Insights into the Populations of Proteolytic and Amino Acid-Fermenting Bacteria from Microbiota Analysis Using In Vitro Enrichment Cultures.

Abstract

This study evaluated the effects of nitrogen source composition and monensin on the populations of proteolytic and amino acid-fermenting bacteria using in vitro enrichment culture. The experiment was designed with a 2 × 2 factorial arrangement: two nitrogen sources, casein (Cas) and tryptone (Try), and two levels of monensin, 0 (C) and 5µmol/L (M), resulting in four treatments: Cas-C, Cas-M, Try-C, and Try-M. Ruminal fluid collected from three cannulated Holstein dairy cows was used as the inoculum. Each treatment culture was consecutively transferred six times after 24h of incubation. The results showed that ammonia concentration was lower in Cas than in Try, and it was reduced by monensin addition. In the 6th transfer enrichment cultures, the 16S rRNA gene copy numbers of total bacteria were reduced by monensin but was unaffected by nitrogen sources. Principal component analysis showed that the bacterial communities differed among the treatments. At the genus level, Peptostreptococcus accounted for as much as 41% of the total bacteria in Try-C, but it made up less than 0.02% in the other three treatments. A Pearson correlation analysis showed that the relative abundance of Peptostreptococcus was positively correlated with ammonia concentration. Overall, the results suggest that nitrogen source composition and monensin can affect ruminal ammonia production by modulating the ruminal proteolytic bacterial communities, and some hyper-ammonia-producing bacteria of the genus Peptostreptococcus may be among the main culprits contributing to the high ammonia concentration in the rumen.