Effects of different doses of Bacillus subtilis Natto on in vitro rumen fermentation parameters

Abstract Text: This experiment was conducted to investigate the effects of different doses (0 (contrl), 0.2×10¹¹cfu/ml (BSL),1×10¹¹cfu/ml (BSM), 5×10¹¹cfu/ml (BSH)) of Bacillus subtilis Natto culture (BSN) on rumen fermentation in vitro. Ruminal fluid was collected from three lactating Holstein dairy cows (BW=558±10 kg, DIM = 153 ±16 d) fed diets contained (DM basis) Chinese wildrye (3.7%), Alfalfa hay (28.4%), Corn silage (26.7%), Corn (22.6%), Soybean meal (11.8%), Cottonseed fuzzy (5.1%), CaHPO₄ ( 0.6%), NaCl (0.5%), Premix (0.6%). Diets were mixed with phosphate buffer (1:2), incubated (90ml) anaerobically at 39°C for 0h, 12h, 24h, 36h and shaken at 50rpm. Each substrate (500mg DM basis) which had same concentrate mixture with donors was added to. Four sets of bottles were sealed anaerobically under CO₂ atmosphere with butyl rubber stoppers and capped with aluminum. One set (0h) were sampling immediately after mixed, other sets were incubated 12h, 24h, 36h respectively. Each set contained four levels of supplements, and for each level, three bottles were incubated. Statistical analysis was carried out by ANOVA (GLM) using SAS (SAS9.2). Medium pH was unaffected ( P = 0.61) by BSN in all treatments, ammonia-N tended to increase linearly with increasing BSN dose ( P < 0.01). Compared with the control, the molar proportion of acetate(46.66 vs 57.32, 60.96 and 68.17, P < 0.01), propionate (19.23 vs 22.88, 24.48 and 29.98 linear, P < 0.01), iso-butyrate (0.83 vs 2.18, 2.50 and 3.70, P < 0.01), iso-valerate (1.55 vs 4.50, 5.02 and 7.70, P < 0.01), valerate (1.52 vs 4.38, 5.04 and 6.89, P < 0.01) and total VFA (79.93 vs 102.77, 110.12 and 131.03, P < 0.01) was enhanced significantly with increasing concentrations of BSN. The molar proportion of butyrate of BSL treatment was increased (P < 0.05) and BSM and BSH treatment were increased significantly (P < 0.01). The molar proportion of acetate:propionate was increased (P < 0.05) in BSL and BSM treatment compared with control, however, decreased significantly in BSH treatment (P < 0.01). The results indicated that the Bacillus subtilis Natto culture can stimulated in vitro fermentation by increase the molar proportion of VFA and change the rumen fermentation type by change the molar proportion of acetate:propionate, and the diets should consist of a minimum of 10¹⁰cfu/g (DM) of BSN to make sure it’s effect.

Keywords: Bacillus subtilis Natto, in vitro, rumen fermentation