Morphology of the Atlantic salmon (Salmo salar) tongue

The Atlantic salmon (Salmo salar) is a freshwater and marine fish of the family Salmonidae, widely farmed in aquaculture facilities in several countries. The salmon are carnivorous, but in aquaculture, alternative foods have been experienced. It is well known that feeding in captivity should cause adaptation and modifications of the morphological characteristics of the oral cavity, especially of tongue; therefore, the aim of this study was to investigate, by light, laser confocal and scanning electron microscopy, the morphological characteristics of the tongue dorsal surface, considering the importance of the correlations between feeding habits and the anatomy of the tongue. Scanning electron microscopy demonstrates the presence of caniniform teeth with oro-aboral orientation surrounded by numerous filiform papillae, single, fused or arranged in row. Oro-aborally, the papillae show an appearance like a rosette and they disappear at level of the root. Light and laser confocal microscopy demonstrates that the mucosa is covered by a non-keratinized stratified pavement epithelium with, in the deepest layer, the presence of a triangular structure whose apex is cranially directed and base facing aborally. In this structure, spindle-shaped cells are present, with a vimentin immunoreactivity, that for their characteristics could be adult mesenchymal stem cells. The obtained data could be useful not only for further studies on the nutrition, but it is interesting the detection of tissues typical of the embryo-fetal phase in the adult specimens tongue, thus giving a basis for studies of potential applications, if any, regarding cell therapies for different clinical indications.     

Comparing efficiency of metabolizable energy utilization by rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar) using factorial and multivariate approaches

A study was conducted to compare utilization of ME for growth vs. maintenance in rainbow trout and Atlantic salmon. Fish were hand-fed to satiation one of four isoenergetic diets (DE = 20 MJ/kg, as-fed basis) with different digestible protein (DP) to DE ratios (24, 22, 20, and 18 g/MJ). Intake of ME (kJ/d), energy deposited as protein (PD, kJ/d), and energy deposited as lipid (LD, kJ/d) were determined by a comparative slaughter technique. Data were analyzed by a factorial approach or by multivariate analysis of PD and LD on ME. Maintenance energy requirements (ME(m)) and efficiency of ME utilization for PD (k(p)) and LD (k(f)) were estimated with both approaches. For the multivariate analysis, an additional parameter, the fraction of ME intake above maintenance used for PD (X) was defined as linear function of BW, with slope (d) and intercept (c) estimated simultaneously with the above parameters. Estimates were highly dependent on the approach and assumptions used. The ME(m) and k(p) values were higher and less accurate with the factorial approach than with multivariate analysis. The factorial approach estimated unrealistic k(f) values (k(f) > 1). With the multivariate analysis, ME(m) did not differ between species (20 kJ x d(-1) x kg(-0.8)). On the other hand, k(p) was significantly higher (e.g., 0.52 +/- 0.06 vs. 0.43 +/- 0.06; P < 0.05) for salmon than for trout and independent of diet, but k(f) was 0.81 (+/-0.13) regardless of species or diet. The ME intake above ME(m) used for PD (c) was higher in salmon than trout (57 vs. 55%; P < 0.05). The change in partitioning of ME for PD due to the change in BW was negative for trout (d = -0.18), but positive for salmon (d = 0.16). The d values agreed well with the increase of LD:PD ratio with BW for trout and the decrease of LD:PD with BW for salmon, which may have been related to the maturation status of this fish and the associated loss of body lipid observed by maturing salmon. In conclusion, ME(m) and cost of LD were similar for rainbow trout and Atlantic salmon, but the cost of PD was lower for salmon than for trout.     

Furunculosis in rainbow trout (Salmo gairdneri) raised in sea water

Ulcerative skin lesions were encountered in rainbow trout raised in sea water by a commercial concern in the Western Cape, South Africa. Grossly, the lesions resembled furunculosis but, histopathologically, they differed from typical furunculosis in that bacterial colonies were rarely found in the organs, and also the kidneys and spleens were minimally involved. The causative organism was identified as an achromogenic Aeromonas salmonicida that shared characteristics with all 3 subspecies, salmonicida, masoucida and achromogenes. This is the first report of an outbreak of this disease in South Africa.     

Tissue distribution of 14C-diflubenzuron in Atlantic salmon (Salmo salar)

Diflubenzuron is a potent inhibitor of chitin synthesis, with potential use against salmon lice infestations. The absorption, distribution and elimination of the substance in Atlantic salmon was examined after a single, oral dose of 75 mg/kg bodyweight. The kinetic properties were studied by whole-body autoradiography, liquid scintillation counting and thin layer chromatography, using a 14C-labelled isotope of the substance. The drug was poorly absorbed from the intestine, but reached a concentration of more than 4 micrograms/g in the mucus layer of the skin 2 days after administration. If maintained for several days, this concentration is probably sufficient to control all moulting stages of sea lice in Atlantic salmon. The main route of excretion was via the bile.     

Clinical pathology of myodegeneration (pancreas disease) in Atlantic salmon (Salmo salar)

The clinical pathology and histopathology of two groups of Atlantic salmon with severe degenerative myopathy (pancreas disease) is described and compared with a third healthy group. One affected group was anorexic and had low plasma protein and albumin levels while the other was feeding and had normal levels. Both diseased groups had plasma and tissue vitamin E and selenium levels lower than the healthy group. Similarly, creatine kinase values were raised in affected groups. If representative of the syndrome as a whole, the results suggest that the myopathy of pancreas disease has a basis in a vitamin E-selenium deficiency, but whether primary or induced is not clear. The results also demonstrate that the myopathy and pancreatic atrophy do not inevitably lead to anorexia or any other clinically obvious sign of disease, despite both cardiac and oesophageal involvement.     

Further studies on the alpha-precipitins of the Atlantic salmon (Salmo salar)

The alpha-precipitins of the Atlantic salmon have been isolated by a combination of chromatography and gel filtration. A monospecific antiserum has been raised and this has been used to measure the concentration of the antigen in 925 fish sera by CELISA. The results were analysed for the effects of sex and disease state of the animal, time of year and year of collection, and the part of the UK from which they were collected. Significant differences were found in all cases. The components of alpha-precipitin were investigated by the isoelectric focusing of pure alpha-precipitin, whole serum and serum heated for 18 h at 95 degrees C. Multiple banding occurred in the acid region of the gels; the number, relative strength and position of the bands depended upon the animal from which the serum was isolated.     

Effect of an immunopotentiator on Aeromonas salmonicida infection in rainbow trout (Salmo gairdneri)

The immunoactive peptide FK-565 (heptanoyl-y-D-glutamyl-(L)-mesodiaminopimelyl-(D)-alanine) was found to induce protection against intraperitoneal Aeromonas salmonicida infection in rainbow trout (Salmo gairdneri Richardson). The survival rate was as high as 60% when FK-565 was given intraperitoneally as a single dose (1mg/kg) one day before bacterial challenge. A non-specific stimulation of phagocytic cells by FK-565 at an early stage of the bacterial infection may contribute to the resistance observed. The phagocytic activity of peritoneal phagocytic cells as well as phagocytic cells of the pronephros were stimulated by FK-565 in vivo and in vitro, respectively, as compared to untreated control fish. Furthermore, decreased activity of phagocytic cells previously immunosuppressed with cyclophosphamide was rapidly restored by application of FK-565.     

Glandular kallikrein in the innate immune system of Atlantic salmon (Salmo salar)

Glandular Kallikrein is a serine-protease with trypsin-like activity and is able to generate bioactive peptides from inactive precursors. We have evaluated the presence of this protease in the different organs of the Atlantic salmon (Salmo salar). The results clearly indicate that GK and PRL are generated in the same pituitary cells based on a co-localization by confocal microscopy. Based on probed cross-reactivity between C. striata and C. carpio glandular anti-GK antibodies, we used a homologous antibody to detect the presence of GK in several salmon tissues. We have evaluated the GK expression in healthy and defied fish. P. salmonis and V. ordalii. The GK immunoreaction in organs such as leukocytes, gills and skin is considerably increased in defied fish compared to healthy fish. This increase was present in the cells of the excretory kidney and in the intercellular tissue, where the development of hematopoietic and lymphocytic lines in fish take place. One of the most interesting organs to study was the skin, bearing in mind that this is a primary barrier to all pathogens. The skin of the defied fish exhibited an increase in immunoreactivity for glandular kallikrein similar to the protease found in mucus. An immunoreactive tissue kallikrein-like protein was identified and partially separated by perfusion chromatography. Enzymatic activity of salmon muscle prokallikrein was determined before and after trypsin activation. Kallikrein activity was characterized with respect to their ability to cleave the chromogenic leaving group, p-nitroanilide, from the peptidyl kallikrein and trypsin substrate. These findings constitute a important contribution to reveal the role of kallikrein in the innate immune system of fish.     

Comparative pharmacokinetics of oxytetracycline in rainbow trout (Salmo gairdneri) and African catfish (Clarias gariepinus)

A comparative pharmacokinetic study was conducted in rainbow trout (Salmo gairdneri) and African catfish (Clarias gariepinus) following intravenous (i.v.) and intramuscular (i.m.) administration of oxytetracycline (OTC) at a dose rate of 60 mg/kg body weight. Trout and catfish were kept in aerated tap water in tanks at constant temperatures of 12 degrees C and 25 degrees C, respectively. The two- and three-compartment open models adequately described plasma drug disposition in African catfish and rainbow trout respectively, following i.v. OTC administration. Compared to catfish (COP = 86 +/- 10 micrograms/ml) an eightfold higher extrapolated zero time concentration was obtained in trout (COP = 753 +/- 290 micrograms/ml). A significant difference was observed with respect to the relatively large apparent distribution volumes (Vd(area] after i.v. OTC administration (trout, mean value: 2.1 l/kg; catfish, mean value: 1.3 l/kg). The mean final elimination half-lives of both fish species were greater than previously reported in mammals (trout, 89.5 h; catfish, 80.3 h). A mean maximum plasma concentration (Cmax = 56.9 micrograms/ml) was obtained in trout at 4 h after i.m. administration of OTC. In catfish a lower Cmax of 43.4 micrograms/ml was determined at about 7 h. No significant difference was observed with respect to bioavailability following i.m. administration of OTC (trout, 85%; catfish, 86%).     

Distribution of emamectin benzoate in Atlantic salmon (Salmo salar L.)

The aims of this study were to investigate the content of emamectin in blood, mucus and muscle following field administration of the recommended dose, and correlation with sea lice infection on the same fish (elimination study). The tissue distribution of tritiated emamectin benzoate after a single oral dose in Atlantic salmon was also investigated by means of whole-body autoradiography and scintillation counting (distribution study). In the elimination study, concentrations of emamectin benzoate reached maximum levels of 128, 105 and 68 ng/g (p.p.b.) for blood, mucus and muscle respectively, on day 7, the last day of administration. From day 7, the concentration in the blood declined until concentration was less than the limit of detection on day 77. The concentration was higher in mucus compared with plasma (P < 0.05) except on days 7 and 21. The concentration of emamectin benzoate decreased gradually from the end of treatment (day 7) to day 70 with half-lives of 9.2, 10.0 and 11.3 days in muscle, plasma and mucus respectively. The distribution study demonstrated a high quantity of radioactivity in mucous membranes (gastrointestinal tract, gills) throughout the observation period (56 days). Activity was high in the epiphysis, hypophysis and olfactory rosette throughout the study. The highest activity was observed in the bile, indicating this to be an important route for excretion. The distribution study confirmed the results from the elimination study with respect to concentrations in blood, skin mucous and muscle.     

Effects of metals on the chemiluminescent response of rainbow trout (Salmo gairdneri) phagocytes

Quantification of an induced chemiluminescent (CL) response in phagocytes is currently being evaluated as an indicator system for determining those environmental pollutants that may predispose fish to disease. A CL assay was developed using phagocytes from the pronephros of rainbow trout (Salmo gairdneri). The CL response of phagocytes to phorbol myristate acetate, a chemical inducer of CL, was shown to be dose-dependent. The response to five species of bacteria was also evaluated. Staphylococcus aureus and Aeromonas hydrophila produced the most intense CL responses and the longest duration of response (100 min.) Yersinia ruckeri induced an immediate strong CL response of short duration (20 min.) whereas Vibrio anguillarum and Aerococcus viridans failed to stimulate CL under the test conditions employed. The effect of sub-toxic levels of Cu, Al, and Cd on the CL response of phagocytes to S. aureus was examined using phagocytes exposed to the metals immediately before assay or after 1 hr or 24 hr exposure times. Copper caused a significant decrease in CL to the baseline level under all treatment conditions upon stimulation with S. aureus. Similar results were obtained with Al except that the decrease in CL, although significant, was not to the baseline level. In contrast, Cd caused a significant increase in CL when added 1 hr prior to or immediately before the assay; but, following a 24 hr exposure, the results were variable, in that either no change or a decrease was observed. The addition of Cu to phagocytes already exhibiting a strong CL response to S. aureus caused an immediate decrease in CL to that seen with the negative controls.     

The ontogeny of humoral immunity in rainbow trout, Salmo gairdneri

The ontogeny of the humoral immune response to a 'thymus dependent' and a 'thymus independent' antigen, human gamma globulin (HGG) and Aeromonas salmonicida (AS) respectively, was investigated in the fry of rainbow trout, Salmo gairdneri, by direct immersion vaccination in the antigens (dose 5 mg/L HGG; 10(8) cells/ml AS; 30 minutes) at known ages/weights from 7 days post hatch, and at 1, 2, 3, and 4 months post hatch. Half the fry in each group were tested for antibodies 4 weeks after vaccination, the remainder were reimmunised and tested again after a further 4 weeks. Appropriate controls to test for tolerance induction and memory responses were included. The results indicate that fry are capable of mounting a humoral immune response very early in ontogeny. There is a period of 'unresponsiveness' which persists for longer against HGG, than against AS, though it was not thought to be tolerance as such. Memory could be detected to HGG in fry given a first immunisation at 2 months. The results are compared with preliminary experiments in which fry were first thymectomised 4 weeks before the first immunisation. In fry thymectomised at 1 month post hatch, and tested for primary and secondary responses at 2 and 3 months, the primary response to HGG is unaltered, but the secondary response is reduced. Both the primary and secondary response to AS is unaltered. When thymectomy is performed later, the effect on the secondary response to HGG is no longer apparent, but the primary response to AS is slightly reduced.     

Effect of immunization route on mucosal and systemic immune response in Atlantic salmon (Salmo salar)

This study aimed to assess systemic and mucosal immune responses of Atlantic salmon (Salmo salar) exposed to two protein-hapten antigens – dinitrophenol (DNP) and fluorescein isothiocyanate (FITC) each conjugated with keyhole limpet haemocyanin (KLH) – administered using different delivery strategies. Fish were exposed to the antigens through different routes, and were given a booster 4 weeks post initial exposure. Both systemic and mucosal antibody responses were measured for a period of 12 weeks using an enzyme-linked immunosorbent assay (ELISA). Only fish exposed to both antigens via intraperitoneal (IP) injection showed increased systemic antibody response starting 6 weeks post immunization. No treatment was able to produce a mucosal antibody response; however there was an increase in antibody levels in the tissue supernatant from skin explants obtained 12 weeks post immunization from fish injected with FITC. Western blots probed with serum and culture supernatant from skin explants showed a specific response against the antigens. In conclusion, IP injection of hapten-antigen in Atlantic salmon was the best delivery route for inducing an antibody response against these antigens in this species. Even though IP injection did not induce an increase in antibody levels in the skin mucus, there was an increased systemic antibody response and an apparent increase of antibody production in mucosal tissues as demonstrated by the increased level of specific antibody levels in supernatants from the tissue explants.