The objectives of this study were to evaluate the effect of hexanoic acid (HA) supplementations (0, as the control, CON; 0.05%, HA1; 0.1%, HA2; 0.2%, HA3) on β-carotene, and ascertain the way and key factors of HA influencing β-carotene content of alfalfa (Medicago sativaL.) after ensiled in an oxygen-free and dark conditions for 10, 40, and 80 d (from May to August, 2021). This was achieved by examining the dynamic change of β-carotene, activities of β-carotene-related enzymes, and bacterial community succession of ensiled alfalfa, using operoncrtNMidentification,crtEgene quantitation, and single-molecule real-time sequencing technology. The results revealed that when compared with the fresh material, terminal alfalfa silage treated with different level of HA supplementations (0, 0.05, 0.1, 0.2%; fresh weight basis) increased β-carotene content up to 2.86, 85.8, 159, and 133%, accordingly. Meanwhile, alfalfa silage treated with higher levels of HA (0.1 and 0.2%) showed superior effects compared to those treated with lower levels of supplementation (0 and 0.05%). HA supplementation specifically facilitated the increase abundance of Lactobacillus kullabergensisand the emergence of L.senioris.Multiple linear regression models inferred thatL.kullabergensis,L.apis, L.saniviri, L.senioris,peroxidase, phytoene desaturase, and lycopene β-cyclasepositively regulated β-carotene. Conversely,LactobacillusrenniniandL.brevisadjusted β-carotene, negatively. Positive regulations of the above bacterial species and enzymes had a stronger role in increasing β-carotene than L.renniniandL.brevis.In conclusion, the β-carotene increase of ensiled alfalfa may be regulated by HA supplementation viamultiple positive factors, including 4 specialLactobacillusspecies (L.kullabergensis, L.apis, L.saniviri, and L.senioris), and 3 vegetative β-carotene-related enzymes (peroxidase, phytoene desaturase, and lycopene β-cyclase).

BMP gene family  /  expression regulation  /  BMP7  /  myogenesis  /  adipogenesis