Abstract
The objective of this experiment was to evaluate the effects of urea hydrolysis rate on ruminal bacterial diversity level and cellulolytic bacteria abundance in vitro. To control urea hydrolysis rate, urea and urease inhibitor (acetohydroxamic acid, AHA) were supplemented to a 2 × 2 factorial design, with urea supplemented at 0 or 20 g/kg dry matter (DM) of substrate, and AHA equivalent to 0 or 450 mg/kg DM of substrate. Ruminal fluid was collected from three Chinese Holstein dairy cows, fed a TMR, and incubated at 39 °C for 12 h after the addition of urea and AHA. Rumen fermentation parameters, which indicated the rate of ammonia formation (including ammonia-nitrogen (NH3-N) and urea-nitrogen concentrations, urease activity, and microbial crude protein) were measured by chemical analysis. Bacterial diversity was analyzed by denaturing gradient gel electrophoresis (DGGE). Total bacteria and cellulolytic bacteria abundance was detected by quantitative PCR. Results showed that AHA addition significantly decreased the rate of ammonia formation when urea was supplemented. Urea and AHA supplementation significantly increased the bacterial community diversity level according to the Shannon–Weiner index of 16S DGGE images. Furthermore, ruminal bacterial profiles were separated by ammonia release rate when urea was supplemented, according to the DGGE and hierarchical cluster analysis. Urea supplementation reduced the abundance of cellulolytic bacteria, such as Ruminococcus albus, R. flavefaciens, Fibrobacter succinogenes, and Butyrivibrio fibrosolvens, but inhibition of urea hydrolysis by AHA addition alleviated the reductions during the early period of incubation. In conclusion, slow release of ammonia induced by urease inhibitor influenced the ruminal bacterial diversity level and lessened the inhibition of total bacteria growth at the incubation of 12 h and F. succinogenes during the early period of incubation.
Highlights
During the early and middle lactation periods in dairy cows, high dietary energy, and crude protein (CP) are required to maintain high milk production
Inhibition of urea hydrolysis and ammonia formation The urea-N concentration could not be detected in the treatment with urea supplementation only by 1 h postincubation, but it gradually decreased until 6 h of incubation in the treatment with both urea and acetohydroxamic acid (AHA) supplementation (Table 2)
The concentration of NH3-N was increased by urea supplementation through the whole incubation (P < 0.05), but it was numerically decreased by AHA addition within 2 h of incubation
Summary
During the early and middle lactation periods in dairy cows, high dietary energy, and crude protein (CP) are required to maintain high milk production. Rapid hydrolysis of dietary urea by bacteria in the rumen leads to a dramatic increase in the concentration of ammonia in the first few hours after feeding. Various strategies are implemented to reduce the rapid hydrolysis of dietary urea into ammonia in the rumen, such as the application of urease inhibitor to diet (Upadhyay, 2012). It is reported that urease inhibitor can effectively delay urea hydrolysis (slow release urea) and decrease the rate of ammonia formation in the rumen without a negative effect on nitrogen metabolism (Zhang, Shan & Bao, 2002). Slow release urea can increase milk fat, protein, and milk energy output of dairy cows during lactation period (Highstreet et al, 2010). There is limited information on rumen bacterial population induced by slow hydrolysis of urea
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