proteins and Lipopolysaccharide biosynthesis predicted to show significant different abundances among before, after and healthy controls group according to Kyoto Encyclopedia of Genes and Genome pathway analysis. The Crohn’s disease-depleted genera including Roseburia, Ruminococcus and Lachnospira were negatively correlated with Lipopolysaccharide biosynthesis proteins (P = 0.001 for Roseburia, P = 0.002 for Ruminococcus and P = 0.025 for Lachnospira) and Lipopolysaccharide biosynthesis (P = 0.0002 for Roseburia, P = 0.002 for Ruminococcus and P = 0.021 for Lachnospira). B: Roseburia; C: Ruminococcus; D: Lachnospira.