Dietary Supplementation of Bacillus subtilis Improves In Vitro Rumen Microbial Fermentation When Using Macadamia integrifolia Husk as a Substrate

Macadamia integrifolia husk (MIH) is a major byproduct of Macadamia integrifolia processing; however, there is limited information available on its rumen fermentation characteristics and associated bacterial communities when used as a feed for ruminants. This study evaluated the effects of different Bacillus subtilis (BS; ≥1013 CFU/g) inclusion levels on MIH using an in vitro technique. A total of six healthy goats (female, 7-month-old) with an average body weight of 15.20 ± 0.20 kg were selected as rumen fluid donors, and the rumen fluid was collected before morning feeding. Four BS inclusion levels were studied: a control group with 0 g/kg (CON), and three treatment groups supplemented with 1.5 g/kg (low dose, BSL), 3.0 g/kg (medium dose, BSM), and 4.5 g/kg (high dose, BSH) of BS. MIH supplemented with BS improved the gas production at all the observed incubation times (p < 0.05). The in vitro dry matter degradability was increased by 3.5%, 2.0%, and 3.1% in the BSL, BSM, and BSH groups, respectively, compared to the CON group at a 48 h incubation time (p = 0.049). The in vitro neutral detergent fiber degradability was increased by 6.0% and 7.8% in the BSL and BSM groups, respectively, compared to the CON group at a 48 h incubation time (p = 0.027). The in vitro acid detergent fiber degradability was increased by 11.7%, 16.8%, and 3.2% in the BSL, BSM, and BSH groups, respectively, compared with the CON group at the 48 h incubation time (p = 0.014). The concentrations of ammonia-N, microbial protein, total VFAs, acetate, propionate, butyrate, iso-butyrate, valerate, and iso-valerate were quadratically changed with increasing BS supplementation levels (p < 0.05). The relative abundances of Bacteroidota and Pseudomonadota were highest in the BSL group and lowest in the BSM group (p < 0.05), whereas Bacillota and Kiritimatiellota were highest in the BSH group and lowest in the BSL group (p < 0.05). Synergistota was highest in the BSM group and lowest in the BSH group (p < 0.05). The relative abundances of norank_p_Bacteroidota and norank_o_Bacteroidoles were highest in the BSL group, whereas they were lowest in the BSM group (p < 0.05). The relative abundances Hoyesella and Succinivibrio were highest in the BSL group, whereas they were lowest in the BSH group (p < 0.05). The relative abundances of Prevotella, Succiniclasticum, and Selenomonas were highest in the BSM group, whereas they were lowest in the BSL group (p < 0.05). These in vitro results indicate that supplementation with Bacillus subtilis could improve the utilization of MIH in goats, which is possibly associated with the altered rumen bacteria communities. The supplementation of 1.5 to 3.0 g/kg Bacillus subtilis (≥1013 CFU/g) on Macadamia integrifolia husk could improve its utilization as a feasible alternative feed for ruminants using an in vitro technique.