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In mammals, CD83 is a surface marker on mature dendritic cells and vital to lymphocyte activation. In teleost, studies on the function of CD83 are very limited. In this study, we examined the potential involvement of turbot ( Scophthalmus maximus ) CD83, SmCD83, in vaccine-induced immunity. For this purpose, turbot were immunized with pORF75, a DNA vaccine against megalocytivirus, in the presence or absence of pSmCD83, a plasmid that constitutively expresses SmCD83. Immune response and protection analysis showed that the presence of pSmCD83 significantly (i) enhanced the activation of head kidney macrophages (HKM) and immune gene expression, (ii) inhibited viral replication in fish tissues following megalocytivirus challenge and increased the survival of the vaccinated fish, and (iii) stimulated production of specific serum antibody and the cytotoxicity of peripheral blood leukocytes. To further examine the effect of SmCD83, pORF75 was administered into turbot in which SmCD83 was knocked down. Subsequent analysis showed that in fish with SmCD83 knockdown, vaccine-induced HKM activation and antibody production were severely reduced, and, consistently, the protectivity of pORF75 was drastically decreased. Taken together, these results indicate for the first time that teleost CD83 is required for the induction of protective immune response by DNA vaccine. [ABSTRACT FROM AUTHOR]

Highlights: [•] Three CpG ODNs that inhibited megalocytivirus replication in flounder were selected. [•] ODN C7 was able to promote leukocyte and phagocyte activation. [•] Knockdown of toll-like receptor 9 expression blocked C7-mediated immune response. [•] C7 exhibited adjuvant property and augmented the protection of a DNA vaccine. [•] C7 enhanced the expression of vaccine-induced genes of innate and adaptive immunity. [ABSTRACT FROM AUTHOR]

Abstract: Heat shock protein (Hsp) 70 is a molecular chaperone that plays an important role in protein folding and transport. In addition, Hsp70 is also involved in regulation of innate and adaptive immune response. In this study, we examined the biological activity and the immunomodulatory property of an Hsp70 homologue, PoHsp70, from Japanese flounder (Paralichthys olivaceus). Recombinant PoHsp70 purified from Escherichia coli exhibits apparent ATPase activity; however, a mutant PoHsp70, PoHsp70M, that bears mutation of the ATPase-associated domain, was completely abolished in activity. Expression of PoHsp70 was upregulated in a time-dependent manner by vaccination of flounder with a DNA vaccine, pSia10, that expresses a Streptococcus iniae antigen, Sia10. To examine whether PoHsp70 possessed any adjuvant potential, the DNA vaccine plasmids pSia10Hsp70 and pSia10Hsp70M were constructed. pSia10Hsp70 co-expresses Sia10 and PoHsp70, while pSia10Hsp70M co-expresses Sia10 and PoHsp70M. Following vaccination of flounder, production of Sia10 plus PoHsp70 and Sia10 plus PoHsp70M was detected in pSia10Hsp70- and pSia10Hsp70M-vaccinated fish respectively. At one month post-vaccination, comparable levels of serum antibodies were detected in fish vaccinated with pSia10Hsp70, pSia10Hsp70M, and pSia10. Subsequent protection analysis showed that, following S. iniae challenge, pSia10Hsp70 induced a survival rate that was significantly higher than that induced by pSia10, while pSia10Hsp70M induced a survival rate similar to that induced by pSia10. These results indicate that PoHsp70 is an effective adjuvant and that the adjuvanticity of PoHsp70 requires the intrinsic ATPase activity. [Copyright &y& Elsevier]

Abstract: Oligodeoxynucleotides (ODNs) containing unmethylated CpG motifs in certain contexts are known to be immunostimulatory in vertebrate systems. CpG ODNs with immune effects have been identified for many fish species but, to our knowledge, not for turbot. In this study, a turbot-effective CpG ODN, ODN 205, was identified and a plasmid, pCN5, was constructed which contains the CpG motif of ODN 205. When administered into turbot via intraperitoneal (i.p.) injection, both ODN 205 and pCN5 could (i) inhibit bacterial dissemination in blood in dose and time dependent manners, and (ii) protect against lethal bacterial challenge. Immunological analyses showed that in vitro treatment with ODN 205 stimulated peripheral blood leukocyte proliferation, while i.p. injection with ODN 205 enhanced the respiratory burst activity, chemiluminescence response, and acid phosphatase activity of turbot head kidney macrophages. pCN5 treatment-induced immune responses similar to those induced by ODN 205 treatment except that pCN5 could also enhance serum bactericidal activity in a calcium-independent manner. To examine whether ODN 205 and pCN5 had any effect on specific immunity, ODN 205 and pCN5 were co-administered into turbot with a Vibrio harveyi subunit vaccine, DegQ. The results showed that pCN5, but not ODN 205, significantly increased the immunoprotective efficacy of DegQ and enhanced the production of specific serum antibodies in the vaccinated fish. Further analysis indicated that vaccination with DegQ in the presence of pCN5 upregulated the expression of the genes encoding MHC class IIα, IgM, Mx, and IL-8 receptor. Taken together, these results demonstrate that ODN 205 and pCN5 can stimulate the immune system of turbot and induce protection against bacterial challenge. In addition, pCN5 also possesses adjuvant property and can potentiate vaccine-induced specific immunity. [Copyright &y& Elsevier]

Abstract: Edwardsiella tarda is an opportunistic pathogen that can infect humans, animal, and fish. Two E. tarda antigens, Eta6 and FliC, which are homologues to an ecotin precursor and the FliC flagellin, respectively, were identified by in vivo-induced antigen technology from a pathogenic E. tarda strain isolated from diseased fish. When used as a subunit vaccine, purified recombinant Eta6 was moderately protective against lethal challenge of E. tarda in a Japanese flounder model, whereas purified recombinant FliC showed no apparent immunoprotectivity. Similarly, DNA vaccines based on eta6 and fliC in the form of plasmids pEta6 and pFliC induced, respectively, moderate and marginal protection against E. tarda infection. To improve the vaccine efficacy of eta6, a chimeric DNA vaccine, pCE6, was constructed, which encodes Eta6 fused in-frame to FliC. pCE6 was found to induce significantly higher level of protection than pEta6. Likewise, another chimeric DNA vaccine, pCE18, which expresses FliC fused to a previously identified E. tarda antigen Et18, elicited significantly stronger protective immunity than the DNA vaccine based on et18 alone. Fish immunized with pEta6 and pCE6 produced specific serum antibodies and exhibited significantly enhanced expression of the genes encoding elements that are involved in both innate and adaptive immune responses. Furthermore, the induction magnitudes of most of these genes were significantly higher in pCE6-vaccinated fish than in pEta6-vaccinated fish. [Copyright &y& Elsevier]