Determination of the bacterial communities in the rumen and their impact on protein breakdown and carbohydrate utilization.

摘要

Ruminal microbial fermentation requires an elaborate collaboration of several diverse species, which begins with the colonization of plant surfaces, facilitating the breakdown of cellulose and protein. Consequently, identifying the microbial community structure and interactions occurring among the members of the consortium is important for understanding dynamics of bacterial communities that are important for the nutritional status of the host animal.;This thesis uses a general microbial ecology approach to investigate aspects of bacterial communities involved in protein breakdown and carbohydrate utilization in the rumen. Degenerate PCR, 16S rRNA sequencing, and phylogenetic analysis of the strains identified were genetic tools used to elucidate information on the composition of this community. Two specific genes were used as indicators for microbial interactions. Serme-like proteases genes were targeted to determine whether proteolytic activity existed for four predominant ruminal bacteria. The results of this work demonstrated that diversity of ruminal bacteria can be inhibitory to PCR based approaches, with numerous non-specific products generated and few putative serine proteases identified. It was concluded that the serine-like protease gene isolated could not be used to identify proteolytic bacteria within the rumen microbial community. The luxS gene was identified from members in the microbial community that may be involved in interspecies communication via quorum sensing during colonization. The luxS study found four gene types among species in the genera Prevotella, Butyrivibrio, Streptococcus, and Ruminococcus , with genotypes also obtained from ruminal fluid samples of uncultured bacteria.;A second approach to examine community structure was used to determine the efficiency of anaerobic cultivation techniques. Bacterial isolates from ruminants on a low forage diet with high concentrate supplements were identified as mostly Streptococcus and Butyrivibrio species. Phylogenetic analysis identified three major groups consisting of a monophyletic Streptococcus clade and two Butyrivibrio clades, which reveal that cultivation techniques are only able to identify a subset of the community that utilizes starch.;This thesis provides a framework for the next step in understanding rumen nutrition from a micro-ecological perspective. Providing a window into the overall diversity of the rumen is extremely important for understanding increased nutritional efficiency and health of ruminants.