Molecular characterization and functional analysis of IL-18 in snakehead (Channa argus) during Aeromonas schubertii and Nocardia seriolae infections.

As a proinflammatory cytokine of the interleukin-1 (IL-1) family, IL-18 plays important roles in host protection against bacterial, viral, and fungal infection. We cloned the open reading frame of snakehead (Channa argus) IL-18 (shIL-18) and found that it contained 609 base pairs and encoded 202 amino acid residues. The shIL-18 included a conserved IL-1-like family signature and two potential IL-1ß-converting enzyme cutting sites; one was conserved in all analyzed IL-18s, but the other was unique to shIL-18. Unlike other IL-18s, shIL-18 also contained a predicted signal peptide. In this study, shIL-18 was constitutively expressed in all tested tissues, and its expression was induced by Aeromonas schubertii and Nocardia seriolae in the head kidney and spleen in vivo and by lipoteichoic acid, lipopolysaccharides, and polyinosinic-polycytidylic acid in head kidney leukocytes in vitro. Moreover, recombinant shIL-18 upregulated the expression of interferon-γ, IL-1ß, and tumor necrosis factor-α1 and -α2 and promoted the proliferation of leukocytes. Taken together, these results showed that IL-18 played crucial roles in host defense against bacterial infection in fish, as it does in mammals.

Two types of TNF-α and their receptors in snakehead (Channa argus): Functions in antibacterial innate immunity.

Tumor necrosis factor-α (TNF-α) is a pluripotent mediator of pro-inflammatory and antimicrobial defense mechanisms and a regulator of lymphoid organ development. Although two types of TNF-α have been identified in several teleost species, their functions in pathogen infection remain largely unexplored, especially in pathogen clearance. Herein, we cloned and characterized two types of TNF-α, termed shTNF-α1 and shTNF-α2, and their receptors, shTNFR1 and shTNFR2, from snakehead (Channa argus). These genes were constitutively expressed in all tested tissues, and were induced by Aeromonas schubertii and Nocardia seriolae in head kidney and spleen in vivo, and by lipoteichoic acid (LTA), lipopolysaccharides (LPS), and Polyinosinic-polycytidylic acid [Poly (I:C)] in head kidney leukocytes (HKLs) in vitro. Moreover, recombinant shTNF-α1 and shTNF-α2 upregulated the expression of endogenous shTNF-α1, shTNF-α2, shTNFR1, and shTNFR2, and enhanced intracellular bactericidal activity, with shTNF-α1 having a greater effect than shTNF-α2. These findings suggest important roles of fish TNFα1, TNFα2, and their receptors in bacterial infection and pathogen clearance, and provide a new insight into their function in antibacterial innate immunity.

Membrane IgM+ plasma cells in grass carp (Ctenopharyngodon idella): Insights into the conserved evolution of IgM+ plasma cells in vertebrates.

Teleost fish are the most primitive bony vertebrates that contain B cells; thus, comparative analysis of teleost naïve/mature B cells and plasma cells can provide helpful evidence for understanding the evolution paradigms of these two B-cell subpopulations in vertebrates. In this study, we developed monoclonal antibody against grass carp IgM and identified two different IgM+ cell subsets: IgM+ lymphocytes (Lym), resembling naïve/mature B cells, and IgM+ myeloid cells (Mye), resembling plasma cells. Like plasma cells in mammals, the size of IgM+ Mye is significantly larger than that of IgM+ Lym, as revealed by flow cytometric analysis and transmission electron microscopy. The IgM+ Mye were further verified as plasma cells because they showed gene expression patterns similar with those of human plasma cells and a great capacity to secrete IgM. Like mammalian IgM+ and IgA+ plasma cells, not IgG+ plasma cells, grass carp IgM+ Mye also expressed membrane immunoglobulins, a feature conserved in IgM+ plasma cells in vertebrates. Furthermore, recombinant CD40L or IL-21 alone could induce the plasma cell generation and IgM secretion, while the combination of CD40L and IL-21 had greater effect on IgM secretion, but not on plasma cell generation. This study fills an important gap in the knowledge of plasma cells in teleost fish and provides critical insights into the conserved evolution of IgM+ plasma cells in vertebrates.

Characterization of grass carp CD40 and CD154 genes and the association between their polymorphisms and resistance to grass carp reovirus.

In bony fish, CD40 and CD154 are two very important costimulatory molecules involved in T and B cell cooperation in thymus-dependent antibody production. In the current study, we identified the cDNAs of CD40 and CD154 and analyzed their genomic structures in grass carp. Quantitative real-time PCR indicated that the CD40 and CD154 were mainly expressed in immune organs. After challenge with grass carp reovirus (GCRV), these two genes were up-regulated at 72 h in head kidney and spleen. Moreover, seven and five single nucleotide polymorphisms (SNPs) were identified in the CD40 and CD154 respectively. Statistical analysis indicated that three SNPs in the coding region of the CD40 were significantly associated with the resistance of grass carp against GCRV. These results indicated that CD40 and CD154 play important roles in the responses to GCRV in grass carp. The SNP markers in the CD40 associated with the resistance to GCRV may facilitate the disease-resistant breeding of grass carp.

Effects of high hydrostatic pressure and thermal processing on bioactive compounds, antioxidant activity, and volatile profile of mulberry juice.

The aim of this study was to investigate the effects of high hydrostatic pressure and thermal processing on microbiological quality, bioactive compounds, antioxidant activity, and volatile profile of mulberry juice. High hydrostatic pressure processing at 500 MPa for 10 min reduced the total viable count from 4.38 log cfu/ml to nondetectable level and completely inactivated yeasts and molds in raw mulberry juice, ensuring the microbiological safety as thermal processing at 85 ℃ for 15 min. High hydrostatic pressure processing maintained significantly (p < 0.05) higher contents of total phenolic, total flavonoid and resveratrol, and antioxidant activity of mulberry juice than thermal processing. The main volatile compounds of mulberry juice were aldehydes, alcohols, and ketones. High hydrostatic pressure processing enhanced the volatile compound concentrations of mulberry juice while thermal processing reduced them in comparison with the control. These results suggested that high hydrostatic pressure processing could be an alternative to conventional thermal processing for production of high-quality mulberry juice.