Bacillus Subtilis Supplemented Diet Improves Weight Gain and Caecal Luminal Microbiota in Meat Chickens

Necrotic Enteritis (NE) outbreaks cause an estimated US$6 billion financial loss per annum to the global poultry industry (Wade & Keyburn, 2015). With discontinuation of antibiotic growth promoters (AGPs) in Europe and implicit withdrawal of AGPs worldwide, a global research effort has been made to find alternative solutions to minimise diseases in animal production including NE. The current study investigated the benefits of a Bacillus subtilis derived probiotic (Bs29784) in NE challenged broiler chickens. The study used 312 male day-old Ross 308 in floor pens in separated blocks of the same room from d 0 to d 35. The treatments groups included: two control diets without or with NE challenge (CtrlNC and CtrlNE, respectively); two treatment diets of probiotic and antibiotic supplements both with NE challenge (ProNE and AntNE). Diets comprised standard wheat and soybean-meal based control diet for CtrlNC and CtrlNE, and addition of respective additives for either ProNE (Bs29784 500 mg/kg) diet or AntNE (Zn Bacitracin 4.4 mg/kg) diet. NE challenge procedures commenced with Eimeria spp. 1 mL/bird per os at d 9 with a suspension of 5000 sporulated oocysts of E. acervulina, and E.maxima, and 2500 sporulated oocycts of E. brunetti and Clostridium perfringens EHE-NE18 (approx. 10⁸ CFU/mL) and 1 ml/bird per os at d 14 and d 15 (Wu et al., 2010). Performance was measured, and samples for 16S next-generation sequencing (NGS) in caecal content and gene expression analysis from jejunal tissue were collected on d 16 and/or d 35. Data analysis applied a one-way ANOVA and significance at P < 0.05 with Tukey`s test.

NE challenge (CtrlNE) significantly suppressed the performance parameters (27% weight gain reduction, 13 points FCR increase at d 16, and 12% weight gain reduction, 6 points FCR increase at d 35) compared to the CtrlNC. By d 35, the probiotic and antibiotic treatments enabled significantly higher weight gain than the CtrlNE. Caecal microbiota d 16 showed reduced OTUs counts in all the challenged groups regardless of additives, but no difference was observed at d 35. For microbial abundance analysis, Venn diagram showed that each group has their specific OTUs at d 16 (6, 3, 3 and 2 for CtrlNC, AntNE, ProNE and CtrlNE, respectively), whereas only CtrlNC showed 1 specific OTU at d 35. Principal component analysis (PCA) of OTUs abundance revealed different diversity between the respective treatment groups. The ProNE and AntNE grouped together while the CtrlNC and CtrlNE grouped separately at d 16, indicating the similarity of OTUs between the ProNE and AntNE birds that were however different from both CtrlNC and CtrlNE birds. Interestingly, the OTUs structure at d 35 changed with distinct grouping of the CtrlNC or AntNE from CtrlNE and ProNE, latter two grouped closely although minimal overlapping. This may imply recovered microbiota balance in both the CtrlNE and ProNE while the strong antimicrobial effects in AntNE induced the different microbiota structure. The taxonomic composition indicated an increase of E. coli in all challenged groups at d 16, but a balanced composition was observed among all groups at d 35 with abundance of F. prausnitzii in ProNE. Besides, ProNE treatment exhibited upregulation of the genes at d 16 (P < 0.05): CLDN1, IL12-b, MUC5ac, TGF-4B, TLR21, TLR5, JAM2 and INFy genes, suggesting enhanced immunity and intestinal integrity. It is suggested that Bs29784 may enable improved health of meat chickens under NE challenge.