Establishment and characterization of a Gobiocypris rarus cell line for studying type II grass carp reovirus infection

The type II Grass Carp Reovirus (GCRV-II) causes severe hemorrhagic disease with high mortality in economically important fish species. However, existing cell culture systems exhibits neither efficient viral replication nor observable cytopathic effects. In this study, a novel cell line (GRLJ) was established utilizing cells obtained from the lower jaw of Gobiocypris rarus (rare minnow), which was characterized in terms of morphology, proliferation, chromosomes, transfection efficiency, susceptibility to GCRV-II, and innate immune response. GRLJ cells with an epithelioid phenotype infected with GCRV-II exhibited a significant viral particle in the cytoplasm, and upregulation of viral genes ( vp4 , vp56 ). High-titer infection (1.9 × 10⁴ copies/μL) caused complete lethality in rare minnow by 10 days post-infection (dpi), with evident histopathology. The calculated LD₅₀ for GRLJ-derived GCRV-II was 2.6 × 10 ³ copies/μL. The expression of immune-related genes, such as tlr3 , tbk1 , ifnα, ifn2 , p65 and pkr were significantly upregulated at 3 and 7 dpi. These results highlight the potential of GRLJ cells as a model for studying GCRV-II replication and host-pathogen interactions. • A new Gobiocypris rarus (rare minnow) lower jaw cell line (GRLJ) was established. • GRLJ cells show epithelioid morphology and high transfection efficiency. • GRLJ cells were susceptible to grass carp reovirus type II (GCRV-II). • Immune-related genes were upregulated in GRLJ cells after GCRV-II infection.