Taxifolin impairs colonization of Clostridium perfringens in association with iron limitation-related physiological stress

Abstract

ABSTRACT Flavonoids are plant-derived polyphenols with diverse biological activities; however, the mechanisms by which individual compounds inhibit intestinal pathogens remain unclear. In this study, we found that taxifolin (dihydroquercetin) showed pronounced inhibitory activity against Clostridium perfringens, a major gram-positive enteropathogen, whereas most other bacterial species tested were less affected. Notably, at concentrations below those causing complete growth inhibition, taxifolin significantly impaired mucin adhesion and biofilm formation, indicating suppression of colonization-associated phenotypes rather than bactericidal activity. In an antibiotic-pretreated mouse infection model, oral administration of taxifolin significantly reduced C. perfringens intestinal colonization during the early phase of infection. Mechanistically, proteomic analysis of C. perfringens revealed marked alterations in surface-associated and extracellular proteins, including stress response factors (MreB, LytR, and ClpB), and enzymes linked to iron- and redox-dependent metabolism, consistent with an iron-limiting stress response. Scanning electron microscopy further revealed pronounced cell elongation in taxifolin-treated cells, consistent with impaired cell division under stress conditions. Importantly, Fe2+ supplementation partially restored bacterial growth, normal morphology, mucin adhesion, and biofilm formation, whereas supplementation with other divalent ions failed to restore growth. Similar phenotypes were observed following treatment with the iron-specific chelator 2,2′-bipyridyl. Collectively, our findings support a working model in which iron limitation-associated physiological stress contributes to the inhibitory effects of taxifolin on C. perfringens growth and colonization-related phenotypes. This study provides insights into the interaction between dietary flavonoids and clostridial pathogens, highlighting the potential of taxifolin as a plant-derived compound for limiting C. perfringens intestinal colonization. IMPORTANCE Clostridium perfringens, a clinically important pathogen in both humans and animals, causes various histotoxic and enteric diseases, including gas gangrene and foodborne or non-foodborne diarrhea. Its control remains challenging due to increasing antimicrobial resistance and the need to preserve beneficial gut microbiota. Here, we identified taxifolin, a naturally occurring flavonoid, as a potential inhibitor of C. perfringens that suppressed its growth and colonization-associated phenotypes while exerting comparatively limited effects on most other tested bacterial species, including commensal bacteria. Proteomic and physiological analyses revealed stress responses associated with iron metabolism, redox balance, and cell division, which were accompanied by reduced mucin adhesion, biofilm formation, and bacterial proliferation. Our findings suggest that taxifolin can reduce the colonization capacity of C. perfringens and support its potential use as a plant-derived approach for limiting C. perfringens intestinal colonization in humans and animals. Clostridium perfringens, a clinically important pathogen in both humans and animals, causes various histotoxic and enteric diseases, including gas gangrene and foodborne or non-foodborne diarrhea. Its control remains challenging due to increasing antimicrobial resistance and the need to preserve beneficial gut microbiota. Here, we identified taxifolin, a naturally occurring flavonoid, as a potential inhibitor of C. perfringens that suppressed its growth and colonization-associated phenotypes while exerting comparatively limited effects on most other tested bacterial species, including commensal bacteria. Proteomic and physiological analyses revealed stress responses associated with iron metabolism, redox balance, and cell division, which were accompanied by reduced mucin adhesion, biofilm formation, and bacterial proliferation. Our findings suggest that taxifolin can reduce the colonization capacity of C. perfringens and support its potential use as a plant-derived approach for limiting C. perfringens intestinal colonization in humans and animals.