2.2 Sample collection and preparation

At the end of the experiment, 2 squabs were randomly selected per replicate (a total of 12 squabs for each group) and sacrificed for postmortem analysis. Before this, the squabs were fasted overnight (12 h), and blood samples were collected from the necks of the squabs. The separated serum samples were stored at −80°C for subsequent analysis of blood biochemical, antioxidant, and immune indices. The squabs were subsequently slaughtered, and their ileum tissues were collected. Some of the intestinal tissues were fixed in a 4% paraformaldehyde solution for morphological observation. The remaining part was stored at −80°C for the detection of intestinal antioxidant indices, immune indices, tight junction proteins, and mRNA expressions of antioxidant-related genes. Additionally, the ileum contents of the squabs were collected and stored at −80°C for the analysis of intestinal digestive enzyme activity and microbial diversity.