The objective of this experiment was to evaluate the effect of different EOC (0.1425% cobalt lactate + 1.13% oregano essential oil + 98.7275% carrier) levels on in vitro rumen fermentation and microbial changes. Six EOC levels (treatments: 0 mg·L⁻¹, CON; 50 mg·L⁻¹, EOC1; 100 mg·L⁻¹, EOC2; 400 mg·L⁻¹, EOC3; 800 mg·L⁻¹, EOC4 and 1500 mg·L⁻¹, EOC5) were selected to be used to in vitro incubation. The in vitro dry matter digestibility (IVDMD), in vitro neutral detergent fiber digestibility (IVNDFD), in vitro acid detergent fiber digestibility (IVNDFD), pH, ammonia-nitrogen (NH₃-N) concentration, total volatile fatty acid (TVFA) concentration and microbial protein (MCP) concentration were measured after 48 h incubation, after which the groups with significant nutrient digestibility and fermentation parameters were subjected to 16S rRNA sequencing. The results showed that the total gas production (GP) of the EOC5 group was higher than that of the other groups after 12 h of in vitro incubation. TVFA, NH₃-N and MCP concentrations were also shown to be higher in group EOC5 than those in other groups (p < 0.05), while NH₃-N and MCP concentrations in the EOC2 group were lower than those in other groups significantly (p < 0.05). The molar ratio of acetic acid decreased while the molar ratio of propionic acid increased after the addition of EOC. 16S rRNA sequencing revealed that the rumen microbiota was altered in response to adding EOC, especially for the EOC5 treatment, with firmicutes shown to be the most abundant (43.1%). The relative abundance of Rikenellaceae_RC9_gut_group was significantly lower, while the relative abundance of uncultured_bacterium_f_Muribaculaceae and Succiniclasticum was significantly higher in the EOC5 group than those in other groups (p < 0.05). Comprehensive analysis showed that EOC (1500 mg·L⁻¹) could significantly increase gas production, alter sheep rumen fermentation parameters and microbiota composition.

The effects of incorporating yeast culture (YC) into pelleted feeds on sheep production and the potential impact on rumen microbial populations, microbial metabolism, and fermentation have not been extensively studied. This study aimed to evaluate the effect of YC on growth performance, rumen tissue development, rumen fermentation, and rumen microflora in sheep and to explore the potential microbial mechanisms involved. Fifty healthy 3-month-old male lambs of small-tailed Han sheep, with an average weight of 28.44 ± 0.63 kg, were randomly divided into five groups: control (0% YC), 3% YC, 6% YC, 9% YC, and 12% YC. The pre-feeding period lasted for 15 days, followed by an official feeding period of 60 days. On the last day of the formal feeding period, six lambs that exhibited the best growth performance were randomly selected from the control group and the 9% YC group. These sheep were slaughtered, then the rumen epithelial tissue and rumen contents were collected for the measurement of rumen fermentation, microbial populations, and metabolites. Compared to the control group, the YC-treated groups showed higher daily and final body weight gains, as well as increased levels of propionic acid, butyric acid, and total volatile fatty acids (p < 0.05). YC supplementation also enhanced rumen papilla length and width (p < 0.05). Additionally, YC increased the relative abundance of certain microbial species (p < 0.05). These results suggest that supplementing 9% YC in pelleted diets for small-tailed Han sheep may enhance growth performance and improve the rumen environment.

This study used oregano essential oil (EO), cobalt (Co) and synergistic of both of them additives instead of antibiotics to explore the effects of different additives on the corn silage fibre structure degradation, nutrient degradation rate and rumen fermentation characteristics of sheep. The additives used in the study as a product were provided by Ralco, Inc., USA. The Latin square experimental design was adopted with 5 treatments. The following treatments were fed through the rumen fistula: carrier (Car), carrier + EO, carrier + Co, carrier + EO + cobalt (EOC) and no additives as the control (CON). These treatments were fed to five sheep for five rounds at a dose of 80 mg/kg. At 48 h of degradation, the area of degraded corn leaf and husk reached 100%, SEM results showed that except for Car and CON treatment, there were obvious destroyed fibre structure differences under EO, Co and EOC treatment, showing total disappearance of leaf epidermal cells, palisade tissue and spongy tissue, especially under EOC. Under EOC treatment, DMD_48 h, CPD_48 h, NDFD_48 h and ADFD_48 h increased by 3.33%, 3.89%, 7.64% and 5.67%, respectively, compared with CON, while NDFD_48 h and ADFD_48 h increased by 4.70% and 6.05% under Co (p > 0.05). The NH₃-N concentration in rumen fluid decreased by 19.36%, 27.20% and 19.48%, while the microbial protein (MCP) content increased by 6.32%, 4.29% and 5.97% (p > 0.05) under EO, Co and EOC treatments compared with CON at 48 h (p < 0.05). The total VFA concentration under EO, Co and EOC treatment was lower than CON at 6 and 48 h (p = 0.016, p = 0.022) and the propionate and acetate concentrations were higher under EOC treatment than CON at 48 h (p > 0.05). Feeding EOC increased the fibre structure degradation and nutrient degradation rate of corn silage and improved rumen fermentation in sheep, resulting in better effects than feeding EO or Co alone.

The effects of isochlorogenic acid (ICGA) on ewes rumen environment, microbial diversity, and immunity at different physiological stages (estrus, pregnancy and lactation) were studied in this experiment. Twenty healthy female Hu lambs of 1.5 months with similar body weight (17.82 ± 0.98 kg) and body condition were selected and randomly divided into two groups: the control group (CON) and the ICGA group (ICGA). The lambs of CON were fed a basal diet, while the lambs of ICGA were supplemented with 0.1% ICGA based on the basal diet. Lambs rumen fermentation characteristics, microbial diversity and immunity at estrus, pregnancy, and lactation stages were determined and analyzed, respectively. The results showed that the rumen pH in CON increased first and then decreased as lambs grew (p < 0.05). However, it showed the opposite change in ICGA. The content of ammonia nitrogen (NH₃-N) showed the highest at estrus stage in both groups, but it was significantly higher in ICGA than that in CON (p < 0.05). The Acetic acid/propionic acid (A/P) ratio at estrus stage and the volatile fatty acids (VFAs) at pregnancy stage in ICGA were significantly higher than those of the CON (p < 0.05). The 16S rDNA sequencing analysis showed that the Shannon, Chao 1 and ACE indexes of the ICGA were significantly higher than those of the CON both at estrus and lactation stages (p < 0.05), while they showed higher at the pregnancy stage in CON (p > 0.05). Principal component analysis (PCA) showed that there were significant differences in rumen microorganism structure between CON and ICGA at all physiological stages (p < 0.01). At the phylum level, compared with the CON, Firmicutes relative abundance of three physiological stages decreased (p > 0.05) while Bacteroidota increased (p > 0.05). The relative abundance of Synergistota at estrus stage and Patescibacteria at the lactation stage increased significantly too (p < 0.05). At the genus level, compared with the CON, the relative abundance of Prevotella at three stages showed the highest (p > 0.05), while the relative abundance of Succiniclasticum, unclassified_Selenomonadaceae and Rikenellaceae_RC9_gut_group showed different abundances at different physiological stages in ICGA. Compared with the CON, the lambs of the ICGA showed higher blood IgG, IgM, and TNF- α contents at three physiological stages and higher IL-6 contents at pregnancy stage (p < 0.05). Conclusion: Adding ICGA could regulate ewes rumen fermentation mode at different physiological stages by increasing rumen NH₃-N at estrus, VFAs at pregnancy, and the ratio of A/P at lactation. It optimizes rumen microbial flora of different physiological stages by increasing Bacteroidota relative abundance while reducing Firmicutes relative abundance, maintaining rumen microbial homeostasis at pregnant stage, increasing the number of beneficial bacteria in later lactating and ewes blood immunoglobulins content at three physiological stages.

The experiment was conducted to investigate the potential effects of steviol glycosides on growth performance, rumen fermentation processes, and microbial diversity in Hu sheep. A single-factor design was used for the trial. Twenty healthy weaned Hu lambs, possessing comparable body weights averaging 18.31 ± 1.24 kg, were randomly allocated into two distinct groups: the control group (CON) and the experimental group (STE), with each comprising 10 lambs. The CON was fed the basal diet, and the STE was supplemented with 0.07% steviol glycosides based on the basal diet. During the experimental period, variations in body weight and feed intake were closely monitored and recorded. After feeding for 90 d, blood was collected to determine blood biochemical indices, and rumen fluid samples were gathered for an in-depth analysis of rumen fermentation parameters and microbial diversity. The outcomes revealed no statistically significant differences in growth performance or serum biochemical indices between the two groups (p > 0.05). Rumen pH in STE and CON was within the normal range. The rumen ammonia nitrogen (NH₃-N) and acetic acid (AA) content of STE decreased significantly compared with CON (p < 0.05). No significant variations were observed in the levels of other volatile fatty acids (VFAs) between the two groups (p > 0.05). The rumen microbial OTUs count, as well as the Shannon, Simpson, Chao1, and Ace indices, were notably lower in the STE group compared to the CON group (p < 0.05). Additionally, at the phylum level, the relative abundance of Firmicutes, Bacteroidetes, and Proteobacteria collectively accounted for over 97% of the total phylum composition. In comparison to the CON group, the STE group exhibited an increase in the relative abundance of Proteobacteria (p < 0.05), accompanied by a significant reduction in the relative abundance of Patescibacteria and Desulfobacteria (p < 0.05). At the genus level, there was a notable increase in the relative abundance of Prevotella_7 and Succinivibrionaceae_UCG_001 in the STE group, whereas the relative abundance of Rikenellaceae_RC9_gut_group significantly decreased (p < 0.05). According to the correlation analysis between rumen microflora and VFAs, the relative abundance of Succinivibrionaceae_UCG_001 displayed a significant negative correlation with AA (p < 0.05), whereas Lactobacillus exhibited a notable positive correlation with isobutyric acid (IBA) (p < 0.05). In summary, steviol glycosides had no significant effect on the production performance and blood biochemical indexes of Hu sheep. Steviol glycosides can improve rumen fermentation parameters and rumen microflora structure of Hu sheep and have a certain effect on rumen microbial diversity and composition.

The sensory quality and flavor of lamb meat, critical to market competitiveness, are influenced by rumen microbial fermentation and dietary management strategies. Coated betaine (CBet), a rumen-protected methyl donor, exerts systemic nutritional regulation in ruminants. This study explored the effects of CBet supplementation on lamb meat quality using 18 Dorset ♂ × Hu sheep ♀ F1 crossbred lambs, randomly assigned to either a control group (basal diet) or a 0.20% CBet-supplemented diet for 60 days (n = 9 per group). The results demonstrated that CBet significantly increased ruminal concentrations of total volatile fatty acids (TVFAs), acetic acid, propionic acid, and butyric acid (p < 0.05). Additionally, CBet supplementation enhanced muscle redness (a*), crude fat, crude ash, heptadecanoic acid (C17:0), and tricosanoic acid (C23:0) (p < 0.05) while decreasing shear force and the concentration of cis-13,16-docosadienoic acid (C22:2) (p < 0.05). Furthermore, CBet elevated characteristic flavor compounds (e.g., nonanal) and their relative odor activity values (ROAVs), and decreased undesirable odors (e.g., dodecanal) (p < 0.05). As illustrated in the graphical abstract, these improvements were mediated through regulatory effects of CBet on rumen microbiota composition, muscle fatty acids, amino acids, and volatile flavor compounds. Specifically, CBet significantly increased the relative abundances of Firmicutes, Proteobacteria, Prevotella, and Bifidobacterium in the rumen (p < 0.05) and altered the Firmicutes/Bacteroidota ratio. In conclusion, dietary supplementation with 0.20% CBet effectively enhances lamb meat quality and flavor, effects closely associated with changes in the abundance of key ruminal microbial taxa.