Genipin dismantles WSSV's metabolic support system by targeting PC and PEPCK in shrimp.

White spot syndrome virus (WSSV) is the most devastating pathogen in shrimp aquaculture, causing massive economic losses worldwide. Current control strategies are limited due to the lack of adaptive immunity in crustaceans. In this study, the antiviral mechanism of genipin, a natural compound with known anti-WSSV activity, was investigated using a dual proteomic approach in Pacific whiteleg shrimp (Litopenaeus vannamei). In vitro fluorescence analysis confirmed that genipin specifically suppressed the expression of WSSV structural proteins. Through a four-group design (control, WSSV-infected, genipin-treated, and genipin-treated infected), 715 "drug-reversed" proteins were identified, whose WSSV-induced dysregulation was significantly restored by genipin. Protein set enrichment analysis and over-representation analysis revealed that genipin globally suppresses ribosome biogenesis and central carbon metabolic pathways, including the TCA cycle--pathways critical for viral replication. A concentration-gradient experiment further revealed differential regulation: genipin dose-dependently down-regulated cytoskeletal and metabolic components while up-regulating stress-responsive and adhesion modules, forming a "core response network" (CRN). Integrative cross-dataset analysis pinpointed two metabolic gatekeepers, pyruvate carboxylase (PC) and phosphoenolpyruvate carboxykinase (PEPCK), which exhibited hallmark virus-induced up-regulation and genipin-mediated reversal. Both enzymes occupy central positions linking the CRN to core metabolic pathways, suggesting a signal-metabolic coupling mechanism. Collectively, these results establish genipin as an agent that dismantles WSSV's metabolic support system potentially through modulation of PC and PEPCK, providing a promising strategy for combating viral diseases in shrimp aquaculture.