An immersion challenge model for type II grass carp reovirus (GCRV-II) induces effective infection and activates...

Grass carp is the most produced freshwater aquaculture species in China. However, high-density farming is often accompanied by severe disease outbreaks, particularly grass carp hemorrhagic disease caused by grass carp reovirus (GCRV), resulting in significant economic losses to the industry. To support disease research and control, we developed an immersion challenge model in which fish were exposed to type II grass carp reovirus (4.23 × 10³ copies/μL) for 30 min under controlled conditions (28 °C, dissolved oxygen > 6 mg/L). Across multiple independent immersion challenges, this method resulted in infection mortality rates ranging from 70% to 86%, and consistently induces typical symptoms of grass carp hemorrhagic disease, including hemorrhage at the fin base and body surface, hemosiderin deposition in the spleen, and intestinal damage. Transcriptome sequencing of spleen collected at 0, 12, 36, 72, and 120 h post-infection (hpi) yielded high-quality data from 15 samples. Compared to the 0 hpi group, 7182, 3856, 5770, and 5123 differentially expressed genes (DEGs) were identified at 12, 36, 72, and 120 hpi, respectively. These DEGs were significantly enriched in pathways related to innate and adaptive immunity, such as inflammatory responses, immunoglobulin receptor binding, and Toll-like receptor signaling pathway. Further analysis revealed significant upregulation of key immune gene families, including TLRs, RLRs, IFNs, IRFs, SOCSs, antigen-presenting molecules, and immunoglobulins. In conclusion, the immersion challenge method established in this study effectively activates both the innate and adaptive immune system in grass carp, providing a reliable experimental model and theoretical foundation for elucidating the pathogenic mechanisms of grass carp hemorrhagic disease, advancing vaccine development, and disease-resistant breeding.