Transcriptome analysis reveals temperature-dependent early immune response in flounder (Paralichthys olivaceus) after Hirame novirhabdovirus (HIRRV) infection

Abstract

Hirame novirhabdovirus (HIRRV), as a highly pathogenic fish virus, is frequently prevalent in a variety of aquaculture fish in recent years, which seriously threatens the healthy development of aquaculture industry. Epidemiological studies show that the outbreak of HIRRV is obviously temperature dependent. Virus proliferation experiments in vitro and in vivo at different temperatures indicate the antiviral response of flounder is a main reason affect the replication of HIRRV. The RNA-Seq was used to analyze the different antiviral response in flounder which infected with HIRRV at different temperatures, the experiment set two temperatures of 10 °C and 20 °C. The flounder infected with HIRRV by artificial injection, and the spleens were collected at 24 h after infection. Meanwhile, the fish injected with EPC supernatant at different temperatures were set as control groups. It can obtain four pairwise comparison groups if determine a single variable, and the differentially expressed genes were further selected. The results showed that after infection with HIRRV at 10 °C and 20 °C, the differentially expressed genes in spleen were significantly enriched in inflammatory and immune-related pathways like Arachidonic acid metabolism, Cytokine-cytokine receptor interaction, Toll-like receptor (TLR) signaling pathway, RIG-I-like receptor (RLRs) signaling pathway, NOD-like receptor (NLR) signaling pathway and Cytosolic DNA-sensing pathway etc. In addition, the expression of phagocytes, lysosomes, endocytosis related genes were significantly upregulated at high temperature whether HIRRV positive or not. But compared to the infected flounder at 10 °C, some genes of RLRs signaling pathway were significantly upregulated at 20 °C, it can be speculated that RLRs pathway may be related to the anti-HIRRV response of flounder. Therefore, key genes of RLRs signaling pathway including mda5, lgp2, mita, mavs, irf3, irf7, ifn I-3 and ifn-γ were selected, and the temporal expression patterns of these genes in infected flounder at different temperatures were further detected by qRT-PCR. The results showed that HIRRV infection can significantly stimulate and activate the RLRs pathway of flounder, and the response level of this pathway was significantly higher at 20 °C than 10 °C. In general, this study provides important data for the further study about the pathogenesis of HIRRV infection in flounder.