Oral treatment for amoebic gill disease (AGD) in Atlantic salmon, Salmo salar

Neoparamoeba spp. is a marine amphizoic protozoan parasite which infects the gills of marine cultured Atlantic salmon, Salmo salar, worldwide causing amoebic gill disease (AGD). Amoebic gill disease is a significant health issue affecting the production of sea-caged Atlantic salmon in Australia with farms experiencing outbreaks regularly throughout the year. It accounts for 10-20% of the gross value of production due not only to the cost of treating and managing the disease, but also to loss of fish condition, increased feed conversion ratio (FCR), lost growth and sometimes mortalities. The current mitigation strategy for AGD is the administration of a freshwater bath to affected sea-caged fish. However, this method is becoming less effective with an apparent increase in bathing frequency over the past few years. The increase in baths has fuelled a rise in already high production costs to the Australian Atlantic salmon industry to approximately 20% annually. This thesis aims to identify an improved method of treatment for AGD either through development of a stand alone in-feed treatment or an in-feed treatment used in conjunction with the current freshwater treatment strategy. This thesis investigates in vitro and in vivo effects of bithionol and bithionol sulphoxide on both Neoparamoeba spp. and Atlantic salmon. Initially, toxicity to Neoparamoeba spp. was examined in vitro using isolated gill amoeba and exposing them to seawater, freshwater, alumina (10 mg L-1), bithionol and bithionol sulphoxide at 10, 5, 1, 0.5 and 0.1 mg L. The assays were observed for 72 h with viable amoeba counts using trypan blue exclusion conducted at 0, 24, 48 and 72 h. Both bithionol and bithionol sulphoxide were toxic to Neoparamoeba spp. in vitro at all concentrations examined. A similar toxicity to freshwater water was observed with bithionol and bithionol sulphoxide at 10 and 5 mg L-1 following a 72 h treatment. However, freshwater was the most effective with only 6% viable amoebae seen after 24 h and no viable amoeba observed a further 24 h later. Once identified as toxic to Neoparamoeba spp. in vitro, an assessment of the toxicity of bithionol to Atlantic salmon and the efficacy as an AGD treatment was evaluated. This was conducted via a bath treatment to Atlantic salmon and rainbow trout, Oncorhynchus mykiss, held in either fresh or seawater using concentrations between 1 and 35 mg L-1 to examine toxicity. To examine efficacy, a bath treatment of AGD-affected Atlantic salmon and rainbow trout at 1 to 25 mg L -1 was also evaluated. To examine toxicity, fish were bathed for 1, 3 and 6 h in bithionol, an anti-protozoal at 0, 1, 5, 10, 25 and 35 mg L -1 , with toxicity determined by time to morbidity and histological examination of internal organs. Efficacy was examined by bathing AGD-affected Atlantic salmon and rainbow trout for I h at bithionol concentrations of 1 to 25 mg L -1 . Efficacy was determined by examining gill amoeba counts and identifying percent lesioned gill filaments at I and 24 h after bath exposure to bithionol. Only bithionol at 1 mg L-1 was considered non-toxic with no signs of morbidity. Bithionol appeared to be more toxic in seawater than freshwater, exhibiting a higher rate of morbidity, and had no acute effects on gill Na+/K+ ATPase and succinic dehydrogenase, or plasma osmolality and chloride concentration. Bithionol reduced the percentage of lesioned gill filaments to the same level as freshwater. Bithionol was examined as an in-feed treatment for AGD with and without the administration of a freshwater bath. Bithionol when fed as a two week prophylactic or therapeutic treatment at 25 mg kg' feed delayed the onset of AGD pathology and reduced the percent lesioned gill filaments. Administration of a 3 h freshwater bath at 28 days post-exposure significantly reduced amoebae numbers to a similar level across all treatments; in contrast gross gill score and percent lesioned filaments were reduced proportionally. Hence, the control was significantly higher than both bithionol treatments. Following the freshwater bath, clinical signs of AGD recurred at a similar level across all treatments although controls clinical signs were significantly higher than the bithionol treatments to begin with. Palatability was not a problem with mean feed intake of bithionol over the trial duration higher compared to both the oil and plain controls. This thesis has identified that bithionol at 25 mg kg-I feed, when fed as a two week prophylactic or a therapeutic treatment, delayed and reduced the intensity of AGD pathology. Such findings as the identification of bithionol as a possible infeed treatment for AGD and its effectiveness against numerous other parasites suggests that bithionol could be worth examining in other aquatic animal diseases. Furthermore, bithionol warrants further investigation as a potential in-feed treatment for AGD in Atlantic salmon especially in regards to a combination therapy with the current freshwater mitigation.