Antiparasitic efficacy of aqueous green tea extracts and its key constituents against Cryptocaryon irritans in the Large Yellow Croaker (Larimichthys crocea)

Cryptocaryoniasis remains a major threat to marine teleost species worldwide. To develop eco-friendly prophylactic treatment and prevention strategies, this study systematically evaluated the efficacy of aqueous green tea extract (AGTE) against three Cryptocaryon irritans life stages and its therapeutic potential during C. irritans infections of cultured large yellow croakers (Larimichthys crocea). In vitro parasiticidal assays demonstrated a significant positive correlation between AGTE concentration, treatment duration, and mortality rates across the three parasite life stages. Notably, at 50 mg/L, AGTE achieved a stage-specific efficacy of >80 % theront mortality within 2 h, 55.56 % protomont mortality within 4 h, and 61.11 % tomont mortality within 12 h. Increasing the AGTE concentration to 75 mg/L enhanced efficacy to 95 % theront mortality within 1 h, 52.78 % protomont mortality within 2 h, and 100 % tomont mortality within 12 h. Ultrastructural analysis revealed significant changes in tomonts following 12 h exposure to 75 mg/L AGTE, including disrupted internal organization, expanded periplasmic space between the cyst wall and cytoplasm, reduced mucocyst counts, and the formation of pathological vacuoles. Transmission electron microscopy further showed mitochondrial degeneration in the 75 mg/L AGTE exposed tomonts characterized by disrupted outer membranes, disorganized cristae, and matrix dissolution with vacuolation. In vivo challenge trials showed complete mortality (100 %) in L. crocea in untreated controls by day nine (relative tomont number [RTN] = 85.5), whereas AGTE-treated groups exhibited dose-dependent protection: 25 mg/L (6.67 % survival), 75 mg/L (76.67 %), and 150 mg/L (60.0 %) at trial termination (day 14). Notably, tomont elimination occurred in the 75 mg/L and 150 mg/L groups, while residual tomonts persisted in 25 mg/L treatment tanks (RTN = 7.5 at day 14). Bioactivity-guided characterization identified epigallocatechin (EGC) as the predominant catechin in AGTE, demonstrating complete tomont eradication at 40 mg/L following 12 h of immersion. Comparative analysis revealed that gallic acid (GA) required double the concentration of EGC (80 mg/L) to achieve equivalent efficacy.