Transcriptomic Profiling of Porcine Duodenal, Jejunal, and Ileal Organoids in Response to Porcine Epidemic Diarrhea Virus

Porcine epidemic diarrhea virus (PEDV) is a highly pathogenic virus that causes severe gastrointestinal disease in neonatal piglets, often leading to high mortality. To better understand PEDV pathogenesis, we developed porcine intestinal apical-out organoids derived from the duodenum, jejunum, and ileum that support viral replication and enable long-term experimental manipulation. In this study, we investigated the region-specific responses of these organoids to PEDV infection, focusing on regional characteristics, gene expression, and susceptibility to infection. PEDV replicated efficiently in the apical-out organoids, with significantly higher viral loads in jejunal and ileal organoids than duodenal organoids, indicating region-specific susceptibility. Bulk RNA sequencing and a differential gene expression analysis revealed unique transcriptomic responses across regions. The jejunal and ileal organoids exhibited stronger activation of pathways related to cellular processes, immune regulation, and antiviral defense than the duodenal organoids. Notably, viral entry receptor genes such as <italic>ANPEP</italic>, <italic>ACE2</italic>, and <italic>DPP4</italic> were expressed at higher levels in jejunal and ileal organoids under uninfected conditions, suggesting an innate predisposition for viral entry in these regions. Further analysis identified key upregulated genes involved in immune modulation, inflammation regulation, and tissue integrity, such as <italic>SLIT2</italic>, <italic>MMD2</italic>, and <italic>PKHD1</italic>, along with downregulated genes, including <italic>IL-1A</italic>, <italic>MMP13</italic>, and <italic>GNA15</italic>, that help control inflammation and minimize tissue damage. In conclusion, PEDV infection in porcine intestinal organoids elicits region-specific responses, with increased susceptibility and antiviral activation in jejunal and ileal organoids driven by the differential expression of viral entry receptors and immune-regulatory genes.