FIGURE 1.

Experimental flow chart. The experiment was divided into three steps. The first step is pre-incubation: 1 g of each stool sample was suspended in 10 ml of PBS and allowed to stand for 5 min, after which 0.5 ml of the suspension was added to blood culture bottles containing sheep’s blood and rumen fluid. The bottles were divided into two groups: the supplemented group (S) and non-supplemented group (NS). The supplemented group was then further divided into the aerobic supplemented group (O₂S) and the anaerobic supplemented group (AS). Similarly, the non-supplemented group was divided into the aerobic non-supplemented group (O₂) and the anaerobic non-supplemented group (A). The second step was subculturing and enrichment: samples were taken from the pre-cultures every 3 days and subcultured aerobically and anaerobically on solid YCFA media, and the pH of the pre-cultures was measured using a pH meter. When colonies appeared on the plates, individual colonies were picked and transferred to liquid culture enrichment medium (YCFA) in 24-well plates. The last step was bacterial storage and identification: the enrichment cultures were frozen in glycerol and inoculated to solid medium at the same time. Then, the colonies were tested by MALDI-TOF mass spectrometry. Unidentified colonies were evaluated by 16S rRNA gene sequencing. All bacteria were incubated aerobically for 1 day or anaerobically for 3 days at 37°C.