Cepharanthine suppresses LMBV replication via cell cycle arrest and immunomodulation

In recent years, the viral disease caused by largemouth bass virus (LMBV) has inflicted severe economic losses on China's largemouth bass (Micropterus salmoides) aquaculture, significantly restricting its development. As targeted antiviral drugs and commercial vaccines remain in the investigational stage, the discovery of novel antiviral agents to control LMBV infections is urgently needed. Consequently, this study systematically evaluated the antiviral efficacy and underlying immunopharmacological mechanisms of cepharanthine (CE), a natural bisbenzylisoquinoline alkaloid, against LMBV infection using integrated in vitro and in vivo models. In vitro, CE exhibited potent anti-LMBV activity in epithelioma papulosum cyprinid (EPC) cells, with a half-maximal inhibitory concentration (IC₅₀) of 0.2136 μM and negligible cytotoxicity within the effective concentration range. Furthermore, CE treatment induced a significant, dose-dependent reduction in progeny viral titers (TCID₅₀). Additionally, CE suppressed intracellular viral replication by inducing G0/G1 phase arrest and apoptosis, effectively blocking viral DNA synthesis and prematurely eliminating infected cells. In vivo, CE administration (40 mg/kg) significantly protected LMBV-infected largemouth bass, reducing cumulative mortality from 73.33% to 46.67%. Concurrently, CE alleviated histopathological damage, diminished viral loads in primary target organs (liver, spleen, and kidney), and dynamically modulated the mRNA expression levels of key immune-related genes (IL-1β, IL-6, TNF-α, and IFN-γ). Furthermore, transcriptomic profiling revealed that CE globally reversed LMBV-induced gene expression patterns, down-regulating core replication, cell cycle, and apoptotic pathways.Collectively, our findings demonstrate that CE is a promising multi-functional antiviral agent for the clinical management of ranavirus infections in aquaculture.