Administration of recombinant attachment protein (r22C03) of Neoparamoeba perurans induces humoral immune response against the parasite in Atlantic salmon (Salmo salar).

This study investigated the use of a recombinant protein of Neoparamoeba perurans, the causative agent of Amoebic gill disease (AGD), as an immunogen to generate systemic and mucosal antibody responses against the parasite. Genes encoding N. perurans homologs of mannose-binding protein (MBP) from Acanthamoeba spp. have been identified. From these, a Neoparamoeba MBP - like EST has been identified and produced as a recombinant fusion protein. Attachment of N. perurans to the gill might be reduced by antibody-mediated interference of this protein, but this is dependent on the presence and level of functional antibodies in the mucus. Fish were immunized with the protein via i.p. injection with Freund's complete adjuvant (FCA); and serum and skin mucus samples were collected before and after immunization. Antibodies (IgM) present in samples were characterized via Western blot and their levels measured with an ELISA. The immunization was able to induce a systemic IgM response 8 weeks after primary exposure and a mucosal response 4 weeks post initial immunization, which were specific to the recombinant protein but not to antigens obtained from crude amoebic preparations. However, adherence of the antibodies to the parasite was observed using immunocytochemistry, and both, serum and skin mucus IgM, were able to bind the surface of formalin-fixed N. perurans. This finding may contribute to further research into the development of a vaccine for AGD.

Effect of immunization route on mucosal and systemic immune response in Atlantic salmon (Salmo salar).

This study aimed to assess systemic and mucosal immune responses of Atlantic salmon (Salmo salar) exposed to two protein-hapten antigens - dinitrophenol (DNP) and fluorescein isothiocyanate (FITC) each conjugated with keyhole limpet haemocyanin (KLH) - administered using different delivery strategies. Fish were exposed to the antigens through different routes, and were given a booster 4 weeks post initial exposure. Both systemic and mucosal antibody responses were measured for a period of 12 weeks using an enzyme-linked immunosorbent assay (ELISA). Only fish exposed to both antigens via intraperitoneal (IP) injection showed increased systemic antibody response starting 6 weeks post immunization. No treatment was able to produce a mucosal antibody response; however there was an increase in antibody levels in the tissue supernatant from skin explants obtained 12 weeks post immunization from fish injected with FITC. Western blots probed with serum and culture supernatant from skin explants showed a specific response against the antigens. In conclusion, IP injection of hapten-antigen in Atlantic salmon was the best delivery route for inducing an antibody response against these antigens in this species. Even though IP injection did not induce an increase in antibody levels in the skin mucus, there was an increased systemic antibody response and an apparent increase of antibody production in mucosal tissues as demonstrated by the increased level of specific antibody levels in supernatants from the tissue explants.