Study On The Mechanism Of Inflammation Induced By Micropterus Salmoides Rhabdovirus And Its Antiviral Drugs

In China,the largemouth bass(Micropterus salmoides)is a significant freshwater economic farmed fish,with an annual farmed production of more than 600,000 tons.Diseases in farmed largemouth bass have become more prevalent in recent years,particularly Micropterus salmoides rhabdovirus(MSRV),which has caused huge economic losses to largemouth bass farming industry.This study demonstrated that inflammation mediated by NF-κB,MAPK,and PI3 K pathways was the pathogenic mechanism of MSRV infection through in vivo infection and transcriptome sequencing.This information could be used to clarify the pathogenic mechanism of MSRV infection and develop effective prevention and control strategies for MSRV.Additionally,screening broad-spectrum antiviral drugs based on the inflammation pathway that effectively resist MSRV infection could offer fresh perspectives on how to prevent and manage Micropterus salmoides rhabdovirus disease.The precise findings are as follows.1.MSRV infection mediated inflammatory responses through NF-κB,MAPK and PI3 K signaling pathways.MSRV induced inflammatory responses and increased expression of TNF α,IL-8,IL-1β,COX-2,IFN-γ and IL-10 in vivo infection and cell-level assays.Transcriptome sequencing suggested inflammation-generating signaling pathways.The results of inhibitor experiments showed that,the inhibition of the MAPK(including p38 MAPK,ERK,JNK)pathway by specific inhibitors significantly inhibited the expression of IFN-γ and IL-10 in MSRV-infected cells compared with the control group,while the expression of pro-inflammatory factors was not significantly affected,indicating that MAPK was involved in regulating the expression of IL-10 and IFN-γ in the MSRV-induced inflammatory response.PDTC,an inhibitor of NF-κB pathway,notably weakened the increased expression of TNF α,IL-8,IL-1β,IFN-γ and IL-10 induced by MSRV infection,and suppressed the NF-κB(p65)migration from cytoplasm to nucleus,indicating that the NF-κB signaling pathway is involved in the process of MSRV infection-induced inflammatory response.When the PI3 K pathway was inhibited,the expression of TNF α,IL-8,IL-1β,COX-2,IL-10 and IFN-γ and the nuclear translocation of AKT were also significantly inhibited,indicating that the PI3 K signaling pathway is involved in the process of MSRV infection-induced inflammatory response.2.Myricetin modulated NF-κB p65 signaling to exert anti-inflammatory and antiviral activity.Antiviral strategies targeting inflammatory pathways were evaluated,and broad spectrum antiviral drugs were screened.Specifically,myricetin,naringenin,andrographolide as well as resveratrol were found to significantly inhibit NF-κB promoter activity.The quantitative analysis of MSRV G protein and plaque assay determined that myricetin had good antiviral activity.At the same time,myricetin was effective in attenuating MSRV infection-induced CPE and inhibiting inflammatory factors overexpressed.The results of western blot and grayscale analysis demonstrated that myricetin significantly inhibited the phosphorylation and nucleation of the NF-κB p65 subunit.Overall,myricetin may exert its anti-inflammatory and anti-viral activities through modulation of NF-κB signaling.3.The direct anti-MSRV activity of Myricetin.In order to further investigate the best method of myricetin dosing,pretreatment,co-incubation and treatment groups were set up,and it was found that myricetin could inhibit the replication of MSRV,its antiviral effect was the best under co-incubation and significantly inhibited the CPE caused by MSRV infection.Then,the effects of myricetin on MSRV infection was detected in the adsorption of virus,and the results demonstrated that myricetin was able to significantly inhibite the adsorption of MSRV at a concentration of 100 μM and had direct inhibition of MSRV activity.