Evaluation of immunogenicity and protective efficacy of the outer membrane porin F (OprF) against Pseudomonas plecoglossicida in large yellow croaker (Larimichthys crocea).

Large yellow croaker (Larimichthys crocea) farming dominates the marine aquaculture industry in China. However, the epidemic outbreaks of visceral white nodules disease (VWND), caused by bacterial pathogen Pseudomonas plecoglossicida, have emerged as a significant concern within the large yellow croaker industry. Although vaccination is considered to be an effective method for preventing and controlling P. plecoglossicida infection, there is currently no commercially available vaccine targeting this bacterium. In the present study, the outer membrane porin F (OprF) of P. plecoglossicida was characterized and revealed a high sequence similarity with that of other Pseudomonas species. The recombinant OprF protein (rOprF) produced in Escherichia coli was then evaluated for its immunogenicity and protective role against P. plecoglossicida in large yellow croaker. The rOprF was identified to have immunogenicity by Western blot using large yellow croaker anti-P. plecoglossicida sera. Additionally, the indirect immunofluorescence assay (IIFA) provided evidence indicating the surface exposure of OprF in P. plecoglossicida. Fish vaccinated twice via intraperitoneal (IP) injection with the purified rOprF combined with commercial adjuvant ISA 763A VG exhibited a relative percent survival (RPS) of 70.60% after challenge with virulent P. plecoglossicida strain through immersion. The administration of rOprF resulted in a notable increase in specific serum antibody levels and serum lysozyme activity compared to the control groups. The immune-related genes in the spleen and head kidney of rOprF-vaccinated fish were remarkably upregulated compared with the PBS-vaccinated sham group after the P. plecoglossicida challenge. In summary, the findings of this study suggest that rOprF exhibits considerable potential in inducing a robust immune response, making it a viable candidate for vaccination against P. plecoglossicida infection in large yellow croaker.

Transcription factor EB (TFEB) participates in antiviral immune responses independent of mTORC1 in macrophage of large yellow croaker (Larimichthys crocea).

Transcription factor EB (TFEB) plays an integral role in the production of proinflammatory cytokines and chemokines in response to pathogen stimulation in mammals. However, the role of TFEB in antiviral immune responses and the potential regulatory mechanisms in fish remain poorly understood. Here, we cloned and characterized Larimichthys crocea TFEB (LcTFEB) with 524 amino acids and a typical basic helix-loop-helix-leucine zipper domain. LcTFEB could translocate into the nucleus upon starvation and had a comparatively high expression in immune tissues. Similar to the expression of antiviral immune genes, the transcriptional expression and activity of LcTFEB showed a trend of increasing and then decreasing with the prolongation of stimulation. Inhibition of LcTFEB using siRNA dramatically increased the polyinosinic-polycytidylic acid (poly (I:C))-induced interferon response and pro-inflammatory cytokines mRNA expression levels, whereas pharmacological activation and overexpression of LcTFEB exhibited the reverse effects. Mechanically, LcTFEB might promote the expression of IFNh as negative feedback to limit the virus-induced inflammatory responses. Notably, although inhibition of mTORC1 exacerbated poly (I:C)-triggered inflammatory responses, the effects of LcTFEB were independent of mTORC1. Overall, this study revealed an unidentified critical role of LcTFEB in the regulation of antiviral immune responses and promoted the understanding of TFEB in the antiviral immunity of fish macrophages.