Gut bacterial species, serum metabolites, and serum cytokines associated with broodiness in chickens

ABSTRACT Increasing evidence suggests that the gut microbiota, serving as a “virtual endocrine organ”, potentially modulates reproductive behavior in poultry via the gut-brain and gut-ovary axes. Broodiness in hens inhibits egg-laying activity and causes major economic losses in native chicken breeds, but its micro-physiological basis remains unclear. This study used shotgun metagenomic sequencing to delineate the cecal bacterial species associated with brooding status in Chinese Kangle chickens. We identified 34 cecal bacterial species exhibiting significantly varying abundances between the broodiness and control groups, including six species (e.g., Bacteroides sp. An51A and Phocaeicola barnesiae ) that were significantly enriched in the broodiness group. Additionally, 28 species significantly enriched in the control group were screened. Among them, Subdoligranulum variabile and Oribacterium asaccharolyticum served as key biomarkers for distinguishing brooding status in Kangle chickens and were associated with functional shifts in the cecal microbiome. Non-targeted metabolomic analysis identified 17 differential metabolites, among which seven (e.g., (13E) -11a-hydroxy-9,15-dioxoprost-13-enoic acid and d-arabitol) were defined as metabolic markers of the broody state and were significantly associated with Subdoligranulum variabile and Oribacterium asaccharolyticum . In addition, our results suggest that serum cytokines, such as IFN-γ and IL-22, are potentially associated with the broody state and the alterations in both serum metabolites and the gut microbiota (e.g., Subdoligranulum variabile and Oribacterium asaccharolyticum ). These findings provide a new insight into the mechanisms underlying reproductive behavior in poultry and offer a theoretical basis for alleviating broodiness through microecological interventions, thereby improving the reproductive efficiency of indigenous chicken breeds.