Abstract:

[Objective] This study was conducted to assess the effect of Bacillus subtilis culture supernatant on sheep β-defesin-1 （SBD-1） expression in ovine ruminal epithelial cells （ORECs）. [Method] The corresponding supernatants of Bacillus subtilis cultures that had no obvious toxic effects on ORECs were screened using the cell counting kit-8 （CCK-8） assay to determine the ranges of proper stimulating concentration and time. After stimulating ORECs with the different concentrations of Bacillus subtilis culture supernatants that had no obvious toxic effects for 8 h, the optimal stimulating concentration for inducing SBD-1 expression was determined by using qPCR and ELISA assays. In addition, this concentration of Bacillus subtilis culture supernatant was subsequently used to stimulate ORECs for various durations, and the optimal stimulating time for inducing SBD-1 expression was determined by using qPCR and ELISA assays. [Result] ①The corresponding supernatants of Bacillus subtilis cultures at concentrations ranging from 10⁷ to 10¹¹ CFU/mL exhibited no obvious toxic effects on ORECs, and the range of proper stimulating time was 2-24 h. ②The best performance in inducing SBD-1 expression was observed in the corresponding supernatant of Bacillus subtilis cultures at a concentration of 10¹¹ CFU/mL. ③After treatment with the screened optimal stimulating concentration of Bacillus subtilis culture supernatant, the optimal stimulating time for inducing SBD-1 expression was determined as 24 h. [Conclusion] Bacillus subtilis culture supernatant could induce the SBD-1 expression in ORECs.

Key words: ovine ruminal epithelial cells, Bacillus subtilis culture supernatant, β-defensin-1, qPCR