Establishment and characterization of a cell line from Yunlong grouper (Epinephelus moara ♀ and Epinephelus lanceolatus ♂) larva and its response to infectious spleen and kidney necrosis virus infection

The Yunlong grouper is a novel hybrid species resulting from the crossbreeding of Epinephelus moara ♀ and Epinephelus lanceolatus ♂. This hybrid species exhibits a combination of desirable traits inherited from both parent species. This study established the first Yunlong grouper cell line from larvae, named Glar (Grouper larvae). The cells were cultured in L-15 medium containing 15 % serum in 24 °C and have culture more than 80 passages. Chromosome karyotype analysis showed that the chromosome number was 2n = 48, and transcriptome analysis indicated that the cell line is mainly composed of fibroblast. After cryopreservation in liquid nitrogen for 6 months, cell viability remained above 70 %. The cells can be transfected, as fluorescence was observed after transfection with both Cy3-labeled scramble siRNA and the pEGFP-N1 plasmid. Furthermore, after infection with two iridoviruses , spotted knifejaw iridovirus (SKIV-SD) and spotted seabass iridovirus (SBIV-V12), Glar cells showed significant cytopathic effect (CPE), and viral particles were observed using transmission electron microscopy . Transcriptome analysis revealed that five days post-infection with SBIV-V12, differentially expressed genes were primarily enriched in pathways related to cell replication, the cell cycle, DNA repair , the p53 signaling pathway , and the Fanconi anaemia pathway, with 20 of 21 genes in the p53 pathway showing upregulation. The p53 signaling pathway plays a crucial role in antiviral immunity . Therefore, this cell line can serve as a valuable tool for in vitro research on the prevention and control of iridovirus, as well as for studying the disease mechanisms and potential therapeutic strategies for the Yunlong grouper. • This study establishes the first Yunlong grouper cell line, Glar. • The Glar cell line is capable of efficient plasmid and siRNA transfection, enabling the expression of foreign gene. • The Glar cell line serves as a substrate in vitro for Infectious spleen and kidney necrosis virus amplification.