Protective activity of galacto-oligosaccharides against intestinal damage and inflammation induced by enterotoxigenic Escherichia coli F4+ and evaluation of prebiotic potential

Post-weaning diarrhea in piglets is frequently caused by enterotoxigenic <i>Escherichia coli</i> (ETEC) F4⁺. The objective of this study was to examine the possible protective effect of galacto-oligosaccharides (GOS) on ETEC F4⁺-induced intestinal injury. Growth inhibition of ETEC F4⁺ in the presence of 2% GOS was assessed, as well as the ability of GOS to reduce pathogen adhesion and invasion in the intestinal Caco-2 cell line. GOS ability to counteract ETEC F4⁺ adhesion was also assessed <i>ex vivo</i> in piglet small intestinal villi. Protective activity of GOS against ETEC F4⁺-induced membrane damage in Caco-2 cells was evaluated through transepithelial electrical resistance (TEER), phenol red apparent permeability (Papp) and immunolocalization of tight junction proteins occludin and ZO-1. Inflammation was assessed by quantification and immunolocalization of phosphorylated-p65 protein, indicative of NF-κB activation. Finally, GOS prebiotic activity on probiotic strains <i>Lactobacillus amylovorus</i>: ATCC 33198 and DSM 16698, as well as <i>Limosilactobacillus reuteri</i> subsp. <i>porcinus</i> DSM 110571, was also investigated. The results showed that GOS significantly reduced ETEC F4⁺ adhesion and invasion in Caco-2 cells, as well as adhesion to piglet intestinal villi. Furthermore, GOS markedly decreased ETEC F4⁺ induced membrane damage, as evidenced by improvement of TEER, phenol red Papp and tight junction protein immunolocalization. A reduction of p65 phosphorylation and nuclear translocation in the presence of GOS indicated a diminished activation of the NF-κB pathway. GOS also exhibited promising prebiotic activity toward the tested probiotic strains. Taken together, these <i>in vitro</i> findings suggest the potential of including GOS in piglet weaner diets to prevent ETEC F4⁺-induced intestinal injury.