Assessment of cecal microflora

At the end of the experiment, cecal digesta samples were collected from eight birds from each treatment group. Briefly, 1 g of fresh digesta was collected from the middle portion of the cecum, stored in bottles with a CO₂ current, and transported to the laboratory to estimate bacterial counts, as described by Long et al. (2021b). Digesta samples were suspended in 99 mL sterile 0.9% saline and homogenized in a stomacher for 5 min. Each homogenate was diluted 10-fold (10% w/v) in sterile ice-cold normal saline. Diluted samples (0.1 mL) were inoculated onto selective agar for bacterial enumeration. E. coli was incubated on MacConkey agar at 37 °C for 24 h. Lactobacillus spp. were incubated on Luria-Bertani agar and Briggs liver agar in an anaerobic incubator at 37 °C for 48 h. Agars were purchased from Beijing Ruizekang Technology Co., Ltd. (Beijing, China).