The Effect of Binding Feed Enzymes to Spores of Bacillus Subtilis (var. Natto) on In Vivo Digestibility and In Situ Disappearance

The objective of this study was to determine whether binding a commercial xylanase feed enzyme to the surface of Bacillus subtilis spores would improve feed digestibility compared to free enzyme. An in vivo study was conducted using 4 treatments: control (CON; no enzyme), B. subtilis spores (SPR; 1.0 × 10¹⁰ spores/kg DMI), free enzyme (ENZY; 2.5 ml of commercial xylanase enzyme/kg DMI), and spore-bound enzyme (SBE; 2.5 ml of commercial xylanase enzyme bound to 1.0 × 10¹⁰ spores/kg DMI) applied to alfalfa grass hay. The treatments were fed to 4 ruminally and duodenally cannulated heifers in a 4 × 4 Latin square design, with 21 d experimental periods and a 7 d washout between periods. Ruminal in situ disappearance was determined on d 13 to 18, where triplicate polyester bags containing alfalfa grass hay (5 g) were removed after 0, 4, 8, 12, 24, 36, 48, 72, 96 and 120 h of incubation. Total collection of feces and feed refusals occurred on d 19 to 21 to measure total tract digestibility. Rumen and fecal samples were collected and processed for 16S rRNA gene sequencing to analyze bacterial microbiota. There were no differences in ruminal in situ rates of disappearance of DM, OM, NDF or ADF (P ≥0.086). Effective rumen degradability of DM and OM were greater for ENZY, SBE, and SPR compared to CON (P ≤ 0.048), but there was no difference between ENZY and SBE (contrast P ≥ 0.146). Total tract and ruminal DM and OM digestibility were not affected by treatment (P ≥ 0.067); however, intestinal DM and OM digestibility were increased for SBE (SBE vs ALL, P ≤ 0.043). There was also an increase in total tract digestibility of NDF and ADF for SBE (SBE vs ALL, P ≤ 0.048). Rumen microbiota diversity was affected by treatment, with differences observed between CON and SPR (P = 0.050), CON and SBE (P = 0.033), and SPR and SBE (P = 0.009). There was no difference in fecal microbiota diversity between CON and SPR (P = 0.113) or SPR and SBE (P = 0.406), but fecal microbiota was different between CON and SBE (P = 0.009) as well as ENZY and SBE (P < 0.001). These results suggest that spore-bound enzymes are more stable than free enzyme, increasing total tract digestibility of NDF and ADF. The effects likely resulted from enhanced post-ruminal digestion and alteration of the gastrointestinal microbiota.