Paramoebiasis of sea-farmed salmonids in Tasmania : a study of its aetiology, pathogenicity, and control

Amoebic gill disease (AGD) is the most significant disease of farmed sea-caged salmonids in Tasmania. The research reported here provides the first substantiated evidence for a Paramoeba species as the cause of this economically important disease. A total of 680 cultures of amoebae were prepared during an extensive sampling programme of diseased Atlantic salmon, resulting in 61 successfully purified and subcultured amoeba isolates. This library of amoebae comprised the protozoan genera Platyamoeba, Vannella, Flabellula, Heteroamoeba, Vexillifera, Acanthamoeba and Paramoeba. Fixed and frozen sections of gills from fish with AGD were immunostained with polyclonal antisera against the predominant genera associated with gills. Only Paramoeba was detected in large numbers on gill tissue and always in close association with gill hyperplasia, a characteristic pathognomonic of AGD. Antisera to Paramoeba were highly specific and did not cross react with other genera of gill-associated amoebae. Specificity of the antisera has enabled the development of a rapid and highly accurate immunofluorescent antibody test for the identification of clinical cases of AGD in farmed fish and is now the major screening tool of farmed Atlantic salmon in Tasmania. Despite evidence that Paramoeba is the cause of AGD, fulfillment of Koch's postulates could not be achieved when naive rainbow trout or Atlantic salmon were exposed to freshly isolated cultures of Paramoeba with limited laboratory passage. Infection could be established however in cohabitation trials when naive fish were exposed to fish with AGD. A sensitive enzyme linked immunosorbent assay was developed using Paramoeba isolated from fish with AGD. In a preliminary survey of several Atlantic salmon populations naturally exposed to Paramoeba, between 50-100% of fish had circulating serum antibody to Paramoeba; no antibody could be detected in gill mucus. The presence of anti-Paramoeba antibodies in the serum of fish exposed to Paramoeba and/or infected with AGD provided additional evidence for the role of Paramoeba in AGD. AGD is controlled by bathing fish in freshwater. The standard treatment regime for fish was validated by determining the rate of inactivation of Paramoeba in freshwater. In addition, 37 potential anti-amoebic compounds were screened for their amoebistatic and amoebicidal activity against Paramoeba using assays developed in this study to determine contact and growth inhibition effects. From these in vitro trials it was established that Paramoeba were totally inactivated by exposure to freshwater within 120 minutes. Hydrogen peroxide at 100ppm inactivated Paramoeba within 30 minutes as did exposure to 0.lppm ozone for four hours. Of the remaining compounds tested, 8-hydroxyquinoline, chloro-iodo-hydroxyquinoline and pyrimethamine at 30µg/ml for four hours' exposure were able to inactivate Paramoeba effectively and may have potential as medicated bath treatments for AGD. These trials also identified several chemicals able to inhibit the growth of Paramoeba at concentrations of <30µg/ml and may have potential as in-feed treatments for AGD.