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Pseudomonas plecoglossicida is an aerobic Gram-negative bacterium, which is the pathogen of “Visceral white spot disease” in large yellow croaker. P. plecoglossicida is a temperature-dependent bacterial pathogen in fish, which not only reduces the yield of large yellow croaker but also causes continuous transmission of the disease, seriously endangering the healthy development of fisheries. In this study, a mutant strain of fusA was constructed using homologous recombination technology. The results showed that knockout of P. plecoglossicida fusA significantly affected the ability of growth, adhesion, and biofilm formation. Temperature, pH, H2O2, heavy metals, and the iron-chelating agent were used to treat the wild type of P. plecoglossicida; the results showed that the expression of fusA was significantly reduced at 4°C, 12°C, and 37°C. The expression of fusA was significantly increased at pH 4 and 5. Cu2+ has a significant inducing effect on the expression of fusA, but Pb2+ has no obvious effect; the expression of fusA was significantly upregulated under different concentrations of H2O2. The expression of the fusA gene was significantly upregulated in the 0.5~4-μmol/l iron-chelating agent. The expression level of the fusA gene was significantly upregulated after the logarithmic phase. It was suggested that fusA included in the TBDR family not only was involved in the transport of ferredoxin but also played important roles in the pathogenicity and environment adaptation of P. plecoglossicida.

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Fish is considered as a supreme model for clarifying the evolution and regulatory mechanism of vertebrate immunity. However, the knowledge of distinct immune cell populations in fish is still limited, and further development of techniques advancing the identification of fish immune cell populations and their functions are required. Single cell RNA-seq (scRNA-seq) has provided a new approach for effective in-depth identification and characterization of cell subpopulations. Current approaches for scRNA-seq data analysis usually rely on comparison with a reference genome and hence are not suited for samples without any reference genome, which is currently very common in fish research. Here, we present an alternative, i.e. scRNA-seq data analysis with a full-length transcriptome as a reference, and evaluate this approach on samples from Epinephelus coioides-a teleost without any published genome. We show that it reconstructs well most of the present transcripts in the scRNA-seq data achieving a sensitivity equivalent to approaches relying on genome alignments of related species. Based on cell heterogeneity and known markers, we characterized four cell types: T cells, B cells, monocytes/macrophages (Mo/MΦ) and NCC (non-specific cytotoxic cells). Further analysis indicated the presence of two subsets of Mo/MΦ including M1 and M2 type, as well as four subsets in B cells, i.e. mature B cells, immature B cells, pre B cells and early-pre B cells. Our research will provide new clues for understanding biological characteristics, development and function of immune cell populations of teleost. Furthermore, our approach provides a reliable alternative for scRNA-seq data analysis in teleost for which no reference genome is currently available.