Simple SummaryThe supplementation of diets for broiler chickens has increased due to the increasing demand of consumers for antibiotic-free broiler products. Nevertheless, the benefits of probiotics for intestinal barrier and immune functions, as well as on growth performance in chickens, are still controversially discussed. In performing a meta-analysis, we found that dietary supplementation with probiotics of various genera/species can enforce intestinal barrier function. Moreover, our meta-regressions indicated that in pathogen-challenged birds, probiotics might effectively help reduce gut inflammation by suppressing the expression of pro-inflammatory cytokines. Probiotics further sustained the intestinal histomorphology and hence digestive and absorptive processes in challenged and non-challenged chickens.AbstractData published in the literature about the favorable effects of dietary probiotics on gut health in broiler chickens are inconsistent. To obtain a more comprehensive understanding, we conducted a meta-analysis to assess the effects of probiotics on the gut barrier and immune-related gene expression, histomorphology, and growth in chickens that were either challenged or non-challenged with pathogens. From the 54 articles published between 2012 and 2022, subsets of data, separately for non-challenged and challenged conditions, for response variables were created. The mean dietary probiotic concentrations ranged from 4.7 to 6.2 and 4.7 to 7.2 log10colony-forming unit/kg under non-challenged and challenged conditions, respectively. Probiotics increased the expression of genes for mucins and tight junction proteins in the jejunum and ileum at weeks 3 and 6. The stimulatory effect of probiotics on tight junction protein expression was partly stronger in challenged than in non-challenged birds. Meta-regressions also showed an anti-inflammatory effect of probiotics under challenged conditions by modulating the expression of cytokines. Probiotics improved villus height at certain ages in the small intestine while not influencing growth performance. Dietary metabolizable energy, crude protein, and days post-infection modified the effects of probiotics on the observed variables. Overall, meta-regressions support the beneficial effects of probiotics on gut integrity and structure in chickens.