Abstract
With the growing demand for antibiotic-free and sustainable poultry production, plant-derived antimicrobials have emerged as promising alternatives. However, a systematic understanding of the combined effects of oregano essential oil (OEO) and Macleaya cordata extract (MCE) on the broiler gut microbiome remains lacking. This study employed an integrated "structure-function-phenotype" framework to investigate the individual and combined (OMS) effects of OEO and MCE on gut microecological remodeling and its coupling with host growth, metabolic, and immune phenotypes. A total of 960 one-day-old broiler chicks were individually weighed and then randomly allocated to four treatments using body-weight-stratified randomization, with 6 replicate pens per treatment and 40 birds per pen, to ensure similar initial body weight across groups. Over a 42-day trial, we evaluated growth performance, serum biochemistry, antioxidant status, and immune parameters. Compared to the control, the OMS treatment significantly enhanced average daily feed intake (ADFI) and average daily gain (ADG), increased serum total protein (TP), and decreased blood urea nitrogen (BUN), triglycerides (TG), total cholesterol (TC), and alkaline phosphatase (ALP). However, the feed-to-gain ratio (F/G) was also higher in the OMS group, indicating that the improvement in growth rate did not translate into enhanced feed efficiency but was primarily driven by increased feed consumption. OMS also improved overall antioxidant capacity and key enzyme activities, elevated immunoglobulin levels, and reduced pro-inflammatory cytokines. Notably, OMS maintained Lactobacillus dominance, enriched Bacteroides, Enterococcus, and Butyricicoccus, and reduced Escherichia-Shigella. Functional predictions via PICRUSt2 suggested enhanced metabolic pathways related to antioxidant and immune functions; however, these results represent inference-based predictions and should be interpreted cautiously. Overall, the combination of OEO and MCE exerted synergistic benefits on growth, physiological health, and gut microbiota, supporting its potential as a phytogenic strategy for antibiotic-free broiler production.