Effects of Dietary Chitosan oligosaccharides Supplementation on Th17/Treg Balance and Gut Microbiota of Early Weaned Pigeon Squabs

Our previous study found that early weaning is associated with decreased growth performance, intestinal barrier impairment, and an imbalance in Th17/Treg in pigeon squabs. Chitosan oligosaccharides (COS) has been substantiated to regulate gut microbiota and restore Th17/Treg equilibrium in mammals, thereby ameliorating growth performance. However, the potential effects of COS in altricial birds remain unclear. 300 healthy 7-day-old American king pigeon squabs were selected with similar body weights and randomly divided into 5 groups. The 5 treatment groups were as follows: the control group (CON), fed with artificial pigeon milk; four supplementation groups, fed with artificial pigeon milk +100 (COS1), 150 (COS2), 200 (COS3), and 250 (COS4) mg/kg COS, respectively. Results showed that dietary supplementation of COS significantly enhanced the growth performance of weaned squabs. Compared to the CON group, the COS groups exhibited increased villus length and villus area in the jejunum and ileum, accompanied by improvements in morphological structure and mucosal permeability. COS was found to reduce the levels of Th17-associated cytokines and increase the levels of Treg-associated cytokines. COS downregulated the expression of retinoic acid receptor-related orphan receptor C (RORC), a key transcription factor of Th17 cells, while upregulated the expression of Forkhead box protein P3 (FOXP3), a key transcription factor of Treg cells. Dietary COS supplementation increased gut bacterial diversity, altered the relative abundance of several bacteria taxa and enhanced the concentration of short-chain fatty acids (SCFAs). Correlation analysis demonstrated a close association between gut microbiota, SCFAs, and indicators related to the Th17/Treg balance. Moreover, we found that SCFAs correlated more strongly with Th17/Treg-related indexes than gut microbiota. These results demonstrated that COS could relieve early weaning stress in pigeon squabs and the optimal dosage of dietary COS supplementation was suggested to be 200mg/kg. In addition, COS had a protective effect on maintaining intestinal immune balance by modulating microbiota and Th17/Treg related signaling pathways, in which SCFAs might play a crucial role as messengers.