Background:Previous studies have evaluated the metabolic status of animals fed direct-fed microbial (DFM) using enzyme-based assays which are time-consuming and limited to a few metabolites.In addition,little emphasis has been placed on investigating the effects of DFM on hindgut microbiota.We examined the effects of dietary supplementation of a blend of Saccharomyces cerevisiae-based DFM and fermentation products on the plasma concentrations of carbonyl-containing metabolites via a metabolomics approach,and fecal bacterial community,via 16S rRNA gene sequencing,of beef steers during a 42-day receiving period.Forty newly weaned steers were randomly assigned to receive a basal diet with no additive (CON;n =20) or a basal diet supplemented with 19 g of CommenceTM (PROB;n =20) for a 42-day period.CommenceTM (PMI,Arden Hills,MN) is a blend of 6.2 x 1011 cfu/g of S.cerevisiae,3.5 x 1010 cfu/g of a mixture of Enterococcus lactis,Bacillus subtilis,Enterococcus faecium,and Lactobacillus casei,and the fermentation products of these aforementioned microorganisms and those of Aspergillus oryzae and Aspergillus niger.On d 0 and 40,rectal fecal samples were collected randomly from 10 steers from each treatment group.On d 42,blood was collected for plasma preparation.Results:A total number of 812 plasma metabolites were detected.Up to 305 metabolites [fold change (FC) ≥ 1.5,FDR ≤ 0.01] including glucose,hippuric acid,and 5-hydroxykynurenamine were increased by PROB supplementation,whereas 199 metabolites (FC ≤ 0.63,FDR ≤ 0.01) including acetoacetate were reduced.Supplementation of PROB increased (P ≤ 0.05) the relative abundance of Prevotellaceae UCG-003,Megasphaera,Dorea,Acetitomaculum,and Blautia.In contrast,the relative abundance of Elusimicrobium,Moheibacter,Stenotrophomonas,Comamonas,and uncultured bacterium belonging to family p-2534-18B5 gut group (phylum Bacteroidetes) were reduced (P ≤ 0.05).Conclusions:The results of this study demonstrated that supplementation of PROB altered both the plasma carbonyl metabolome towards increased glucose concentration suggesting an improved energy status,and fecal bacterial community,suggesting an increased hindgut fermentation of the beef steers.