Multi-omics analysis reveals the potential for fermented Cordyceps militaris mushroom substrate in laying hens.

This study examines how varying levels of fermented Cordyceps militaris mushroom substrate (CMMS) in laying hen diets affect production performance, digestive health, immunity, cecal microbiota, metabolites, and quorum-sensing functions. Fermentation reduced CMMS dry matter, NDF, and phosphorus content (p < 0.05). Replacing 30% of the diet with fermented CMMS significantly improved laying rate, egg weight, feed intake, and feed efficiency (p < 0.05), while enhancing yolk color, Haugh units, and lipase activity. A 20% substitution increased nutrient digestibility and immunoglobulin levels (p < 0.05). Metagenomic analysis revealed increased abundance of Phocaeicola, Alistipes, and Parabacteroides (p < 0.05) with enhanced energy metabolism and specific gene families. Metabolomic analysis identified 1,529 differentially expressed metabolites, with carboxylic acids being most prevalent (21.20%), and enhanced taurine/hypotaurine metabolism and GPI-anchor biosynthesis. Parabacteroides showed negative correlations with certain metabolites, while Alistipes correlated positively with PemK/MazF family genes (p < 0.001). CMMS fermented feed proportions influence cecal microbiota, their metabolites, and quorum sensing in laying hens, affecting production, digestibility, immunity, metabolism, and health, demonstrating CMMS potential as alternative poultry nutrition.