Introduction

Between 50% and 70% of patients with Crohn’s disease (CD) will require an ileocecal resection. Surgery is not curative, and endoscopic recurrence is frequently observed at the anastomosis site and/or in the neo-terminal ileum.
The objective of this study was to analyse the mucosa-associated ileal microbiota at the time of surgery and during the colonoscopy performed 6 months later and to identify predictive factors of post-operative endoscopic recurrence.

Aims & Methods

Patients were recruited from 15 centres within the REMIND group. Since September 2010, the REMIND group has established a homogeneous, prospective, multicenter cohort (POP-REMIND, NCT03458195) of patients undergoing surgery for ileal and/or ileocolonic Crohn’s disease. Samples were collected from the surgical specimen and during endoscopy, centrally stored in a biobank, and analysed to identify factors associated with recurrence.
Inclusion criteria were: age ≥18 years, ileal or ileocecal CD requiring intestinal resection. Data were collected using an electronic case report form (eCRF). Post-operative treatment was prescribed according to a predefined algorithm. Clinical, therapeutic, biological, and endoscopic data (Rutgeerts classification) were collected 6 months after surgery.
The composition of the mucosa-associated ileal microbiota was analysed at the time of surgery in 140 patients (M0) and in 125 samples at the colonoscopy performed 6 months later (M6), using ultra-deep 16S and ITS2 sequencing (MiSeq, Illumina) to characterize bacterial and fungal microbiota.
We specifically selected patients who had not received antibiotics during the perioperative period to avoid biasing the analysed signal. Sequencing data were processed using the DADA2 pipeline. Alpha and beta diversity and microbiota composition were analysed. Bacterial taxa associated with clinical parameters were identified using MaAsLin2 (Multivariate Association with Linear Models), accounting for CD phenotype and treatments.

Results

We observed numerous differences in the ileal bacterial and fungal microbiota between M0 and M6, and between patients who experienced recurrence (>i2a) and those who were in remission at M6. At M6, the Rutgeerts score was the main factor associated with variability in microbiota composition. The abundance of certain species, notably a decrease of Faecalibacterium prausnitzii, or increase of Fusobacterium nucleatum and Akkermansia muciniphila, was differentially associated with a whole range of levels of the Rutgeerts score or with more specific scores characterizing the inflammation of the anastomosis or the ileum. In addition, machine learning approaches displayed better performances when considering the Rutgeerts score as a multilevel prediction instead of a binary classification, further confirming the need to consider its full range. Finally, we investigated the microbial community dynamics, which highlighted a denser network organisation after surgery and in the absence of recurrence, along with changes in keystone species.

Conclusion

The ileal microbiota associated with post-operative recurrence in CD follows a dynamic process characterized by several waves of alterations. The link between ileal microbiota and disease recurrence varies depending on whether the lesions are located at the anastomosis or ileum sites, supporting a different underlying pathogenesis for the inflammatory process at these two locations.