Effects of pre-colonization patterns of complex probiotics on chick gut microbiota development, growth performance, and blood parameters

Modern commercial poultry production can disrupt natural microbial transfer from hens to newly hatched chicks, potentially affecting early gut microbiota establishment and chick development. However, how different pre-colonization approaches shape early gut microbiota development remains unclear. This study evaluated the effects of spray and padding pre-colonization with a complex probiotic preparation containing Bacillus subtilis and Enterococcus faecium on gut microbiota, growth performance, and blood parameters in chicks. A total of 1,000 hatching eggs were allocated to control (Con), spray (Treat. S), and padding (Treat. P) groups. Probiotics were applied on incubation day 19. Gut microbiota and environmental samples were collected on days 1, 14, and 28 post-hatch. Growth performance was evaluated during the 28-day brooding period, and serum biochemical, antioxidant, immune, and inflammatory parameters were measured on day 28. Gut microbial diversity, composition, differential taxa, predicted functions, and environmental–intestinal microbial associations were analyzed. Treat. P showed higher body weight than Con ( P < 0.05). Proteobacteria dominated on day 1, whereas Firmicutes predominated on days 14 and 28, with Lactobacillus becoming the major genus. Beta diversity differed among groups on day 1 ( P = 0.02) but converged by day 28 ( P = 0.952). Treat. P was enriched in Bradyrhizobium and Lactobacillus ingluviei , whereas Helicobacter was enriched in Treat. S. Predicted butanoate, propanoate, and tricarboxylic acid cycle pathways were enriched in Treat. P. Treat. P also showed higher serum IgA and antioxidant capacity and lower pro-inflammatory cytokines ( P < 0.05). Environmental and intestinal microbial communities were associated on day 14 ( r = 0.574, P = 0.007). Padding pre-colonization was associated with higher body weight than the control and showed distinct effects on gut microbial composition, predicted functions, and blood immune and antioxidant parameters. Compared with spray pre-colonization, padding was associated with better performance in several measured outcomes. These findings suggest that the padding approach may have greater potential than spray application for influencing early gut microbiota development and chick physiological responses. Not applicable.