Baohuoside I Combated Cryptocaryon irritans via Dual Targeting of Parasite Apoptosis and Host Defense Enhancement

Cryptocaryon irritans Brown, 1951, a ciliated protozoan, is the pathogen of cryptocaryoniasis (white spot disease) in marine fish, causing substantial morbidity and mortality, particularly in tropical and subtropical regions. This is the first study to investigate the antiparasitic activity of baohuoside I, a natural flavonoid isolated from Epimedium brevicornu Maxim., against C. irritans. In vitro exposure to baohuoside I suppressed theront viability and tomont hatching in a dose- and time-dependent manner, inducing an apoptosis-like death in both stages, characterized by ciliary detachment, mitochondrial disruption, nuclear condensation, and extensive vacuolization, as evidenced by transmission electron microscopy and Annexin V-FITC/PI staining. Further studies demonstrated that baohuoside I elevated the intracellular Ca²⁺ and reactive oxygen species levels in tomonts, indicating Ca²⁺ overload and oxidative stress. Transcriptomic analysis of infected Larimichthys crocea skin revealed that baohuoside I upregulated immune-related genes while downregulating pro-inflammatory genes, concurrently enhancing host serum acid phosphatase activity and mitigating oxidative stress in enzyme activity assays. In vivo trials showed that oral administration of baohuoside I reduced trophont attachment and improved fish survival. It did not exhibit hemolytic activity at concentrations effective against the parasites. Collectively, these findings elucidate a multi-target mechanism of baohuoside I, highlighting its potential as an eco-friendly therapeutic agent for cryptocaryoniasis control in marine aquaculture.