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Avian influenza virus (AIV) poses a great threat to the poultry industry and public health. However commercial vaccines only provide limited immunity due to rapid virus mutation and rearrangement. Here, we developed an mRNA-lipid nanoparticle (mRNA-LNP) vaccine expressing AIV immunogenic protein hemagglutinin (HA) and also assessed its safety and immune-protection efficacy in vivo. Specifically, its safety was tested by inoculation of SPF chicken embryos and chicks, and there showed no clinical manifestations and pathological changes in both. As for the immune efficacy, the antibody titers, IFN-γ production levels, and viral loads in various organs were analyzed. The results showed that chickens in the mRNA-LNP-inoculated groups produced higher specific antibody titers compared with that in the control group by hemagglutination inhibition (HI) test. Meanwhile, the ELISpot assay demonstrated that the expression of IFN-γ was markedly induced in the mRNA-LNP group, and the viral loads in multiple organs were decreased. In addition, HE shows no obvious pathomorphological changes in the lungs of the mRNA-LNP-inoculated group. While, there was severe inflammatory cell infiltration in the DMEM-treated group instead. Taken together, the vaccine prepared in this study was safe and could trigger potent cellular and humoral immune response to defend against virus infection. [ABSTRACT FROM AUTHOR]

PRRSV and CSFV are both the main pathogens of pigs and pose great threats to the pig industry. Previous studies have shown that PRRSV infection or attenuated virus vaccination can reduce the antibody level of attenuated CSFV vaccine and even cause immune failure. The higher pro-inflammatory cytokines induced by PRRSV might play a significant role in inhibiting the proliferation of CSFV-C. However, the molecular mechanism has not been elucidated yet. Here, the effect of IL-1β, a central mediator of immune-regulating inflammatory responses, on CSFV-C proliferation was investigated, as well as the mechanisms responsible for the production of IL-1β in the PRRSV and CSFV-C co-infection systems. The results showed that co-infection could significantly increase IL-1β production both at mRNA and protein levels with the infection progressing, and the IL-1β upregulation was mainly triggered by PRRSV infection. Additional experiments indicated that IL-1β inhibited the proliferation of CSFV-C in a cell-type independent manner at the replication and release stages. Furthermore, the IL-1β production induced via the TLR4/MyD88 pathway and the downstream signaling pathways NF- κ B, ERK1/2, P38, and JNK were involved by treatment with specific inhibitors or siRNA knockdown assays. Finally, we clarified that the NLRP3 inflammasome played a meaningful role in the maturation and release of IL-1β. Together, the accumulated results provided a deeper understanding of the vaccination failure of CSFV caused by PRRSV co-infection as well as targets for the development of novel approaches for the vaccination and control of CSF. • PRRSV infection significantly increased IL-1β expression with the infection progressing in PAMs. • IL-1β inhibited the proliferation of CSFV-C at the replication and release stages. • The TLR4/MyD88/NF- κ B/MAPK signaling pathways, as well as the NLRP3 inflammasome were required for IL-1β production. [ABSTRACT FROM AUTHOR]

• PRV infection inhibited SGs formation induced by AS and DTT. • EIF2α was dephosphorylated at the early stage of virus infection. • Blocking the interaction between PP1 and GADD34 could impair PRV replication in vitro. Pseudorabies virus (PRV) is one of the most notorious pathogens in the global pig industry. During infection, viruses may evolve various strategies, such as modulating stress granules (SGs) formation, to create an optimal surroundings for viral replication. However, the interplay between PRV infection and SGs formation remains largely unknown. Here we showed that PRV infection markedly blocked SGs formation induced by sodium arsenate (AS) and DL-Dithiothreitol (DTT). Accordantly, the phosphorylation of eIF2α was markedly inhibited in PRV-infected cells, although two eIF2α kinases double-stranded RNA-activated protein kinase (PKR) and PKR-like ER kinase (PERK) were activated during PRV infection. Furthermore, we also found that the dephosphorylation of eIF2α occurred at the early stage of virus infection but without the elevated production of GADD34 and PP1. Moreover, inhibition of PP1 activity by salubrinal could counteract PRV-mediated eIF2α dephosphorylation partially and inhibit virus replication. Our results revealed that, on the one hand, PRV infection activated eIF2α kinases PKR (latter inhibited) and PERK, and on the other hand, PRV encoded-functions dephosphorylated eIF2α and inhibited SGs formation to facilitate virus replication. [ABSTRACT FROM AUTHOR]

• A PAM39 cell line that could be infected with both PRRSV and CSFV-C was established. • Infection with various PRRSV could inhibit the replication of CSFV-C in PAM39 cells. • TNF-α induced by PRRSV infection through the NF-kB signaling pathway inhibited the replication of CSFV-C in vitro. Porcine productive and respiratory syndrome virus (PRRSV) and classical swine fever virus (CSFV) both are major pathogens of swine that pose a great threat to the Chinese pig industry. It has been found that PRRSV infection can lead to vaccination failure of CSFV C strain-derived modified live vaccine (CSFV-C) by interfering with the immune responses to the latter. To investigate whether PRRSV can suppress CSFV-C replication, we created a 3D4/21-based cell line PAM39 that is susceptible to both viruses by expressing PRRSV receptors CD163 and CD169, and then investigated their interplay under the condition of either sequential or simultaneous co-infection. The most significant suppressive effect came from the sequential infection when the cells were first infected by PRRSV and then followed by CSFV-C at an interval of 6 h. In addition, this effect was independent of PRRSV strains. Mechanistically, PRRSV induced an elevated level of a subset of pro-inflammatory cytokines, especially tumor necrosis factor (TNF-α), through the nuclear factor κB (NF-κB) signaling pathway to inhibit the replication of CSFV-C in vitro. Thus, our studies provide an alternative explanation on PRRSV-induced CSFV vaccination failure, and this has an important implication in CSF vaccination and control. [ABSTRACT FROM AUTHOR]