A promising strategy to battle Enterocytozoon hepatopenaei infection in shrimp: dual action of paeonol on spore inhibition and immune activation

Enterocytozoon hepatopenaei (EHP) infection poses a significant challenge to shrimp aquaculture because the substantial economic losses it currently causes and the lack of treatment methods . In this study, ten individual compounds were evaluated for their effectiveness in whiteleg shrimp ( Penaeus vannamei ) against EHP. Among these compounds, paeonol exhibited the strongest inhibitory effect, reducing the EHP copy number by 82.2% at 10 mg/L. Both prophylactic and therapeutic administration of paeonol significantly reduced EHP copy number and enhanced shrimp weight gain. A combined prophylactic-therapeutic regimen resulted in superior outcomes compared with either strategy alone. Shrimp receiving a 0.1% paeonol-supplemented diet for 15 d before infection and continuing for 45 d post-infection showed a 28.7% increase in body weight compared with the untreated group. Paeonol inhibited EHP spore germination in a time-dependent manner and disrupted sporogony by decreasing nuclear number and electron-dense disc formation. Furthermore, paeonol upregulated immune-related genes in the JAK/STAT signaling pathway—including Domeless (DOME), JAK , STAT , and c-type lysozyme ( Lyz-c )—indicating an immunomodulatory role in its anti-EHP activity. Collectively, these findings demonstrate that paeonol targets two critical stages of the EHP life cycle—spore germination and developmental progression—and represents a promising candidate for integrated EHP prevention and control strategies in shrimp aquaculture.