Dietary supplementation of Enterococcus faecalis based parabiotics combined with Bacillus subtilis ameliorate intestinal dysfunction in weaned piglets challenged with ETEC

Post-weaning diarrhea caused by enterotoxigenic Escherichia coli (ETEC) is a major challenge in pig production, necessitating development of effective alternatives to antibiotic growth promoters. This study aimed to evaluate the effects of dietary E. faecalis EF-2001 based parabiotics, supplemented either alone (WB) or alongside BS, β-glucan, and mannan-oligosaccharides (FB) on intestinal health in weaned pigs challenged with ETEC. A total of 24 crossbred weaning pigs were randomly assigned to six treatment groups with four replicates per treatment: positive control (PC, basal diet without challenge), negative control (NC, basal diet with ETEC challenge), WB1 (NC with 0.10% WB alone), FB50 (NC with 0.05% FB), FB75 (NC with 0.075% FB), and FB100 (NC with 0.10% FB). All pigs in challenged groups received oral ETEC inoculation for three consecutive days. The combined supplementation at the highest inclusion level (FB100) completely restored average daily gain and feed efficiency to levels comparable with PC, while markedly reducing diarrhea scores compared to NC. Apparent ileal digestibility of dry matter, crude protein, and gross energy improved progressively with increasing combined supplementation levels. The FB100 group maintained white blood cell counts and pro-inflammatory cytokine levels (IL-6, TNF-α) comparable to PC while increasing serum IgA and IgG concentrations compared to NC. Intestinal morphology was preserved in combined supplementation groups, with maintained villus height and reduced crypt depth resulting in improved villus height-to-crypt depth ratios. Gene expression analysis revealed upregulated claudin-1 in PC and FB100 groups, indicating enhanced intestinal barrier function. Microbiota analysis showed maintained alpha diversity with increased Blautia abundance in the FB75 group and reduced Bacteroidota in the FB100 group compared to NC. These findings demonstrate that dietary supplementation with E. faecalis-based parabiotics combined with B. subtilis, β-glucan, and mannan-oligosaccharides effectively protects against ETEC-induced intestinal dysfunction through integrated mechanisms including barrier enhancement, immune modulation, and microbiota optimization, offering a scientifically validated alternative to antibiotic growth promoters in weaned pig production.