Residual feed intake phenotype modulates growth performance and ruminal microbiome responses to a Bacillus-based direct-fed microbial in beef steers

Dietary supplementation with direct-fed microbials may improve cattle performance through rumen microbiome modulation; however, responses are inconsistent and may depend on host feed-efficiency phenotype. This study evaluated the effects of a Bacillus -based direct-fed microbial (DFM) on growth performance and the ruminal microbiome of beef steers with divergent residual feed intake (RFI) phenotypes. Following a 49-d RFI test in 108 Angus × Hereford beef steers (initial BW = 242 ± 14.4 kg), 40 steers (BW = 332 ± 18 kg) representing negative (n = 20; mean RFI = − 1.66 kg/d, more feed efficient) or positive (n = 20; mean RFI = 1.62 kg/d, less feed efficient) RFI phenotypes were used in a 70-d feeding trial arranged as a 2 × 2 factorial with RFI phenotype (negative vs. positive) and diet (control vs. Bacillus -based DFM; 14 g/steer/d; 1.5 × 10 ⁸ CFU/g; Papillon, Easton, MD) as main effects. Treatments were replicated across 12 pens (3 pens/treatment; 3–4 steers/pen). Rumen fluid collected on d 70 was used for 16S rRNA gene (V3–V4) sequencing; sequences were processed in QIIME 2 with DADA2, and microbiome data were analyzed using MicrobiomeAnalyst (microbiomeanalyst.ca). A diet × RFI interaction was detected for final BW and ADG ( P  = 0.02), with Bacillus supplementation increasing final BW (489 vs. 467 kg) and ADG (2.19 vs. 2.05 kg/d) in negative-RFI steers but not in positive-RFI steers. Dry matter intake differed by RFI phenotype ( P  = 0.03) but was not affected by diet. Chao1 richness and microbial community composition (beta diversity) exhibited diet × RFI interactions ( P  = 0.05 and P  = 0.01, respectively). Corresponding diet × RFI interactions were detected for relative abundance of Bacteroidota , Firmicutes, Prevotella , Ruminococcus , and Lachnospiraceae _NK3A20_group; in negative-RFI steers, Bacillus supplementation increased the relative abundance of Bacteroidota and Prevotella , whereas in positive-RFI steers, supplementation increased the relative abundance of Firmicutes, Ruminococcus , and Lachnospiraceae _NK3A20_group. These results indicate that Bacillus -based DFM responses are phenotype-dependent, with more feed-efficient cattle benefiting more from supplementation, supporting precision supplementation strategies based on feed-efficiency classification.