Differential Effects of Bacillus Species-Fermented Products on Antibiotic Resistome and Virulence Factor Gene Composition in the Cecal Digesta of Broilers

This study investigated the differential effects of Bacillus subtilis- and Bacillus licheniformis-fermented products (defined as SFP and LFPs, respectively) on microbial antibiotic resistance gene (ARG) and virulence factor gene (VFG) composition in the cecal digesta of 35-day-old broilers by using metagenomic sequencing. First, 160 1-day-old unsexed Arbor Acres broiler chicks were randomly allocated to four treatment groups: basal diet (CON group); basal diet + enramycin (10 mg/kg, ENM group); basal diet + SFPs (108 colony-forming units (CFU) of B. subtilis spores/kg, SFP group); and basal diet + LFPs (108 CFU of B. licheniformis spores/kg, LFP group). Principal coordinate analysis of ARG and VFG composition indicated distinct clustering among the cecal samples of the groups. At the antibiotic resistance class level, LFP treatment increased the expression of peptide resistance genes and decreased the expression of aminocoumarin resistance genes compared with the other groups. Compared with the other groups, LFP treatment promoted bcrA, ugd, and efrB expression but suppressed parY expression, whereas SFP treatment inhibited efrA expression. The abundance of the peptide resistance gene bcrA in Lachnoclostridium species was higher in the CON and LFP groups than in the ENM and SFP groups, whereas the abundance of the peptide resistance gene rpoB2 in Bacteroides species was lower in the ENM and LFP groups than in the SFP group. No specific VFGs were regulated only by SFPs or LFPs. SFP and LFP treatment inhibited clpC expression compared with the other groups. clpC abundance in Bacteroides species was lower in the LFP group than in the CON group, whereas its abundance in Faecalibacterium species was lower in the SFP group than in the CON and ENM groups. These results demonstrated that SFPs and LFPs differentially regulate microbial ARG and VFG composition in the cecal digesta of broilers. LFP supplementation modulated more antibiotic resistance classes and ARGs than did SFP supplementation.