Intestinal microbial dysbiosis, intestinal inflammation, and Th17 immunity are all linked to the pathophysiology of spondyloarthritis (SpA); however, the mechanisms linking them remain unknown. One potential hypothesis suggests that the dysbiotic gut microbiome as a whole produces metabolites that influence human immune cells. To identify potential disease-relevant, microbiome-produced metabolites, we performed metabolomics screening and shotgun metagenomics on paired colon biopsies and fecal samples, respectively, from subjects with axial SpA (axSpA, N=21), Crohn’s disease (CD, N=27), and Crohn’s-axSpA overlap (CD-axSpA, N=12), as well as controls (HC, N=24). Using LC-MS based metabolomics of 4 non-inflamed pinch biopsies of the distal colon from subjects, we identified significant alterations in tryptophan pathway metabolites, including an expansion of indole-3-acetate (IAA) in axSpA and CD-axSpA compared to HC and CD and indole-3-acetaldehyde (I3Ald) in axSpA and CD-axSpA but not CD compared to HC, suggesting possible specificity to the development of axSpA. We then performed shotgun metagenomics of fecal samples to characterize gut microbial dysbiosis across these disease states. In spite of no significant differences in alpha-diversity among the 4 groups, our results confirmed differences in gene abundances of numerous enzymes involved in tryptophan metabolism. Specifically, gene abundance of indolepyruvate decarboxylase, which generates IAA and I3Ald, was significantly elevated in individuals with axSpA while gene abundances in HC demonstrated a propensity towards tryptophan synthesis. Such genetic changes were not observed in CD, again suggesting disease specificity for axSpA. Given the emerging role of tryptophan and its metabolites in immune function, altogether these data indicate that tryptophan metabolism into I3Ald and then IAA is one mechanism by which the gut microbiome potentially influences the development of axSpA.

In order to generate meaningful metabolomic and microbiome data, the method of sample collection is critical. This study utilizes and compares two methods for intestinal tissue collection for evaluation of metabolites and microbiomes, finding that using a brush to sample the microbiome provides valuable data. However, for metabolomics assessment, biopsy samples may still be required.

Newer ‘omics approaches such as metatranscriptomics and metabolomics allow functional assessments of the interaction(s) between the gut microbiome and the human host. In order to generate meaningful data with these approaches, though, the method of sample collection is critical. Prior studies have relied upon expensive and invasive means towards sample acquisition such as intestinal biopsy, while other studies have relied upon easier methods of collection such as fecal samples that do not necessarily represent those microbes in contact with the host. In this pilot study, we attempt to characterize a novel, minimally invasive method towards sampling the human microbiome using mucosal cytology brush sampling compared to intestinal gut biopsy on 5 healthy participants undergoing routine screening colonoscopy. We compared metatranscriptomic analyses between the two collection methods, identifying increased taxonomic evenness and beta diversity in the cytology brush samples, and similar community transcriptional profiles between the two methods. Metabolomics assessment demonstrated striking differences between the two methods, implying a difference in bacterial-derived versus human absorbed metabolites. Put together, this study supports the use of a less invasive method of microbiome sampling with cytology brushes, but caution must be exercised when performing metabolomics assessment as this represents differential metabolite production but not absorption by the host.ImportanceIn order to generate meaningful metabolomic and microbiome data, the method of sample collection is critical. This study utilizes and compares two methods to intestinal tissue collection for evaluation of metabolites and microbiome, finding that using a brush to sample the microbiome is superior to tissue biopsy. However, for metabolomics assessment, biopsy may still be required.