Transcriptomic and untargeted metabolomic studies on the anticoccidial activity of eugenol in broilers

Coccidiosis, a protozoan disease caused by Eimeria parasites, significantly impacts the poultry industry. Traditional control methods involve the addition of anticoccidial drugs to feed, which has led to concerns over drug residues. Thus, the search for alternative treatments has become a research priority, with plant essential oils emerging as a promising option. In the study, we evaluated the anticoccidial effects of seven plant-derived products on Eimeria tenella using a broiler cage trial and calculated the anticoccidial index (ACI) to assess their efficacy. The results revealed that eucalyptus oil had the highest ACI (157.79), followed closely by eugenol (155.41), both nearing the 160.00 threshold. Eugenol demonstrated a lower oocyst output compared to eucalyptus oil, leading us to focus on the mechanism of eugenol's anticoccidial activity using transcriptomic and untargeted metabolomic analyses. Transcriptomic analysis of cecal tissue revealed 749 upregulated and 1057 downregulated differentially expressed genes (DEGs). The top three enriched Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were the extracellular matrix (ECM)-receptor interaction, cell adhesion molecules, and cytokine-cytokine receptor interaction, with the latter pathway showing significant expression differences in 40 genes. This suggests that eugenol modulates the immune response in broilers by regulating the expression of various cytokines. Metabolomic analysis identified 103 upregulated and 22 downregulated differential metabolites, with a high enrichment of the gut IgA production-related immune network pathway. Notably, vitamin A acid, a key metabolite in this pathway, was significantly upregulated. ELISA assays confirmed the upregulation of SIgA, a terminal product of this metabolic pathway. Additionally, several anti-inflammatory metabolites and prebiotics, such as fumaric acid, quinolinic acid, succinic acid, and d-raffinose, were significantly upregulated. These results indicate that eugenol modulates the intestinal immune network and levels of various anti-inflammatory metabolites and prebiotics, suggesting its role in anticoccidial activity through the regulation of DEGs and differential metabolites. This study demonstrates that eugenol has the potential to serve as a safe alternative or adjunct to anticoccidial drugs in poultry and deepens our understanding of its anticoccidial activity.