In situ identification and quantification of protein-hydrolyzing ruminal bacteria associated with the digestion of barley and corn grain

Abstract

In this study, BODIPY FL DQ™ casein staining combined with fluorescence in situ hybridization (FISH) was used to detect and identify protein-hydrolyzing bacteria within biofilms that produced active cell-surface-associated serine- and metallo-proteases during the ruminal digestion of barley and corn grain in cows fed barley-based diets at 2 different levels. A doublet coccoid bacterial morphotype associated with barley and corn grain particles fluoresced after BODIPY FL DQ™ casein staining. Bacteria with this morphotype accounted for 3%–10% of the total bacteria attached to surface of cereal grain particles, possibly indicative of an important role in the hydrolysis of the protein matrix within the endosperm. However, the identity of these predominant proteolytic bacteria could not be determined using FISH. Quantitative FISH revealed that known proteolytic species, Prevotella ruminicola, Ruminobacter amylophilus, and Butyrivibrio fibrisolvens, were attached to particles of various cultivars of barley grain and corn, confirming their role in the proteolysis of cereal grains. Differences in chemical composition among different barley cultivars did not affect the composition of proteolytic bacterial populations. However, the concentrate level in the basal diet did have an impact on the relative abundance of proteolytic bacteria and thus possibly their overall contribution to the proteolysis of cereal grains.