The safety and longevity of DNA vaccines for fish

A plasmid that contained the cytomegalovirus (CMV)‐promoter‐driven lacZ reporter gene (pCMV‐lacZ) remained in the epaxial muscle of five of eight goldfish as covalently closed circles, the most functional form of plasmid, for at least 70 days at 22°. It was not present in the gills or elsewhere by polymerase chain reaction and was not integrated. Its expressed protein, Escherichia coliβ‐galactosidase (β‐gal), which was in the injected myofibres, was detected in all the fish at 4–21 days and in about half the fish from 28 days until the end of the experiment at 70 days. The numbers of cells that secreted antibody to β‐gal in the kidney peaked at 14 days. Serum antibody and proliferating kidney cells to β‐gal were in all fish from 14 days with a plateau of the responses from 21 days onwards. The plasmid did not induce autoimmune‐like antibodies to itself or to single‐ or double‐stranded salmon testis DNA. Plasmids can therefore induce long‐term foreign protein expression whilst inducing humoral and cell‐mediated immunity without autoimmunity or integration in goldfish.