In vitro antiviral activity of the RdRp inhibitor 4'-fluorouridine against a porcine teschovirus clinical isolate.

Porcine Teschovirus (PTV) is an economically significant swine pathogen with a global distribution that can cause severe neurological, respiratory, and reproductive diseases. However, no specific vaccines or antiviral drugs are currently available. To address the absence of specific antiviral therapies for PTV, this study aimed to identify a clinically promising antiviral candidate through in vitro compound screening. We began by isolating and purified a contemporary PTV strain (PTV/Shandong/2024) from a clinical swab of a dead piglet. We also established a strain-specific RT-qPCR assay to accurately quantify PTV RNA load. A subsequent targeted screen of a hepatitis C virus (HCV) inhibitor library identified the nucleoside analog 4'-fluorouridine (4'-FLU) as a potent inhibitor of PTV replication in vitro. Treatment with 4'-FLU significantly reduced viral load and cytopathic effects in infected PK-15 cells. The compound exhibited potent antiviral activity, with a half maximal inhibitory concentration (IC₅₀) of 1.234 µM, and low cytotoxicity, with a half maximal cytotoxic concentration (CC₅₀) of 295.7 µM. This resulted in a high selectivity index of 239.6, indicating a wide therapeutic window. Time-of-addition experiments revealed that 4'-FLU acts primarily during the early stages of viral replication. These results provide the first evidence that 4'-FLU suppresses PTV replication in vitro supporting its further evaluation as a candidate antiviral against PTV.