Mb Adams

Preliminary success using hydrogen peroxide to treat Atlantic salmon Salmo salar L. affected with experimentally induced amoebic gill disease (AGD)

Currently, the only effective and commercially used treatment for amoebic gill disease (AGD) in farmed Tasmanian Atlantic salmon is freshwater bathing. Hydrogen peroxide (H₂O₂), commonly used throughout the aquaculture industry for a range of topical skin and gill infections, was trialled in vitro and in vivo to ascertain its potential as an alternative treatment against AGD. Under in vitro conditions, trophozoites of Neoparamoeba perurans were exposed to three concentrations of H₂O₂ in sea water (500, 1000 and 1500 mg L⁻¹) over four durations (10, 20, 30 and 60 min) each at two temperatures (12 and 18 °C). Trophozoite viability was assessed immediately post-exposure and after 24 h. A concentration/duration combination of 1000 mg L⁻¹ for >10 min demonstrated potent amoebicidal activity. Subsequently, Atlantic salmon mildly affected with experimentally induced AGD were treated with H₂O₂ at 12 and 18 °C for 15 min at 1250 mg L⁻¹ and their re-infection rate was compared to freshwater-treated fish over 21 days. Significant differences in the percentage of filaments affected with hyperplastic lesions (in association with amoebae) and plasma osmolality were noted between treatment groups immediately post-bath. However, the results were largely equivocal in terms of disease resolution over a 3-week period following treatment. These data suggest that H₂O₂ treatment in sea water successfully ameliorated a clinically light case of AGD under laboratory conditions.

Host response to the chondracanthid copepod Chondracanthus goldsmidi, a gill parasite of the striped trumpeter, Latris lineata (Forster), in Tasmania

The chondracanthid copepod, Chondracanthus goldsmidi is an ectoparasite of gills, inner opercula and nasal cavities of cultured striped trumpeter, Latris lineata (Forster). Whilst often present in high numbers (up to 60 parasites per host), little is known about its effect on striped trumpeter. In this study C. goldsmidi was associated with extensive epithelial hyperplasia and necrosis. Pathological changes were most pronounced near the parasites attachment site, with papilloma-like growths surrounding the entire parasite resulting in deformation of the filament. The number of mucous cells increased near the parasite attachment sites on both the opercula and gills. Mast cells were absent in healthy gills; in contrast numerous mast cells were identified in the papilloma-like growths. Immunostaining identified piscidin-positive mast cells in the papilloma-like growths, presenting the first evidence of piscidin in the family Latridae.

A phylogeny for the genus Isoodon and a range extension for I. obesulus peninsulae based on mtDNA control region and morphology

Short-nosed bandicoots, Isoodon, have undergone marked range contractions since European colonisation of Australia and are currently divided into many subspecies, the validity of which is debated. Discriminant function analysis of morphology and a phylogeny of Isoodon based on mtDNA control region sequences indicate a clear split between two of the three recognised species, I. macrourus and I. obesulus/auratus. However, while all previously recognised taxa within the I. obesulus/auratus group are morphologically distinct, I. auratus and I. obesulus are not phylogenetically distinct for mtDNA. The genetic divergence between I. obesulus and I. auratus (2.6%) is similar to that found among geographic isolates of the former (I. o. obesulus and I. o. peninsulae: 2.7%). Further, the divergence between geographically close populations of two different species (I. o. obesulus from Western Australia and I. a. barrowensis: 1.2%) is smaller than that among subspecies within I. auratus (I. a. barrowensis and I. auratus from northern Western Australia: 1.7%). A newly discovered population of Isoodon in the Lamb Range, far north Queensland, sympatric with a population of I. m. torosus, is shown to represent a range extension of I. o. peninsulae (350 km). It seems plausible that what is currently considered as two species, I. obesulus and I. auratus, was once one continuous species now represented by isolated populations that have diverged morphologically as a consequence of adaptation to the diverse environments that occur throughout their range. The taxonomy of these populations is discussed in relation to their morphological distinctiveness and genetic similarity.

Genetic variation within the dasyurid marsupial genus Planigale

Genetic variation within the genus Planigale was examined through analyses of 12S rRNA gene sequences and allozymes. The level of genetic divergence between the five currently recognised Planigale species was compared and the magnitude of divergence among populations assessed. This examination of molecular variation within the genus revealed that Planigale contains far more taxonomic diversity than is currently recognised. Specifically, the Pilbara region of Western Australia probably contains two currently unrecognised Planigale species and there is substantial genetic heterogeneity within the widespread species P. maculata. Ambiguity over the status of several genetic and/or morphological forms within the genus indicates that further taxonomic changes are likely to be warranted in the future. This study also demonstrates that the currently accepted geographic ranges of many planigale species require careful re-evaluation and that many specimens in collections are apparently misidentified. This is especially true of P. ingrami, which appears to have a much greater range than is currently recognised, being present in South Australia.

Characterisation of an immunodominant, high molecular weight glycoprotein on the surface of infectious Neoparamoeba spp., causative agent of amoebic gill disease (AGD) in Atlantic salmon.

Amoebic gill disease can be experimentally induced by the exposure of salmonids to Neoparamoeba spp. freshly isolated from infected fish, while cultured amoebae are non-infective. Results from our previous work suggested that one key difference between infectious and non-infectious Neoparamoeba were the highly glycosylated molecules in the glycocalyx. To characterise these surface glycans or glycoproteins we used a monoclonal antibody (mAb 44C12) specific to a surface molecule unique to infective parasites. This mAb recognised a carbohydrate epitope on a high molecular weight antigen (HMWA) that make up 15-19% of the total protein in a soluble extract of infectious parasites. The HMWA consisted of at least four glycoprotein subunits of molecular weight (MW) greater than 150 kDa that form disulfide-linked complexes of MW greater than 600 kDa. Chemical deglycosylation yielded at least four protein bands of approximate MW 46, 34, 28 and 18 kDA. While a similar HMWA complex was present in non-infective parasites, the glycoprotein subunits were of lower MW and exhibited differences in glycosylation. The four glycoproteins subunits recognised by mAb 44C12 were resistant to degradation by PNGase F, PNGase A, O-glycosidase plus β-1, 4-galactosidase, β-N-acetylglucosaminidase and neuraminidase. The major monosaccharides in the HMWA from infectious parasites were rhamnose, fucose, galactose, and mannose while sialic acids were absent. The carbohydrate portion constituted more than 90% of the total weight of the HMWA from infectious Neoparamoeba spp. Preliminary results indicate that immunisation of salmon with HMWA does not lead to protection against challenge infection; rather it may even have an immunosuppressive effect.

Skeletal anomaly assessment in diploid and triploid juvenile Atlantic salmon ( Salmo salar L.) and the effect of temperature in freshwater

Triploid Atlantic salmon tend to develop a higher prevalence of skeletal anomalies. This tendency may be exacerbated by an inadequate rearing temperature. Early juvenile all-female diploid and triploid Atlantic salmon were screened for skeletal anomalies in consecutive experiments to include two size ranges: the first tested the effect of ploidy (0.2-8 g) and the second the effect of ploidy, temperature (14 °C and 18 °C) and their interaction (8-60 g). The first experiment showed that ploidy had no effect on skeletal anomaly prevalence. A high prevalence of opercular shortening was observed (average prevalence in both ploidies 85.8%) and short lower jaws were common (highest prevalence observed 11.3%). In the second experiment, ploidy, but not temperature, affected the prevalence of short lower jaw (diploids > triploids) and lower jaw deformity (triploids > diploids, highest prevalence observed 11.1% triploids and 2.7% diploids) with a trend indicating a possible developmental link between the two jaw anomalies in triploids. A radiological assessment (n = 240 individuals) showed that at both temperatures triploids had a significantly (P < 0.05) lower number of vertebrae and higher prevalence of deformed individuals. These findings (second experiment) suggest ploidy was more influential than temperature in this study.

An opportunistic detection of amoebic gill disease in blue warehou, Seriolella brama Günther, collected from an Atlantic salmon, Salmo salar L., production cage in south eastern Tasmania.

Amoebic gill disease (AGD) is a recurrent health issue affecting the commercial culture of Atlantic salmon, Salmo salar L., in south eastern Tasmania

Changes in the interbranchial lymphoid tissue of Atlantic salmon (Salmo salar) affected by amoebic gill disease

The interbranchial lymphoid tissue (ILT) was recently described in the gills of salmonids. This study examined changes in the ILT during a parasitic infection in marine environment, using amoebic gill disease (AGD) as a model. Atlantic salmon (Salmo salar) experimentally infected with Neoparamoeba perurans were sampled at 0, 3, 7, 14 and 28 days post challenge. Transversal sections of three areas of the gills (dorsal, medial and ventral) were histologically assessed for morphological and cellular changes. AGD induced morphological changes and a cellular response in the ILT of affected fish. These changes included a significant increase in the ILT surface area in fish 28 days after AGD challenge, compared to control fish at the same time point. The length of the ILT increased significantly 28 days post exposure in the dorsal area of the gill arch in the fish affected by AGD. The lymphocyte density of the ILT increased after AGD challenge, peaking at 7 days post exposure; however, by 28 days post exposure, a reduction of lymphocyte density to values close to pre-infection levels was observed. PCNA immunostaining revealed that epithelial hyperplasia was the most likely factor contributing to the ILT enlargement in the affected fish.

In vitro screening of lactic acid bacteria isolated from gastrointestinal tract of Atlantic Salmon (Salmo salar) as probiont candidates

A total of 20 lactic acid bacteria (LAB) were isolated from the gastrointestinal tracts of Atlantic salmon (Salmo salar) and screened for antimicrobial activity against six fish pathogens using either a microtiter plate assay or a well diffusion assay. The results indicated that three LAB exhibited antimicrobial activity against at least two pathogens. Based on their partial 16S ribosomal deoxyribonucleic acid (16S rDNA) sequences, these isolates were identified as Lactobacillus farraginis, Pediococcus acidilactici, and P. pentosaceus. In addition, these LAB were able to tolerate the simulated gastrointestinal tract conditions (stomach and intestine), had a good adhesion capacity to intestinal mucus as well as were able to grow in intestinal mucus of Atlantic salmon. These results suggest that these LAB are potential probionts in aquaculture. However, further studies are required to evaluate probiotic properties in aquatic animals.

Sex reversal of brook trout (Salvelinus fontinalis) by 17α-methyltestosterone exposure: A serial experimental approach to determine optimal timing and delivery regimes

Commercial culture of Brook trout (Salvelinus fontinalis) in Tasmania was partly abandoned due to sexual maturation of male fish early on during the estuarine rearing phase. Maturation adversely affects body mass, flesh quality and immunocompetency effectively. Sex reversal techniques such as the in-feed addition of a synthetic androgen have proven difficult to adapt in brook trout. An appropriate timing, duration and delivery vehicle for administration of 17α-methyltestosterone (MT) to produce phenotypic males (neomales) from genotypically female brook trout required further investigation. In this study, groups of brook trout eggs (n=1000) maintained at 9.5±0.15-10±0.14°C, were immersed in MT (400μgL-1) for four hours on two alternate days (two immersions/group) staggered over a two week period surrounding the hatch of embryos (control groups excluded). The groups were then split and half received MT-supplemented feed for 60days and the other a standard diet. Following an 11 month on-growing period sex phenotypes were determined by gross & histological gonad morphology. The highest proportion of male phenotypes (75%) was found in fish immersed six and four days pre-hatch and subsequently fed a normal diet. Fish fed a MT supplemented diet and immersed in MT showed significantly higher proportions of sterile fish. These data indicate that a pre-hatch immersion-only regime (4-6days pre-hatch at 9.5°C) should be pursued as a target for optimization studies to further refine the effective concentration and duration of exposure to MT for the successful production of neo-male brook trout.