A tetrazole-based immunomodulator confers mitochondrial protection and enhances host resistance against IHNV in rainbow trout.

Infectious hematopoietic necrosis virus (IHNV) remains a serious threat to salmonid aquaculture, largely due to its high transmissibility and the absence of effective antiviral therapies. While previous studies have focused primarily on viral suppression, few have addressed the immunological and cellular responses required for host defense. Here, we report the discovery of a tetrazole-based small molecule, THA, that exerts robust anti-IHNV effects by preserving mitochondrial homeostasis and activating innate immune signaling. Among 20 synthesized tetrazole derivatives, THA exhibited the highest antiviral activity and low cytotoxicity in EPC cells. Mechanistically, THA treatment restored mitochondrial membrane potential disrupted by IHNV infection, inhibited virus-induced nuclear damage, and significantly upregulated the MAVS-TBK1-IRF3 signaling cascade as well as downstream effectors such as IFN1 and Nrf2. In vivo, immersion exposure to THA not only improved survival of IHNV-infected rainbow trout by 33 % but also markedly reduced viral load in the liver. Moreover, prolonged pre-treatment to THA enhanced host resistance and limited horizontal transmission between cohabiting fish. Although THA's antiviral efficacy declined after extended water exposure, repeated dosing retained protective effects. Overall, this study highlights the therapeutic potential of mitochondrial-targeted immunomodulators and provides a foundation for developing tetrazole-based antiviral strategies in aquaculture.