Keywords

Clostridioides difficile, xanthoquinodin, antimicrobials, toxin, iron homeostasis, transcriptomics

Abstract

The bacterial pathogen Clostridioides difficile is the leading cause of antibiotic- and healthcare-associated diarrhea, imposing significant clinical and economic impacts. The emergence of hypervirulent strains, such as PCR ribotype 027 (RT027), has exacerbated disease severity, recurrence rates, antibiotic resistance, and epidemiological spread. Expanding resistance profiles among C. difficile strains continue to limit the effectiveness of current IDSA-SHEA recommended therapies, which are already compromised by high recurrence rates due to antibiotic-induced disruption of the gut microbiota. These challenges highlight the urgent clinical need to identify and characterize antimicrobial compounds targeting alternative pathways in C. difficile physiology. We report the discovery and characterization of five bioactive xanthoquinodin analogues purified from Trichocladium sp., demonstrating potent antimicrobial activity against epidemic RT027 isolates and the susceptibility-testing reference strain ATCC 700057. Beyond direct antimicrobial effects, we demonstrated that xanthoquinodins modulate toxin A production at subinhibitory concentrations, potentially attenuating a key virulence factor of C. difficile. To elucidate the bactericidal mechanism of xanthoquinodin, we performed global transcriptomic profiling upon acute exposure. RNA-seq analyses revealed widespread dysregulation of iron homeostasis, characterized by significant upregulation of iron-acquisition genes and metal-responsive regulons. We also observed perturbations in membrane bioenergetics pathways, suggesting multiple cellular targets. Complementary directed evolution experiments yielded xanthoquinodin-resistant mutants harboring mutations in the iron-uptake gene feoB and the iron-responsive oxidoreductase fldX, providing genetic evidence that iron metabolism interference underlies compound bioactivity. These findings position xanthoquinodins as promising leads for target exploration in C. difficile, meriting further investigation into their mechanisms of action and therapeutic potential.

Completion Date

2025

Semester

Summer

Committee Chair

Self, William

Degree

Master of Science (M.S.)

College

College of Medicine

Department

Burnett School of Biomedical Sciences

Format

PDF

Release Date

2-15-2026

Document Type

Thesis

Campus Location

Orlando (Main) Campus

Subjects

Clostridium difficile; Antibacterial agents--Research; Fungal metabolites--Toxicology; Bacterial diseases--Chemotherapy; Microbial metabolites--Physiological effect

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