Abstract
Rumen microbes produce fermentation products that meet the nutritional needs of the host ruminant animal. Previous research establishes that there are differences in microbial communities of solid versus liquid microenvironments within the rumen. However, this research has utilized 16S rRNA gene sequencing, which is limited because this sequencing strategy only provides information at the genus level. Alternatively, whole genome shotgun sequencing (WGS) facilitates analysis of the whole genome of a microorganism, generating data at the strain level which provides information on the functionality of the microorganism. The objective of our study was to characterize rumen microorganisms differentially associated with solid or liquid rumen microenvironments using WGS. Four ruminally cannulated steers were utilized in a 4×4 Latin Square project and were provided ad libitum access to hay, water, and a trace mineral block, in addition to alfalfa cubes at 200 g/kg of hay intake. On d 14 of each 28-d period, ruminal contents were collected from four areas of the ventral and dorsal sac of the rumen and separated into liquid (n=16) and solid (n=16) microenvironments. DNA was extracted from rumen samples and sequenced using WGS. Our findings indicate there were significant differences in alpha diversity (P< 0.01) and species richness (P=0.01) such that the liquid environment had a greater number of species present and higher diversity than the solid environment. Beta diversity using non-metric multidimensional scaling (NMDS) revealed a significant difference (P≤0.05) between the solid and liquid microenvironment clusters. In the liquid microenvironment, species that have known associations with propionate production and are responsible for the breakdown of polysaccharides were present in significantly higher abundances than in the solid microenvironment – e.g., uncultured Prevotellaceae bacterium (P≤0.01) and Prevotella sp. (P≤0.01). Species that were present in higher abundances in the solid microenvironment included Ruminococcaceae bacterium (P≤0.01) and uncultured Lachnospiraceae bacterium (P≤0.01), both of which are fibrolytic. The species uncultured Methanobrevibacter sp., which is associated with methane production, was found at a higher abundance in the solid microenvironment (P=0.06) compared to the liquid microenvironment. This data provides valuable insights into the different rumen microorganisms present in either solid or liquid microenvironments, building on existing literature with findings from next-generation sequencing.