Discovery of a widespread polyamine-low molecular weight thiol hybrid pathway in Clostridioides difficile

Date of Award

Spring 1-1-2025

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Microbiology

First Advisor

Crawford, Jason

Abstract

Clostridioides difficile infection can cause severe inflammation in the gastrointestinal (GI) tract, leading to diarrhea, colitis, and an increased risk of colorectal cancer. Colonization of C. difficile is associated with community-level changes in the expression of polyamine biosynthesis genes. Polyamines are abundant cationic metabolites that serve indispensable functions for all kingdoms, particularly in gut homeostasis. Catabolism of the polyamine precursors arginine and ornithine offers C. difficile supplemental nutrition while subverting host immunity, yet existing models of C. difficile metabolism are incomplete regarding polyamines with comparable importance in the gut (e.g., spermidine). In this study, we conducted feeding studies with isotope labelled polyamines and discovered a network of low molecular weight thiol (LMWT) molecules termed clostridithiols (CSHs) constructed from polyamines conjugated with N-acetylcysteine (NAC) moieties. Through the analysis of a human microbiota diversity collection, we established that these previously uncharacterized hybrid metabolites are widely detected in Firmicutes and Bacteroidetes. A gain-of-function screen in an alternative CSH producer Bacteroides uniformis enabled the identification and validation of a two gene operon that was conserved in both producing organisms. CSH abundance in GI mucosal biopsies positively correlated with colorectal cancer versus matched healthy control samples. These studies indicate that the human microbiota broadly unite polyamine and LMWT functionalities that may be associated with disease.

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