Use and development of fish cell lines as in vitro models for the study of intracellular and extracellular protozoan pathogens

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Fish and shellfish diseases have been a cause of concern in fisheries and aquaculture because of economic losses as well as the threat of disease to humans.  Pathogens, whether viruses, bacteria or parasites, occur naturally in the aquatic environment and are usually kept under control by the homeostatic mechanisms of living animals. When environmental conditions change due to events such as global warming, pollution, stress of crowding, exposure to foreign molecules, etc, the balancing mechanisms in place change, and the animals become susceptible to disease. Much is known about disease progression, prevention, control and treatment in mammals, but comparatively little is known about these events in fish and shellfish. This research program aims to study two model protozoan parasites: a marine amoeba, Neoparamoeba pemaquidensis, believed to be the causative agent for several diseases in marine finfish and shellfish; and the freshwater microsporidian, Heterosporis anguillarum, an intracellular parasite of eels that has also been shown to affect other species such as perch, walleye, trout, and that has been recently reported to occur in species within the Great Lakes. In vitro cell culture systems have proven successful for aiding in the growth and research of important mammalian and insect protozoan pathogens. This approach has not been exploited to its full potential with fish species due to the relative paucity of convenient fish cell lines. The objective of this proposal is to elucidate the mechanisms of pathogen and host interactions in fish using in vitro approaches with the following goals: 1. Develop relatively simple and readily accessible assay systems for studying pathogen-host cell interactions, 2. Develop accurate qualitative and quantitative means to discern early changes in disease progression using novel methodologies such as analyzing protein expression signatures in parasitic protozoa exposed fish cells, 3. Investigate temporal and physicochemical factors modulating pathogen-cell interactions and the development or prevention of virulence, 4. Compare the utility and accuracy of above approaches in the prevention, diagnosis and treatment of aquatic protozoan diseases.