DIFFERENTIAL ABUNDANCE OF THE FUNGAL MICROBIOME IN PATIENTS WITH ULCERATIVE COLITIS

Background
The fungal microbiome has been increasingly implicated in the pathogenesis of ulcerative colitis (UC). Circulating antibodies to Saccharomyces antibodies are detected in 10-15% of patients with UC, and patients with high fecal Candida are more likely to demonstrate a favorable response to fecal microbial transplantation (FMT) in inducing remission. However, the fungal microbiome remains poorly characterized in UC, especially across the spectrum of endo-histologic activity and following exposure to immunosuppressive drugs. Furthermore, unidentified fungal sequences are commonly recovered from UC cohort studies.

Aims
In this study, we aim to characterize the fungal microbiome in UC patients with varying levels of endoscopic activity, endo-histologic activity, and treatment exposure as well as additionally identify unknown fungal sequences.

Methods
We performed a secondary analysis using the data extracted from the Crohn’s and Colitis Foundation’s Study of a Prospective Adult Research Cohort with IBD, which contains clinical, endoscopic, histologic, and metagenomic data. Using Internal Transcribed Spacer based deep sequencing of fungal rDNA from fecal samples, we classified sequences utilizing the UNITE fungal database and a fitted classifier. These sequences were also analyzed with the Basic Local Alignment Search Tool (BLAST). Phyloseq and DESeq2 in R studio were used to assess fungal diversity and differential abundance of fungi between comparator groups.

Results
We identified 500 unique fungal amplicon sequence variants across the cohort of 82 patients, belonging to phylum Ascomycota (71.5%), Basidiomycota (11%), Mucoromycota (0.16%), or unidentified (17.2%). We found no differences in alpha or beta diversity in the fungal microbiome during endoscopic activity vs remission. However, patients with endoscopic activity (n=25) had relative increases in Saccharomyces (log 2-fold change 4.54, p-adj< 5 x 10-5) and Candida (2.56, p-adj< 0.03), compared to during endoscopic remission. Similarly, patients with endo-histologic activity (n=19) were also enriched for Saccharomyces (2.78, p-adj<0.08), compared to endo-histologic remission. Exposure to immunosuppressants was not associated with any significant changes in differential abundance about phylum or genera. After adjusting for age, gender, and biologic exposure among patients with endoscopic activity, Saccharomyces (7.76, p-adj < 1 x 10-15) and Candida (7.28, p-adj < 1 x 10-8) remained enriched. Among 270 unclassifiable fungal ASVs, we identified the majority of these to be low-prevalence ASVs (in <5% of samples), with 265 representing dietary contaminants (i.e. seed plants) and 5 representing either a fungus (Candida mesenterica), bacteria (Phocaeicola vulgatus, Eubacterium rectale, or Bacteroides ovatus), or a nematode (Meloidogyne chitwoodi).

Conclusion
These data demonstrate that endoscopic and histologic inflammation in UC is associated with relative expansion of Saccharomyces and Candida compared to remission, even after controlling for exposure to biologic exposure. Unidentified fungal sequences are usually of low prevalence and represent dietary contaminants. The role of fungal ASVs as potential biomarkers and targets for personalized approaches to therapeutics in UC should be evaluated in future studies.