The inhibition of high ammonia to in vitro rumen fermentation is pH dependent

Ammonia is an important rumen internal environment indicator. In livestock production, feeding a large amount of non-protein nitrogen to ruminants will create high ammonia stress to the animals, which increases the risk of ammonia toxicity. However, the effects of ammonia toxicity on rumen microbiota and fermentation are still unknown. In this study, an in vitro rumen fermentation technique was used to investigate the effects of different concentrations of ammonia on rumen microbiota and fermentation. To achieve the four final total ammonia nitrogen (TAN) concentrations of 0, 8, 32, and 128 mmol/L, ammonium chloride (NH₄Cl) was added at 0, 42.8, 171.2, and 686.8 mg/100 mL, and urea was added at 0, 24, 96, and 384 mg/100 mL. Urea hydrolysis increased, while NH₄Cl dissociation slightly reduced the pH. At similar concentrations of TAN, the increased pH of the rumen culture by urea addition resulted in a much higher free ammonia nitrogen (FAN) concentration compared to NH₄Cl addition. Pearson correlation analysis revealed a strong negative correlation between FAN and microbial populations (total bacteria, protozoa, fungi, and methanogens) and in vitro rumen fermentation profiles (gas production, dry matter digestibility, total volatile fatty acid, acetate, propionate, etc.), and a much weaker correlation between TAN and the above indicators. Additionally, bacterial community structure changed differently in response to TAN concentrations. High TAN increased Gram-positive Firmicutes and Actinobacteria but reduced Gram-negative Fibrobacteres and Spirochaetes. The current study demonstrated that the inhibition of in vitro rumen fermentation by high ammonia was pH-dependent and was associated with variations of rumen microbial populations and communities.