Theaflavin Suppresses PEDV Replication via Inhibiting PI3K/Akt/mTOR Signaling and Directly Interacting with Viral Proteins

Porcine epidemic diarrhea virus (PEDV) causes acute diarrhea and high mortality in piglets, and conventional control strategies are often limited by frequent viral mutations. Theaflavin (TF), a natural polyphenol from black tea, has demonstrated antiviral effects against several viruses; however, its efficacy against PEDV and its underlying mechanisms remain unclear. In this study, we established a PEDV infection model in Vero and IPI-HB1 cells and confirmed that TF significantly inhibited viral replication in a dose-dependent manner, particularly during internalization and replication. Network pharmacology and molecular docking identified 12 TF-related targets, four of which showed strong binding affinities. TF treatment downregulates the mRNA expression of these targets, contributing to its antiviral effects. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses suggested that the PI3K/Akt/mTOR pathway was central to the mechanism of TF. Western blotting confirmed that TF suppressed Akt and mTOR phosphorylation, while pathway activation reversed its antiviral effects. Furthermore, TF were found to directly bind to the PEDV nucleocapsid (N) and nonstructural protein 5 (Nsp5), suggesting additional virus-targeted activity. Therefore, TF exerts dual antiviral effects against PEDV by modulating host signaling and targeting viral proteins, thus offering a promising natural compound for antiviral drug development.