TWENTY-FIFTH ANNUAL EASTERN FISH HEALTH WORKSHOP MARCH 10-13, 2000   Evidence For A Novel Vertebrate Peroxidase In Channel Catfish (Ictalurus punctatus)     Kesavan Nair Praveen1, William Nauseef2 and Andrew Goodwin1   1Aquaculture/Fisheries Center, University of Arkansas at Pine Bluff, 1200 N University Drive, Pine Bluff, AR 71611; 2 Inflammation Program and Department of Medicine, University of Iowa, Iowa City, IA  52242     Fish leukocytes produce intracellular and extra-cellular superoxide in response to a wide variety of particulate and soluble stimuli. Secondary oxidants derived from superoxide, are critical in bactericidal activity in phagocytes of fish and other animals. Derivatives like hydrogen peroxide (H2O2) and superoxide are bactericidal only at high concentrations in extra-cellular environments. In higher vertebrates, most of the H2O2 produced will be consumed by myeloperoxidase (MPO), an enzyme contained in azurophilic granules of polymorphonuclear leukocytes (PMNs). Myeloperoxidase is a classical heme peroxidase, which uses H2O2 to oxidize many organic compounds. It also oxidizes chloride ions to strong non-radical oxidant hypochlorus acid (HOCl), which is the most powerful antibacterial substance produced by neutrophils. Little is known about the MPO/Chloride/HOCl system of bactericidal activity in fish neutrophils. We investigated the presence and properties of peroxidase in neutrophils separated from the peripheral blood and head kidney of channel catfish by differential density gradient centrifugation. Polyacrylamide gel electrophoresis under non-denaturing conditions for MPO (Gel system 8, Maurer 1971) clearly showed the presence of peroxidases in neutrophils separated from peripheral blood and head kidney. There was no noticeable peroxidase activity in any other cell populations. Catfish peroxidase had a significantly different electrophoretic mobility than human MPO and eosinophilic peroxidases, when analyzed simultaneously on the same native gel system. Catfish peroxidases are antigenically distinct from MPO as a polyclonal antibody developed against human MPO failed to recognize the catfish proteins in western blot. The chlorinating activity of phorbol myristate acetate stimulated fish PMNs was considerably weaker than that of stimulated human PMNs. Purification and spectral characterization of this enzyme need to be addressed.     Return to 25th Annual Eastern Fish Health Workshop Return to Leetown Science Center Home Page