In ovo co-administration of vitamins (A and D) and probiotic lactobacilli modulates immune responses in broiler chickens

There is evidence that probiotic lactobacilli, in addition to essential vitamins, such as vitamin A and D, have immunomodulatory properties that enhance immune response of neonatal chickens against infections. The present study evaluated the effects of in ovo administration of retinoic acid (RA), 25-Hydroxyvitamin D₃ (VitD), and a lactobacilli cocktail on cytokine gene expression, antibody responses and spleen cell subsets in chickens. RA (90 µmol/egg) and VitD (0.6 μg/egg) were administered in ovo, either alone or in combination with lactobacilli (10⁷ CFU/egg), at embryonic d 18. On d 5 and 10 posthatch, gene expression and cellular composition were analyzed in the bursa of Fabricius and spleen. Birds were immunized on d 14 and 21 posthatch with 2 T-dependent antigens, sheep red blood cells (SRBC) and keyhole limpet hemocyanin (KLH), to assess their antibody responses. Sera were collected from the immunized chickens on d 14, 21, 28, and 35 posthatch. The results demonstrated that lactobacilli treatment increased the number of monocyte/macrophages (KUL01⁺) and CD3⁺CD4⁺ T cells in the spleen, and enhanced serum anti-KLH IgM and IgY on d 14 postprimary immunization (P < 0.05). RA significantly increased serum IgY and IgM titers to KLH and enhanced the expression of interferon (IFN)-α, interleukin (IL)-1β, IL-6, IL-8, IL-12, IL-13, and transforming growth factor-β (TGF-β) in the bursa of Fabricius (P < 0.05). The percentage of CD3⁺CD8⁺ T cells, and monocyte/macrophages (KUL01⁺) was elevated in the spleen as well (P < 0.05). These findings reveal that prehatch administration of RA improves immunocompetency of neonatal chickens by increasing the production of cytokines that regulate innate immunity and through enhancing antibody-mediated response against T-dependent antigens.