Evaluation of filamentous bacteriophage as immunogens in Atlantic salmon.

farming industry for several years. The life cycle of salmon dictates that they are reared in fresh water for the initial part of their life with maturation in salt water before harvest. It is while maturing in sea cages that salmon are susceptible to attack by marine parasites. Farmed salmon are present in much higher densities compared to wild populations and the close proximity of fish leads to rapid spread of disease or infection which leads to reduction of output. These problems have led to the development of vaccination trials to protect fish from infection or attack by parasites[ 1,2]. We have been working on a strategy to immunise salmon against a natural ectoparasite of salmon whch accumulates in high numbers on farmed fish. This parasite, Lepeophtheirus salmonis, is a copepod with a circumpolar distribution in the northern hemisphere[3]. It is responsible for loss of production due to stress on the host fish and allows other agents into the salmon's system through lesions giving rise to secondary infections which may be lethal to the fish[4]. Trials canied out have involved injection of putative vaccines into fish which are then challenged with parasites at above natural density. Sufficient control fish are included to assess the effect of the immunogen on the life cycle, morphology and fecundity of the parasites. Antigens used in trials have been in the form of Pgalactosidase fusion proteins produced in a kgtl 1 expression system, maltose binding protein fusions and formalin fixed whole cell E.coli produced using the pMAL expression system. Recently a novel system has been proposed to use epitopes expressed on the surface of filamentous bacteriophage[5] as immunogens[6]. This technique allows screening and production of antigens simultaneously using murine monoclonal antibodies raised against L. salmonis [7]. Isolated clones are then grown, purified and rendered non-infective by ultraviolet irradiation before injection into salmon. The immune response of salmon to filamentous bacteriophage has not previously been determined. The salmon immune system resembles what would be a primary immune response in mammals, with only type IgM antibodies being detected[S]. A preliminary trial immunising with "dead" M13mp19 virions was devised to determine their immunogenic effect in fish. The results of this trial are documented in table 1. Immunisation involved I.M injection with phage in Freud's complete adjuvant. Group 1 received 5 x 1Olo phage per fish while Group 2 received 1 x 1 0 ' 0 phage per fish(1pg and 0.6pg total viral protein respectively). Immune response was measured by ELISA using 1: 10 diluted salmon anti and control sera against M13 coated PVC microtitre plates. Negative controls were obtained using sera from nonimmunised FIsh and normal sheep serum used at 1: loo0 dilution. Positive controls were sheep anti-MI3 serum at a dilution of 1:2500. Responders were taken to be fish who exhibited a 2-fold greater response than the negative control mean. JASON J COULL$, WILLIAM T MELVIN$, MARIE B