Abstract
Ursolic acid (UA), a naturally occurring pentacyclic triterpenoid abundant in various plants, possesses potent biological activities. However, its effects and mechanisms on immune competence in broilers remain unclear. In this study, 320 one-day-old Cobb broilers were randomly allocated to four groups (8 replicates of 10 birds each) for a 42-day trial: a control group (CON) and three treatment groups supplemented with 50, 200, or 400 mg/kg UA (UA 50, UA 200, or UA 400). We employed enzyme-linked immunosorbent assay (ELISA), alcian blue-periodic acid schiff (AB-PAS) staining, immunofluorescence (IF), immunohistochemistry (IHC), qRT-PCR, metagenomics, and untargeted metabolomics to analyze the effects of UA on immune factors, inflammatory cytokines, intestinal barrier function, regulatory T (Treg) cell / T helper 17 (Th17) cell balance, as well as intestinal microbial composition and metabolism in broilers. The results indicated that UA significantly increased immune factor levels while reducing pro-inflammatory cytokine concentrations in broilers. Regarding intestinal barrier function, UA supplementation effectively reduced lipopolysaccharide (LPS) and D-lactic acid levels, promoted goblet cell proliferation, and enhanced the expression of tight junction proteins (Claudin-1, ZO-1). Notably, UA also significantly modulated Treg/Th17 balance. Furthermore, UA supplementation modulated the gut microbial composition, which was marked by an increase in the beneficial Lactobacillus johnsonii and a concurrent suppression of the pathobiont Escherichia coli. Furthermore, UA reduced the enrichment of microbial pathways associated with pathogenic Escherichia coli and Salmonella infection. Further analysis indicated that UA modulated propionate and tryptophan metabolism, thereby increasing the concentrations of propionic acid and the tryptophan metabolites (5-Hydroxyindole-3-Acetic Acid (5HIAA) and Indole-3-Acetic Acid (IAA)). In summary, our findings demonstrate that UA enhances broiler immunity and intestinal barrier function. These benefits appear to be mediated by the UA-driven enrichment of Lactobacillus johnsonii, which promotes the production of propionate and tryptophan-derived metabolites (5-HIAA and IAA), thereby rebalancing the Treg/Th17 balance and ultimately reinforcing intestinal integrity. These findings underscore the potential of UA as a natural supplement for sustainable poultry production.