Phenotypic features and analysis of genes supporting probiotic action unravel underlying perspectives of Bacillus velezensis VTX9 as a potential feed additive for swine

Abstract

Purpose

To date, a total of 13 probiotic Bacillus species are considered as a Generally Recognized as Safe organism (GRAS) approved by the US Federal Food, Drug, and Cosmetic Act (FDCA), which are used for food and feed additives. However, Bacillus velezensis is not considered as a probiotic candidate in swine farming due to a lack of genetic basis of probiotic action-related traits. Therefore, the present study was undertaken to exploit the genetic basis underlying the probiotic traits of B. velezensis VTX9.

Methods

The genome sequencing of B. velezensis VTX9 was performed on a PacBio Sequel platform. The probiotic properties including biosafety, antioxidative capacity, and riboflavin and exopolysaccharide production were evaluated by using genotypic and phenotypic analysis. The secondary metabolite potentials were also predicted.

Results

Strain VTX9 isolated from swine feces proved some probiotic properties including resistance to 3 mM H₂O₂, 0.6 mM bile salt, low pH, and antipathogenic activity. The complete genome of B. velezensis VTX9 consists of a 3,985,800 bp chromosome that housed 3736 protein-coding genes and 5 plasmids with the size ranging from 7261 to 20,007 bp. Genome analysis revealed no functional genes encoding enterotoxins and transferable antibiotic resistance, which confirmed the safety of VTX9. A total of 82 genes involved in gastrointestinal stress tolerance were predicted, which has not been reported previously. The maximum production of riboflavin reached 769 ± 7.5 ng/ml in LB medium after 72 h, which was in agreement with the complete de novo riboflavin biosynthetic pathway exploited for the first time in the B. velezensis genome. Antagonistic activity against pathogenic bacteria was attributed to 10 secondary metabolites clusters. The presence of a large gene cluster involved in biosynthesis of exopolysaccharides underscored further the adhesion and biofilm-forming capabilities of VTX9 in swine intestines.

Conclusion

Our results revealed for the first time that B. velezensis VTX9 has the potential to be a probiotic candidate. The information provided here on the genome of B. velezensis VTX9 opens new opportunities for using B. velezensis as a feed additive for swine farming in the future.