Accumulation of immunomodulatory laminaran (β(1,3)-D-glucan) in the heart, spleen and kidney of Atlantic cod, Gadus morhua L.

The tissue distribution of ³H- and FITC-lammaran in Atlantic cod, Gadus morhua L., was investigated after intravenous administration. Liquid scintillation counting and whole-body autoradiography revealed a substantial accumulation of ³H-labelled laminaran in the heart throughout the experimental period (1 h to 12 days). High amounts of the immunomodulator were also present in the spleen, anterior kidney and posterior kidney during the study. Renal excretion and intestinal exsorption were the main elimination pathways. Fluorescence microscopic examination revealed accumulation of FITC-labelled laminaran in macrophages in the kidney and ellipsoid sheath cells (macrophages and reticular cells) in the spleen. Furthermore, endothelial entrapment in the heart was pronounced. Endothelial cells in the kidney also contained the FITC-labelled immunomodulator. Immunohistochemical analysis, using anti-β(l,3)-D-glucopyranose antibody, also showed endothelial uptake of laminaran in the heart. The present investigation shows that laminaran accumulates and is retained in immunologically relevant cells in the Atlantic cod.     

The immunomodulatory effect of laminaran [β(1,3)-D-glucan] on Atlantic salmon, Salmo salar L., anterior kidney leucocytes after intraperitoneal, peroral and peranal administration

Anterior kidney leucocytes obtained from Atlantic salmon, Salmo salar L., 2 days after administration of laminaran, were assayed for their capacity to reduce nitroblue tetrazolium to formazan, and for their activity of lysosomal acid phosphatase after intraperitoneal (15 mgkg^?1), peroral (150 mg kg^?1) or peranal (150 mg kg^?1) administration. Leucocytes obtained from salmon treated by an intraperitoneal injection of laminaran produced significantly more superoxide anion than cells obtained from fish treated with dextran or sodium chloride immediately after cell isolation. Immediately after extraction, the activity of acid phosphatase in anterior kidney leucocytes obtained from salmon injected with laminaran was significantly higher than in cells harvested from fish treated with dextran or sodium chloride. Furthermore, cells obtained from salmon treated by peroral instillation of laminaran showed significantly enhanced production of superoxide anion compared with leucocytes from fish treated with either sodium chloride or dextran. The acid phosphatase activity in anterior kidney leucocytes from salmon treated by peroral and peranal instillation of laminaran was significantly higher than in cells from fish treated either with sodium chloride or dextran. Finally, fluorescence microscopic examination of tissue sections from fish treated peranally by intubation with fluorescein labelled laminaran revealed fluorescent vesicles in intestinal epithelial cells and in anterior kidney macrophages.     

Differences between Atlantic halibut (Hippoglossus hippoglossus L.) and turbot (Scophthalmus maximus L.) in tolerance to acute low temperature exposure

Juvenile Atlantic halibut (Hippoglossus hippoglossus L.) and turbot (Scophthalmus maximus L.) acclimated to 8–9 °C sea water were transferred to sea water at 1 °C for 7 days. An immediate spasm-like response was noticed in the turbot after the transfer, but not in the halibut. None of the fish died. The turbot suffered from substantial physiological disturbances after the transfer. Their blood plasma Cl^– concentration increased and muscle water content decreased markedly. The plasma glucose concentration was several times higher than the control level from day one onwards. A decrease in the haematocrit was recorded some hours after transfer, followed by a marked increase on day 7. In halibut, the plasma Cl^– concentration fell somewhat during the first days, but returned thereafter to its pretransfer level. The muscle water content was unchanged. Both haematocrit and plasma glucose concentration were unchanged until day 7, when both were significantly higher. The differences in the response to low-temperature challenge between halibut and turbot are probably related to genetic differences in temperature tolerance, which reflect differences in the distribution of the species.     

A comparative study of susceptibility and induced pathology of cod, Gadus morhua (L.), and halibut, Hippoglossus hippoglossus (L.), following experimental infection with Moritella viscosa

In this study experimental challenges with Moritella viscosa, the causative agent of winter ulcers in salmonids, were performed on juvenile Atlantic cod and Atlantic halibut. The challenges involved both intramuscular (i.m.) and intraperitoneal (i.p.) injections and bath with a strain originally isolated from Atlantic salmon. Cod was found to be significantly more sensitive than halibut to the infection. Both fish species were found to be more sensitive to i.m. than i.p. challenges. Both challenges induced a systemic disease in cod and halibut, but only cod was infected with an experimental bath challenge. Pathognomonic signs were found to be comparable with those described in M. viscosa-infected salmon and turbot. The main distinguishing pathological sign was that the cod showed host response to M. viscosa infection resulting in granuloma formation in infected tissues, which is a known response of cod to a infection with another Gram-negative bacterium, Aeromonas salmonicida. Re-isolation of M. viscosa from kidneys of cod and halibut with clear disease signs was problematic and optimization of isolation measures is needed. The results of this study indicate that M. viscosa infection may be a risk factor in cod farming, but that halibut is more resistant.     

Comparative study of antioxidant defence mechanisms in marine fish fed variable levels of oxidised oil and vitamin E

The aim of the study was to compare theantioxidant systems in juvenile marine fish ofcommercial importance to European aquaculture,namely turbot (Scophthalmus maximus),halibut (Hippoglossus hippoglossus) andgilthead sea bream (Sparus aurata). Thepresent dietary trial was specifically designedto investigate the antioxidant effects ofvitamin E under moderate oxidising conditions,including high dietary levels of highlyunsaturated fatty acids and the feeding ofoxidised oils. The objective was to induce astressful pro-oxidant status to enablecharacterisation of the biochemical responsesto peroxidative stress without causingunnecessary suffering to the experimentalanimals or high mortalities during the trials. Both sea bream and turbot showed excellentgrowth, whereas growth was poorer in halibut.Dietary oxidised oil significantly reducedgrowth in turbot and especially in halibut, butnot in sea bream. Vitamin E improved growth insea bream fed oxidised oil but not in turbot orhalibut. However, vitamin E supplementationappeared to improve survival in all threespecies. In sea bream and turbot, liverantioxidant defence enzyme activities weregenerally increased by feeding peroxidised oiland reduced by vitamin E. Conversely, inhalibut, the liver antioxidant defence enzymeactivities were not increased by feedingperoxidised oil and only superoxide dismutasewas reduced by feeding vitamin E. Consistentwith these data, feeding oxidised oil increasedlipid peroxidation products in halibut, butgenerally not in sea bream or turbot.Furthermore, lipid peroxidation products weregenerally reduced by dietary vitamin E in bothsea bream and turbot, but not in halibut. Therefore, halibut liver antioxidant defenceenzymes did not respond to dietary oxidised oilor vitamin E as occurred in turbot and,especially sea bream. This resulted inincreased levels of lipid peroxides in halibutcompared to turbot and sea bream in fish givendietary oxidised oil. In addition, supplementalvitamin E did not reduce lipid peroxides inhalibut as it did in turbot and sea bream. Theincreased peroxidation stress in halibut mayaccount for their poorer growth and survival incomparison to turbot and especially sea bream.Halibut were reared at a lower temperature,although relatively high for halibut, thaneither turbot or sea bream but they were alsoslightly younger/smaller fish and possibly,therefore, more developmentally immature, andeither or all of these factors may be importantin the lack of response of the liver enzymes inhalibut.     

Tissue tropism of nervous necrosis virus (NNV) in Atlantic cod, Gadus morhua L., after intraperitoneal challenge with a virus isolate from diseased Atlantic halibut, Hippoglossus hippoglossus (L.)

Atlantic cod, Gadus morhua , averaging 100 g, were experimentally challenged by intraperitoneal injection of nervous necrosis virus (NNV) originating from Atlantic halibut. Cod tissues, including blood, gill, pectoral fin, barbel, ventricle, atrium, spleen, liver, lateral line (including muscle tissue), eye (retina) and brain, were sampled at day 25 and 130 and investigated by real-time RT-PCR for the presence of NNV. Relative quantifications at day 130 were calculated using the 2^−ΔΔCt method. Immunosuppression by injection of prednisolone-acetate was introduced for a 30-day period, and tissue sampled at day 180 and relative quantification estimated. No mortality or clinical signs of disease were observed in the challenged group. The challenge resulted in detection of NNV in blood, spleen, kidney, liver, heart atrium and heart ventricle at day 25, and by the end of the experiment NNV showed a clear increase in brain and retina, suggesting these to be the primary tissues for viral replication. There was no increase in the relative amount of NNV in blood, atrium, ventricle, spleen, liver and kidney. Corticosteroid implants resulted in a weak increase in virus RNA in spleen, kidney, liver and brain. These findings suggest that Atlantic cod is susceptible to infection with NNV from halibut. The observed tissue tropism patterns suggest an initial viraemic phase, followed by neurotrophy. Head-kidney is the best tissue identified for possible NNV detection by non-lethal biopsy, but detection was not possible in all injected fish.     

Distribution of α-tocopherol in fillets of turbot (Scophthalmus maximus) and Atlantic halibut (Hippoglossus hippoglossus), following dietary α-tocopheryl acetate supplementation

The present study investigated the distribution of α‐tocopherol (vitamin E) in fillets of turbot (Scophthalmus maximus) and Atlantic halibut (Hippoglossus hippoglossus). Turbot and Atlantic halibut were fed commercial diets, supplemented with different levels of α‐tocopheryl acetate at the dietary target levels of 100, 500 and 1000 mg α‐tocopheryl acetate kg^?1 diet. The actual levels were 72, 547 and 969 for turbot, while halibut received 189, 613 and 875 mg α‐tocopheryl acetate kg^?1 diet. Turbot were fed the diets for 24 weeks, while Atlantic halibut were fed for 20 weeks prior to slaughter. At the end of the feeding periods fish had reached a final weight of around 1 kg. Fish were slaughtered and filleted. From the four fillets obtained per fish, 22 samples were taken from designated areas and analysed for their α‐tocopherol content. The average concentrations of α‐tocopherol incorporated in turbot and Atlantic halibut increased with increasing levels of α‐tocopheryl acetate in the diet. Atlantic halibut had significantly (P < 0.05) more α‐tocopherol in positions 2/II and 1/I than in position 9/IX. Turbot had significantly (P < 0.05) more α‐tocopherol in position 2/II than in positions 1/I, 4/IV and 11/XI. By mapping α‐tocopherol concentrations in fish fillets, a high degree of quality prediction may be established. Moreover, this study may help scientists in their choice of sampling position, when investigating if α‐tocopheryl acetate supplementation resulted in successful α‐tocopherol incorporation.     

Comparative composition and shelf-life of fillets of wild and cultured turbot (Scophthalmus maximus) and Atlantic halibut (Hippoglossus hippoglossus)

Turbot and Atlantic halibut are highly valued fish species. However,very little is known about fillet shelf-life characteristics associated withboth species. Thus, fillet -tocopherol content and proximate compositionof wild turbot (1.5 kg) and Atlantic halibut (1.1 kg)caught off the south coast of Ireland and the north-west coast of Iceland,respectively, were investigated. In addition, the susceptibility of fillets, storedunder retail conditions, to lipid oxidation and colour change was studied.Proximate composition analysis showed that turbot had significantly highermoisture (P < 0.001) and lower protein (P < 0.001) contents compared toAtlantic halibut. Atlantic halibut incorporated significantly higher (P <0.001) levels of -tocopherol into fillets than turbot. Over 14 days ofstorage on ice, fillets from Atlantic halibut exhibited significantly lower (P =0.020) levels of lipid oxidation than those of turbot. However, malondialdehyde(MDA) concentrations were generally very low, never exceeding 0.6 gg^–1 fillet. Turbot maintained a significantly higher (P< 0.001) pH over the storage period. The lightness (L^* values) offillets from both species increased over 14 days of storage, but wassignificantly higher (P < 0.001) in Atlantic halibut than in turbot. Turbotdeveloped a relatively intense yellow colour during storage (decrease in hueangle and increase in b^* values), whereas this was not the case forAtlantic halibut. The results of this study demonstrate that fillets of wildAtlantic halibut stored on ice, were less prone to lipid oxidation anddiscolouration than those of wild turbot. However, quality changes in turbotwere very small showing that both fish have tremendous shelf-life capacities interms of lipid oxidation. These findings are considered in the context of knownmaterial for farmed fish.     

A study of the susceptibility of Atlantic halibut,Hippoglossus hippoglossus (L.), to viral haemorrhagic septicaemia virus isolated from turbot,Scophthalmus maximus (L.)

The susceptibility of Atlantic halibut, Hippoglossus hippoglossus (L.), to viral haemorrhagic septicaemia virus (VHSV) was tested. Juvenile halibut of approximately 5 g weight were subjected to challenge by intraperitoneal injection, cohabitation and immersion to a VHSV isolate from an outbreak of the disease in turbot, Scophthalmus maximus (L.). The intraperitoneal injection gave the highest mortality rate of 28% after 50 days. The cohabitee group suffered 19% mortality rate and the immersion group only 2%. Control groups included turbot exposed either by intraperitoneal injection or immersion which suffered mortality rates of 93 and 50%, respectively. The results suggest that halibut are markedly less susceptible to VHSV than turbot.     

Evaluation of Electrical Stunning of Atlantic Cod (Gadus morhua) and Turbot (Psetta maxima) in Seawater

The aim of this study was to assess electrical stunning of Atlantic cod and turbot in seawater to develop a protocol for the process of stunning and killing. An induced general epileptiform insult (unconscious) had a duration of 40 ± 27 s (n =14) in cod (2.6 ± 0.5 kg) and 34 ± 18 s (n = 19) in turbot (520 ± 65 g). Seven cod and 3 turbot displayed a physical reaction, and 11 turbot registered an electroencephalogram (EEG) response to pain stimuli administered 30 s post-stun. The heart rate was 32 ± 6 beats/min in cod and 25 ± 7 beats/min in turbot prior to stunning. Post-stunning, the electrocardiogram (ECG) revealed fibrillation and reduced activity post-stun. EEG, ECG recordings, and behavioral observations indicate that when a bipolar square wave current was applied with a frequency of 133 Hz and 43% duty cycle side to side (turbot) and at 170 Hz and 33% duty cycle (cod) head to tail, both species were stunned in seawater at current densities of 3.2 A/dm² and 2.5 A/dm², respectively. For turbot, a 5 s exposure to electricity followed by chilling in ice water for 15 min is sufficient to prevent recovery. For cod, a killing method needs to be established.     

Retention of furunculosis vaccine components in Atlantic salmon, Salmo salar L., following different routes of vaccine administration

The retention of vaccine components was studied in Atlantic salmon, Salmo salar L., following different routes of vaccination against Aeromonas salmonicida. Frozen tissue was collected from the spleen, head kidney, hind gut and liver of fish that had been vaccinated by intraperitoneal injection (monovalent and trivalent vaccines), immersion and oral administration 2,6,8 and 16 weeks previously. The trivalent injection group showed the highest levels of specific antibodies and was the only group to show protection following challenge with virulent A. salmonicida. Following intraperitoneal injection, there was an initial widespread distribution of Aeromonas lipo-polysaccharide, but by 16 weeks lipopolysaccharide was predominantly found in macrophage populations in the spleen, head kidney and abdominal granulomas. Only small amounts of lipopolysaccharide were retained in the head kidney of the immersion group and no lipopolysaccharide was retained in the oral group. Small and inconsistent amounts of A-layer protein were present in the meianomacrophages of the head kidney of all groups. The relative prominence of A-layer protein in the spleen of the trivalent injection group 8 weeks after vaccination was linked to the high levels of specific antibodies, and possible immune-complex trapping and the enhancement of immunological memory.     

Anesthesia induces stress in Atlantic salmon (<Emphasis Type="Italic">Salmo salar</Emphasis>), Atlantic cod (<Emphasis Type="Italic">Gadus morhua</Emphasis>) and Atlantic halibut (<Emphasis Type="Italic">Hippoglossus hippoglossus</Emphasis>)

Stress in response to anesthesia with benzocaine, MS-222, metomidate and isoeugenol was studied in Atlantic salmon (Salmo salar), Atlantic halibut (Hippoglossus hippoglossus), and Atlantic cod (Gadus morhua) with no concomitant stress from handling or confinement in association with anesthesia or sampling. All of the anesthetics tested induced a stress response in all species, displayed by a release of cortisol to the water. MS-222 anesthesia elicited the highest cortisol release rates, reaching maximum levels 0.5 h post-exposure and returning to basal levels after 3–4 h. Benzocaine anesthesia caused a bimodal response where the initial peak in cortisol release rate was followed by a second increase lasting towards the end of the trial (6 h). This bimodality was more profound in Atlantic salmon than in Atlantic halibut and Atlantic cod. Metomidate anesthesia induced the lowest release of cortisol of the agents tested in both Atlantic halibut and Atlantic cod, but resulted in a bimodal response in Atlantic salmon where the initial increase in cortisol release was followed by a larger increase peaking at 2–2.5 h post exposure before returning to basal levels after 5 h. The stress induced in Atlantic salmon by isoeugenol anesthesia resembled that of MS-222, but did not reach the same elevated level. Overall, the cortisol release was most profound in Atlantic salmon followed by Atlantic halibut and Atlantic cod.     

A cost‐effectiveness analysis of live feeds in juvenile turbot Scophthalmus maximus (Linnaeus, 1758) farming: copepods versus Artemia

The biological benefits of copepods as live feed for marine finfish larvae have already been well established in the literature. Copepods have better biochemical compositions that improve growth, reduce malpigmentations and allow successful farming of ‘new’ marine finfish species. However, their current usage is quite limited. One of the reasons has been lack of economic knowledge concerning the cost‐effectiveness of copepod application compared to other commonly used feed items such as the brine shrimp Artemia. In this study, a cost‐effectiveness analysis is made on two alternative live feed items (copepods and Artemia) in juvenile turbot farming. Unit cost of production and profit are compared between the two feeding regimes using a unique data set from an existing turbot fry production facility in Denmark. The result reveals that copepods are not only biochemically superior but they are also economically a cost‐effective alternative. Thus, a commercial use of copepods will significantly reduce the production costs for turbot. Furthermore, the unexploited economic potential can be utilized for the successful farming of other high‐valued marine finfish species such as tuna, flounders, cod, sole and halibut. Generally, the biochemical superiority coupled with economic benefits can lead to the commercial utilization of copepods as complementary live feed in the short run and in some situations as a substitute in the long run.     

Antibacterial efficacy and pharmacokinetic evaluation of sanguinarine in common carp (Cyprinus carpio) following a single intraperitoneal administration

Sanguinarine (SA), with antimicrobial and antiparasitic activities against fish pathogens, exhibits great potential commercial use in aquaculture. However, little information on pharmacokinetics of SA restricts further application in aquaculture. In this study, pharmacokinetics of SA in common carp (Cyprinus carpio) following a single intraperitoneal administration [10 mg kg^?1 BW (body weight)] was evaluated by high‐performance liquid chromatography (HPLC). The peak concentration (C_max) of SA in kidney was 11.8 μg g^?1, which was higher than in other tissues and plasma. The terminal half‐life in fish tissue and plasma was as follows: 42.3 h (kidney) > 37.2 h (liver) > 20.1 h (gill) > 18.8 h (muscle) > 10.9 h (spleen) > 10.0 h (plasma). Additionally, we determined the bacterial loads in tissues of common carp infected with Aeromonas hydrophila after i.p. administration of SA at 0, 5, 10 and 20 mg kg^?1 BW. The results showed that i.p. administration of SA at 10 mg kg^?1 BW significantly enhanced antibacterial efficacy against A. hydrophila, where the antibacterial ratio in the gill, kidney, spleen and liver on day 5 was 95.13%, 93.33%, 90.09% and 92.82%, respectively. Overall, these results suggested the potential of SA to treat A. hydrophila infection in common carp farming industry.     

Experimental susceptibility of Atlantic cod, Gadus morhua (L.), and Atlantic halibut, Hippoglossus hippoglossus (L.), to different genotypes of viral haemorrhagic septicaemia virus

Viral haemorrhagic septicaemia (VHS) is a well-characterized disease of rainbow trout, Oncorhynchus mykiss, which has also caused economic losses in marine turbot farms in the British Isles. We have previously demonstrated that turbot, Scophthalmus maximus, are susceptible to isolates of viral haemorrhagic septicaemia virus (VHSV) that are endemic in the marine environment, highlighting a potential risk to marine aquaculture. Given the increasing interest in the intensive rearing of additional aquaculture species such as Atlantic cod, Gadus morhua, and Atlantic halibut, Hippoglossus hippoglossus, this study aimed at investigating the susceptibility of these species to VHSV. Both species were found to be largely resistant to VHS following immersion challenge with a selection of 18 isolates, representing the known marine VHSV genotypes. Only one and two VHSV-associated mortalities occurred out of a total of 1710 and 1254 halibut and cod, respectively. These findings suggest that there is a low direct risk to the development of commercial cod and halibut aquaculture from the existing endemic reservoir of VHSV. This study, coupled to field observations has, however, highlighted the fact that both species can become infected with VHSV. The known adaptability of RNA viruses, together with the selection pressures associated with intensive aquaculture would thus advocate a cautious approach to VHSV surveillance within these emerging industries.     

The effect of dietary chitin on growth and nutrient digestibility in farmed Atlantic cod,Atlantic salmon and Atlantic halibut

The effect of adding 0%, 1%, 2% and 5% chitin from prawn shells in the diets for Atlantic cod, Atlantic halibut and Atlantic salmon on growth was investigated. Nutrient digestibility and feed utilization was investigated in salmon and cod. Atlantic cod grew from 186 ± 29 to 383 ± 78 g (N = 960) over 13 weeks. Dietary chitin had no effect on length, weight, condition, liver size or specific growth rate (SGR). The apparent digestibility (ADC) for protein ranged from 84.7% to 86.5%, lipid between 88.8% and 93.1% and dry matter from 96.1% to 96.6%. Feed utilization varied between 1.08 and 1.11 and was not correlated with dietary chitin content. Atlantic salmon tripled their weight from 199 ± 9 to 615 ± 75 g (N = 480) during the 13 weeks. High inclusions of chitin (>1%) reduced both growth rate and condition. Protein and lipid ADC was negatively correlated with dietary chitin. Feed utilization ranged between 0.86 and 0.90 and was not significantly affected by dietary chitin. Faecal protein increased significantly with increasing dietary chitin, while faecal dry matter and lipid did not. Individually tagged Atlantic halibut grew from 1300 ± 470 to 2061 ± 714 g (N = 70) during 6 months. Individual growth rates varied within each group from being slightly negative to 0.81%·day^?1. Diet had no significant effect on growth rates. Atlantic cod and Atlantic halibut seems unaffected by up to 5% chitin additions in the diet, while chitin >1% of diet negatively affects growth and nutrient utilization in Atlantic salmon.     

Growth and age at first maturity in turbot and halibut reared under different photoperiods

The effect of extended photoperiods on growth and age at first maturity was investigated in 166 (79 females and 87 males) individually tagged Atlantic halibut Hippoglossus hippoglossus and in 114 (50 females and 64 males) individually tagged turbot Scophthalmus maximus. The halibut were reared at 11 °C on four different light regimes from 10 February to 6 July 1996: simulated natural photoperiod, (LDN), continuous light (LD24:0), constant 8 h light and 16 h darkness (LD8:16) and LD8:16 switched to continuous light 4 May 1996 (LD8:16–24:0). From 6 July 1996 to 9 February 1998 the LD24:0 and LD8:16–24:0 were reared together under continuous light and the LDN and LD8:16 together under natural photoperiod. The turbot were reared at 16 °C on three different light regimes: constant light (LD24:0), 16 h light:8 h darkness (LD16:8), or simulated natural photoperiod (LDN). After 6 months on the different photoperiods, the turbot was reared together on LDN for approximately 12 months until first maturation. Juveniles subjected to continuous light (halibut) or extended photoperiods (halibut and turbot) exhibited faster growth than those experiencing a natural photoperiod or a constant short day. Moreover, when the photoperiod increased naturally with day-length or when fish were abruptly switched from being reared on short-day conditions to continuous light, a subsequent increase in growth rate was observed. This growth enhancing effect of extended photoperiods was more apparent on a short time scale in Atlantic halibut than in turbot, but both species show significant long-term effects of extended photoperiods in the form of enhanced growth. In both species lower maturation of males was seen in groups exposed to extended or continuous light compared to LDN and this could be used to reduce precocious maturation in males leading to overall increase in somatic growth. This revised version was published online in August 2006 with corrections to the Cover Date.     

Experimental infection of Atlantic halibut,Hippoglossus hippoglossus L., yolk-sac larvae with infectious pancreatic necrosis virus: detection of virus by immunohistochemistry and in situ hybridization*

Three different concentrations (10⁷, 10⁵ and 10³ TCID₅₀ ml^?1) of infectious pancreatic necrosis virus (IPNV) serotype Sp isolated from Atlantic halibut, Hippoglossus hippoglossus L., were used to bath-challenge Atlantic halibut yolk-sac larvae. The larvae challenged with 10⁷ TCID₅₀ ml^?1 suffered significantly higher cumulative mortality than the other challenged groups and the control group, and affected individuals displayed necrosis of the intestine, liver and kidney. In larvae from the groups challenged with 10⁷ and 10⁵ TCID₅₀ ml^?1, IPNV was detected by immunohistochemistry and in situ RNA/DNA hybridization in the intestine, liver and kidney. In addition, some individuals stained IPNV-positive in the heart and eye/ brain region. Detection by in situ hybridization did not appear to be more sensitive than immunohistochemistry. However, background staining was virtually absent in comparison with immunohistochemistry, and the staining seemed to be more distinctly localized to the cytoplasm of infected cells. The results show that farmed halibut yolk-sac larvae can be infected by IPNV immediately after hatching, with resulting high mortality. As the larvae are not immunologically mature at this stage of development, vaccination is not recommended.     

Experimental infection of Atlantic halibut, Hippoglossus hippoglossus L., yolk-sac larvae with infectious pancreatic necrosis virus: detection of virus by immunohistochemistry and in situ hybridization

Three different concentrations (10⁷, 10⁵ and 10³ TCID₅₀ ml^-1) of infectious pancreatic necrosis virus (IPNV) serotype Sp isolated from Atlantic halibut, Hippoglossus hippoglossus L., were used to bath-challenge Atlantic halibut yolk-sac larvae. The larvae challenged with 10⁷ TCID₅₀ ml^-1 suffered significantly higher cumulative mortality than the other challenged groups and the control group, and affected individuals displayed necrosis of the intestine, liver and kidney. In larvae from the groups challenged with 10⁷ and 10⁵ TCID₅₀ ml^-1, IPNV was detected by immunohistochemistry and in situ RNA/DNA hybridization in the intestine, liver and kidney. In addition, some individuals stained IPNV-positive in the heart and eye/brain region. Detection by in situ hybridization did not appear to be more sensitive than immunohistochemistry. However, background staining was virtually absent in comparison with immunohistochemistry, and the staining seemed to be more distinctly localized to the cytoplasm of infected cells. The results show that farmed halibut yolk-sac larvae can be infected by IPNV immediately after hatching, with resulting high mortality. As the larvae are not immunologically mature at this stage of development, vaccination is not recommended.     

A single-dose pharmacokinetic study of oxolinic acid and vetoquinol, an oxolinic acid ester, in Atlantic halibut, Hippoglossus hippoglossus L., held in sea water at 9 °C

The pharmacokinetic properties of the antibacterial agent oxolinic acid were studied after intravenous, intraperitoneal and oral administration to 1.5–3.0 kg Atlantic halibut, Hippoglossus hippoglossus L., held in sea water at 9 °C. Following intravenous injection, the plasma drug concentration-time profile showed two distinct phases. The terminal elimination half-life was estimated to be 52 h, whereas total body clearance (Cl_T) was determined to be 0.044 L kg^–1 h^–1. The volume of distribution at steady state, V_d(ss), was calculated to be 3.0 L kg^–1, indicating good tissue penetration of oxolinic acid in Atlantic halibut. The peak plasma concentration (C_max) and the time to peak plasma concentration (T_max) were estimated to be 1.2 and 2.7 μg mL^–1, and 21.5 and 80 h, respectively, following oral administration of medicated feed or intraperitoneal injection. The corresponding bioavailabilities were calculated to be 15% and 92%, respectively. Oral administration of vetoquinol, the carbitol ester of oxolinic acid, increased the bioavailability of oxolinic acid to 64% and the total bioavailability (oxolinic acid + vetoquinol) to 82%, whereas C_max and T_max values of 6.7 μg mL^–1 and 14.5 h, respectively, for oxolinic acid, and 1.0 μg mL^–1 and 6.3 h, respectively, for vetoquinol were obtained. Based on a minimum inhibitory concentration (MIC) of 0.0625 μg mL^–1 for susceptible strains, a single intraperitoneal injection of 25 mg kg^–1 of oxolinic acid maintains plasma levels in excess of 0.25 μg mL^–1, corresponding to four times the MIC value, for ≈12 days. The corresponding values for a single oral dose of 25 mg kg^–1 of oxolinic acid and vetoquinol were 5 and 10 days, respectively. For resistant strains with a MIC of 1 μg mL^–1, a single oral dose of vetoquinol (25 mg kg^–1) maintained plasma levels in excess of 4 μg mL^–1 for 34 h.