Effects of Carbon Dioxide Exposure on Intensively Cultured Rainbow Trout Oncorhynchus mykiss: Physiological Responses and Fillet Attributes

Rainbow trout Oncorhynchus mykiss (261.6 × 24.7 g initial weight, mean × SEM) at 13.1 × 0.2 C were exposed for 94 d to one of three CO₂ treatments: control (22.1 × 2.8 mg/L), medium (34.5 × 3.8 mg/L), or high (48.7 × 4.4 mg/L). Trout were checked daily for survival, and fish were sampled at 0, 28, 56, and 84 d for physiological responses, growth, and fillet quality assessments. Trout were also challenged to a 15-min crowding stress at 93 d to assess their ability to initiate a stress response during hypercapnia. Chronically exposed trout showed nearly 100% survival through 84 d exposure (1 of 1,500 fish died). Growth and physiological results showed that increasing elevated CO₂, concentrations result in corresponding decreased growth rates and CO₂specific physiological parameters: The medium and high CO₂ treatments had significantly slower growth and subsequently smaller fish by 84 d. Exposed trout also showed significantly ( P < 0.05) decreased plasma chloride for medium and high CO₂ treatments compared to the control from 28 through 84 d. Decreased growth and smaller fish in the medium and high CO₂ treatments resulted in correspondingly smaller fresh and smoked fillet weights. Chronic CO₂ exposure did not result in notable changes in ultimate muscle pH. Exposure to 15-min crowding stress at 93 d resulted in significant changes in hematocrit, plasma cortisoI, glucose, and chloride for all treatment groups. CO₂-specific changes were detected in hematocrit, plasma cortisoI, and plasma chloride responses following the 15-min crowding stress.     

14C-glucose injection in Atlantic cod, Gadus morhua, metabolic responses and excretion via the gill membrane

In the present study cod were separately placed in fish metabolic chambers. Ten individuals were injected with a dose of 1 μCi ¹⁴C-glucose, in addition to 1 g pure glucose per kg live weight, and 10 individuals were kept under the same conditions and sham injected with 9 g L⁻¹ (0.9%) sterile saline solution.
Of the injected ¹⁴C-glucose, 3% was recovered in plasma, 2.9% in liver, 1.7% in red muscle, 18.2% in white muscle, 0.2% in heart, 1.9% in gills and 12.1% in gill-surrounding water. Of the 12.1% in gill-surrounding water, 9.1% was found to be in the form of CO₂, leaving 3% to pure glucose and lactate. These results indicate a relatively high production of CO₂ from glucose showing a utilization for energy, and a possible route of excretion of excess glucose via the gill membrane.
Activity of ¹⁴C-glucose on a concentration basis showed in descending order the following organs to be metabolically active in glucose utilization: gills > plasma (as a transporter) > heart > red muscle > liver > white muscle.
Of the total amount of ¹⁴C-glucose injected, only 0.3% was recovered in the liver lipid fraction, and mainly as triacylglyceroles; minor ¹⁴C-activity was found in mono- and diacylglyceroles and free fatty acids. These results show very low activity of de novo syntheses of lipid from injected glucose in the cod.     

Pond Bottom Soil Respiration during Fallow and Culture Periods in Heavily-Fertilized Tropical Fish Ponds

Benthic respiration in two fish culture ponds at Comayagua, Honduras, was below 1g CO₂/m² per day during culture periods. When ponds were drained for fish harvest, bottom soils were exposed to the air, and soil respiration rates as high as 10g CO₂/m² per day were recorded. High rates of soil respiration following removal of saturated conditions declined as labile organic matter was consumed and soil moisture content declined. Laboratory studies showed that the moisture content greatest for soil respiration was near saturation, and either drying soils or completely saturating them drastically reduced respiration. Although soil respiration rates were much greater during the fallow period than during the culture period, more organic matter was decomposed during the culture period because it was much longer (5 months) than the fallow period (2 weeks).     

Tissue distribution of vitamin D3 in Atlantic salmon Salmo salar: effect of dietary level

Atlantic salmon Salmo salar were given three dietary doses of vitamin D₃ for a period of 11 weeks: diet groups I, II and III received a 0.04, 2.21 and 28.68 μg g⁻¹ diet of vitamin D₃, respectively. The tissue distribution of vitamin D₃ was investigated, and between diet groups II and III, the level of vitamin D₃ found in the tissues was increased by a factor similar to the increased level in the feed. No effects were observed on weight gain, survival, plasma level of calcium, red blood cell count or haematocrit, relative to the dietary levels of vitamin D₃. The results showed that while the plasma concentration of 25(OH)D₃ increased between diet groups II and III, the concentration of 1,25(OH)₂D₃ decreased.     

Sensitivity of juvenile lobsters, Homarus americanus L., to different concentrations of malachite green during various states of the intermoult cycle

Abstract. Survival of the early stages (egg, larva and juvenile) of the American lobster, Homarus americanus L., reared under controlled conditions has often been threatened by a variety of fungi and microbial epibionts. A preliminary study was conducted for the purpose of defining toxicity levels of malachite green for 5th stage juveniles during each of the following intermoult (or fused intermoult) states: A-B, C₄, D₀ and D₂-D₃ Irrespective of exposure time or concentration, all the lobsters treated during intermoult state A-B died. This contrasts with seemingly good survival of lobsters in intermoult states C₄, D₀ and D₂- D₃ treated at concentrations of 900 mg/1 malachite green for exposure periods of 10 min. The findings suggest that an effective treatment combination for lobsters immersed in malachite green may be one which is employed during intermoult state C₄, for about 6min and at concentrations of about 700–800 mg/l.     

Effects of Aeration, Alum Treatment, Liming, and Organic Matter Application on Phosphorus Exchange between Soil and Water in Aquaculture Ponds at Auburn, Alabama

Application of readily-oxidizable organic substrate to laboratory soil-water systems and fish ponds caused anaerobic conditions in bottom soil and water, and concentrations of soluble reactive phosphorus (SRP) increased. Aeration of ponds increased total phosphorus (TP) concentrations by suspending soil particles in the water, but SRP concentrations declined because of increased oxy- genation of bottom water and soil, Alum [Al₂(SO₄)₃·14H₂O] treatment of ponds reduced SRP and TP concentrations in ponds, but the low concentration of alum used, 20 mg/L, had little residual effect on phosphorus concentration. Application of agricultural limestone at 0.2 kg/m² to ponds with soil pH of 5.5 and Ca²⁺ concentration of 5 mg/L did not affect SRP and TP concentration. Unless pond soils were anaerobic at their surfaces, a condition not acceptable in thermally-unstratified fish ponds, soils released little phosphorus to the water. Strong adsorption of phosphorus by soils in intensive ponds with feeding is beneficial, because removal of phosphorus by aerobic soils is a control on excessive phytoplankton growth. In fertilized ponds, phosphorus must be applied at frequent intervals to replace phosphorus removed from the water by soils.     

Metabolic costs of changing the cation–anion difference in the diet of juvenile African catfish Clarias gariepinus (Burchell)

The influence of dietary cation–anion difference (CAD, Na⁺ + K⁺– Cl^–, mEq kg^–1) on energy metabolism and nitrogen losses in juvenile African catfish Clarias gariepinus (Burchell) was examined in fish exposed to different dietary CAD levels (–146, 116, 497, 713 and 828 mEq kg^–1 diet). The experiment was conducted in open circuit balance respiration chambers over a 3-week period. Five 24-h monitoring periods were carried out at 3-day intervals during the experimental period with O₂ consumption, ammonia and nitrate + nitrite (NO_x) and CO₂ production being measured at 5-min intervals for each chamber. The negative dietary CAD (–146 mEq kg^–1) resulted in the highest energy expenditures (83 kJ kg^–0.8· d^–1). With increasing dietary CAD levels, heat loss gradually decreased to minimum values of 56 kJ kg^–0.8 day^–1 at a dietary CAD level of 713 mEq kg^–1. Consequently, metabolizable energy utilization efficiency (MEU, percentage of retained energy over metabolizable energy) quadratically ( P  < 0.05) increased and reached a maximum at a dietary CAD of 713 mEq kg^–1.     

Pharmacokinetics of Dietary 13C-Labeled Docosahexaenoic Acid and Docosapentaenoic Acid in Red Sea Bream Chrysophrys major

The objectives of this study were to investigate: 1) the pharmacokinetics of dietary docosahexaenoic acid (DHA) and docosapentaenoic acid (DPA) using ¹³C-labeled fatty acids; 2) the interorgan transport of DHA in the red sea bream by monitoring the DHA level of several organs; and 3) the relationship between the plasma DHA level and optimum dietary DHA level in the plasma of the red sea bream Chrysophrys major . For this purpose, a mixture of 38.5% of [¹³C]DHA, 8.5% of [¹³C]DPA, and 4.2% of [¹³C]palmitic acid were given to the red sea bream at dose level of 8.0, 16.0, and 47.9 mg/kg by a single oral administration. For [¹³C]DHA, the maximum plasma concentration (t_max) occurred at 2.00–3.00 h after the oral administration. The peak plasma concentration (C_max) and the area under the plasma concentration-time curve to 24 h (AUC_0-24 for [¹³C]DHA level linearly increased with respect to dosage. [¹³C]DHA appeared in each organ (plasma, erythrocyte and the fat body of the orbit, liver, intestine, skin, brain, heart and muscle) at 0.5 h and was observed until 24 h. From the values determined for the pharmacokinetic parameters, the range of the effective plasma DHA level for normal growth of the red sea bream was suggested to be between 21.0 and 40.3 μg/mL. For [¹³C]DPA, the AUC_0-24 and C_max values also linearly increased with the dosage, but t_max did not depend on it.     

In vitro hydroxylation of vitamin D3 and 25-hydroxy vitamin D3 in tissues of Atlantic salmon Salmo salar, Atlantic mackerel Scomber scombrus, Atlantic halibut Hippoglossus hippoglossus and Atlantic cod Gadus morhua

The in vitro metabolism of ¹⁴CD₃ and ³H25OHD₃ was investigated in different tissues from Atlantic salmon Salmo salar , Atlantic mackerel Scomber scombrus , Atlantic halibut Hippoglossus hippoglossus and Atlantic cod Gadus morhua . The tissues analysed were liver, kidney, head kidney, gills, spleen and intestine. The metabolites were extracted in methanol–chloroform and separated by normal-phase high-pressure liquid chromatography (HPLC) followed by scintillation counting. Identification of the metabolites was by comigration with standards on normal and reversed-phase HPLC systems and by protein-binding assays. All tissues from all species analysed produced hydroxylated derivatives identified as 25OHD₃, 24,25(OH)₂D₃ and 1,25(OH)₂D₃. In addition, some unidentified derivatives were recorded, one probably being 25,26(OH)₂D₃. Organs producing great amounts of one metabolite also produced considerable amounts of the other possible derivatives, suggesting a lower degree of specificity in fish organs than in human organs. The predominating metabolite was 24,25(OH)₂D₃ in all organs from salmon and mackerel during incubation with ¹⁴CD₃ and within most organs from all species during ³H25OHD₃ incubation. The latter observation probably results from the need for decreasing rather than increasing the calcium absorption in these species, which live at least some periods of life in a marine environment.     

Purification and characterization of transglutaminase from squid gill

ABSTRACT: The present study used squid gill as a source of transglutaminase (TGase) because it has extremely high TGase activity compared with other tissues. The enzyme was purified using successive chromatographies of Sephacryl S-300 and hydroxyapatite columns. The yield and purification-fold of the enzymatic activity was 12.6% and 14.1-fold, respectively. The molecular mass of the purified enzyme was estimated to be 94 kDa by using sodium dodecylsulfate–polyacrylamide gel electrophoresis analysis. Enzyme activity was enhanced 15-fold with an increase in NaCl concentration. Although the activity was dependent on Ca²⁺ concentration, it was not sufficiently activated even by 50 mM CaCl₂ in the absence of NaCl, but could be fully activated with 10 mM CaCl₂ in 0.7 M NaCl. However, in the absence of substrates, the enzyme was rapidly inactivated. The pH and temperature optima of the enzyme were approximately pH 8.0 and 20°C, respectively. It was stable in the absence of Ca²⁺ at pH 7.5–9.0 and had a rate constant (K _D ) of 1.6 × 10^–5 s^–1 for thermal inactivation at 50°C. These results in which squid gill TGase could be activated at higher concentrations of Ca²⁺ and NaCl than at a physiological concentration, suggest that contact with seawater or body fluid seems to activate the enzyme if the tissue is disrupted.     

Improved Seed Production of Goldstriped Amberjack Seriola lalandi under Hatchery Conditions by Injection of Triiodothyronine (T3) to Broodstock Fish

Survival of the seed of goldstriped amberjack Seriola lalandi was consistently enhanced by intramuscular injection of triiodothyronine (T₃) to the broodstock fish at a dose of 20 mg/kg body weight. Survival rates at the time of transfer to net cages (35 d after hatching) were 2.6–5.0% for the T₃ treatment while only 0.3–2.1% for the control. Survival rates 36 mo after hatching was estimated to be 12 times higher in the T₃-treated group (1.1%) than in the control (0.09%). T₃ concentration in newly spawned eggs increased from control levels of 0.3–1.0 ng/g to 68–127 ng/g 2 d after injection and subsided with successive spawns. T₃ concentration in the treated larvae was about 360 times higher than in the control upon hatching (29 np/p vs 0.08 ng/g), but became indistinguishable from the control 6 d after hatching. These results indicate injection of T₃ to the broodstock of S. lalndi to be of practical value in aquaculture of the species.     

Effect of various medium compositions on survival and hatching rates of embryos of the giant freshwater prawn Macrobrachium rosenbergii cultured in vitro

SUMMARY: The effectiveness of in vitro embryo culture of the giant freshwater prawn Macrobrachium rosenbergii depended on the age of the embryos at the onset of culture and on the concentrations of various compositions in the medium. Embryos that started being cultured on day 0.5 after oviposition were more sensitive to variations in the medium compositions than those that started being cultured on day 10.5 after oviposition. An optimal NaCl level was essential for embryonic development, survival, hatching and survival of the newly hatched larvae. Variations of NaCl or KCl levels dramatically altered embryonic development, and variation of the MgCl₂ + MgSO₄ level significantly lowered survival of the embryos that started being cultured at the early stage of development. In contrast, no significant change in embryonic development was observed upon variation of the CaCl₂ level. Hatching of the embryos required the presence of NaCl and CaCl₂ but not KCl or MgCl₂ + MgSO₄. The ionic requirements of the newly hatched larvae differed from that of the developing embryos. Variations of NaCl, KCl or CaCl₂ but not MgCl₂ + MgSO₄ levels significantly influenced the survival of the newly hatched larvae.     

Production of salmonid amoebic gill disease by exposure to Paramoeba sp. harvested from the gills of infected fish

Atlantic salmon, S almo salar L., were exposed to different concentrations of Paramoeba sp. The lowest concentration which induced amoebic gill disease (AGD) was 230 Paramoeba sp. L^–1 and the severity of infection increased with increasing concentration. The concentration of Paramoeba sp. positively correlated with the number of gill lesions ( R ² > 0.7). This study provides evidence that Paramoeba sp. is the causative agent of AGD and describes an experimental model that enables the severity of the induced disease to be controlled.     

Role of prostaglandin in the control of ovulation in the Japaneseeel Anguilla japonica

ABSTRACT:   The in vitro effects of 17,20β-dihydroxy-4-pregnen-3-one(DHP) and prostaglandins (PGE₁, PGE₂, PGF_1α,PGF_2α) on ovulation in the Japanese eel Anguillajaponica were examined. Oocytes with follicle layers at themigratory nucleus stage (approximately 850–900 µmdiameter) were removed using a polyethylene cannula from artificiallymatured fish. At concentrations of 10 and 100 ng/mL,DHP was found to induce both germinal vesicle breakdown and ovulation.The prostaglandins, except for PGE₁, effectively inducedovulation of previously matured oocytes by DHP treatment in vitro .Prostaglandin F_2α was the most effective. Asignificant increase in ovulation rate was observed even at a concentrationof 0.01 µg/mL PGF_2α.Indomethacin blocked the in vitro ovulation induced by DHPand addition of PGF_2α reversed indomethacin-blockedovulation. Actinomycin D and cycloheximide blocked DHP-induced ovulationand PGF_2α reversed the effects of both inhibitors. Theseresults indicate that DHP induces ovulation through endogenous prostaglandinsynthesis in the follicle layers of the Japanese eel.     

Haematological responses in the European eel, Anguilla anguilla L., to sublethal infestation by Anguillicola crassus in a thermal effluent of the Swedish Baltic

Abstract. Haematological variations were studied in a European eel population, Anguilla anguilla L., naturally infested by the sanguivorous nematode Anguillicola crassus. This occurred off the Swedish Baltic coast in an area receiving heated cooling water from a nuclear power station. The erythrocyte count, haematocrit, leucocrit, haemoglobin concentration, mean corpuscular volume, mean corpuscular haemoglobin and mean corpuscular haemoglobin concentrations, the various cell types in peripheral blood, serum iron concentration, the amount of total serum protein, and scrum protein fractions α ₁ , β₁ , α ₂, β ₂ and γ were all measured. Each variable was correlated to: (1) the intensity of infestation, i.e. number of parasites per host; and (2) a parasitization index, i.e. the weight of parasites per somatic weight of the host. Most variables showed no or only minor reactions to the infestation. However, a marked increase in the γ -fraction of scrum proteins, reduced lymphocyte numbers and increased granulocyte numbers are indicative of a humoral and cellular immune response.     

Digestibility in goldfish fed diets with and without chromic oxide and exposed to sublethal concentrations of cadmium

Effects of three sublethal levels of cadmium (40, 120 and 260 μg L⁻¹) on apparent total dry matter (TDM) and apparent protein (PD) digestibility in goldfish, Carassius auratus (L.), over an 8-week experimental period were investigated using diets with and without chromic oxide (0.5%) marker. The %TDM and %PD were estimated based on crude fibre as the marker for the diet without Cr₂O₃, and Cr₂O₃ and crude fibre as markers for the diet with Cr₂O₃. Generally, digestibility estimations were lower in goldfish fed the diet with the Cr₂O₃ marker. Similarly, for the diet with Cr₂O₃ marker, digestibility estimates based on crude fibre were higher than those based on Cr₂O₃. In all treatments (2 diets × control and 3 cadmium levels), the weekly estimations of %TDM and %PD did not differ significantly ( P > 0.05). The effect of cadmium on weekly %TDM and %PD was not that obvious in all treatments. However, the overall mean %TDM and %PD differed significantly ( P < 0.05) between diets and between markers, at all cadmium concentrations. The present study raises doubts regarding the validity of the use of Cr₂O₃ as a marker in digestibility studies in fish, as had been recently suggested by other authors but for different reasons.     

Toxicity of Ammonia and Nitrite to Penueus monodon Juveniles

Pcnaeus monodon juveniles (35.4 ± 2.2 mm TL) were exposed to seawater (20 ppt) having different concentrations of total ammonia (NH₃+ NH,+) and nitrite. Median lethal concentration (LC₅₀) was determined with static-renewal tests. The LC₅₀, of total ammonia-N, NH₃-N and niMte-N on shrimps decreased with increase of exposure time. The 24, 48, 72, % and 120 h LC₅₀ were 94.96, 61.09, 47.47 45.58 and 38.00 mg/L total ammonia-N (2.68, 1.73, 135, 1.29 and 1.08 mg/L NH₃-N) and 215.85, 185.33, 88.54, 54.76 and 37.97 mg/L nitrite-N, respectively. The 144h LC₅₀ of total ammonia-N and NH₃-N was 36.71 mg/L and 1.04 mgL. The "threshold" of ammonia and nitrite was found at 144h and 120h respectively, from the toxicity curve approaching asymptote. A "safe value" was 3.7 mg/L total ammonia-N, 0.1 mg/L NH₃-N (20 ppt, pH 7.70, 27C) and 3.8 mg/L nitrite-N for P. monodon juvenile .     

Hemolymph Oxyhemocyanin and Protein Levels and Acid-Base Balance in the Tiger Shrimp Penaeus monodon Exposed to Copper Sulfate

Abstract.— Subadult tiger shrimp Penaeus monodon (30.15 ± 2.19 g) were exposed individually in sea water of 25%o to 0 (control), 1.11, 4.76, 10.06 and 19.09 mg/L copper for 24 h. Hemolymph pH, PO₂ (partial oxygen pressure), PCO₂ (partial carbon dioxide pressure), HCO₃, oxyhemocyanin and protein levels were determined. Hemolymph PO₂ increased, whereas hemolymph pH, PCO₂, HC0₃-, oxyhemocyanin. protein and the ratio of oxyhemocyanidprotein levels decreased with increasing concentrations of ambient copper in the range of 1.11 to 19.09 ma/L. It is concluded that P. monodon following 24-h exposure to ambient copper as low as 4.76 mg/L shows a reduction of oxyhemocyanin, protein, and the ratio of oxyhemocyanin to protein, and develops metabolic acidosis in the hemolymph.     

Effect of Exposure to Potassium Permanganate on Stress Indicators in Channel Catfish Ictalurus punctatus

Juvenile channel catfish Ictulurus punctatus were exposed to 1× (0.44 mg/L), 3× (1.32 mg/L), or 5× (2.19 mg/L) the recommended therapeutic concentrations of waterborne potassium permanganate (KMnO₄) for 36 h to determine the toxicity of the chemical. The fish were observed for 14 d after exposure. Gill, liver, and blood samples were collected before exposure, at 12, 24, and 36 h of exposure, and at 48-h intervals for 14 d thereafter. Analysis of homogenized gill tissue showed a transient increase in manganese content that quickly disappeared once exposure was discontinued. Fish exposed to the 3× and 5× concentrations of KMnO₄ experienced 9 and 50.6% mortality, respectively. Plasma cortisol was elevated more than ten-fold at the 5× concentration. Both plasma chloride and osmolality were significantly reduced at the 3× and 5× concentrations but were unchanged at the 1×. Packed cell volumes (PCV) of whole blood rose significantly in response to 3× and 5× concentrations of KMnO₄ Mortality may have been the result of blood electrolyte depletion as indicated by increased PCVs, loss of chloride, and reduced osmolality. All stress indicators measured, except PCV at the 5× concentration, were indistinguishable from unexposed controls within 48 h after exposure was discontinued. At the l× concentration (the concentration most like that employed in a disease treatment) no changes were observed in any stress indicators measured suggesting that KMnO₄ may be safely used as a disease therapeutant for channel catfish.     

Effects of hydrogen peroxide on clearance of formalin-killed Flavobacterium branchiophilum from the gills of rainbow trout, Oncorhynchus mykiss (Walbaum)

The effect of hydrogen peroxide (H₂O₂) treatment on the clearance of formalin-killed Flavobacterium branchiophilum from the gills of rainbow trout was examined. In untreated control fish, clearance was characterized by a rapid initial phase, with 50% reduction in bacterial antigen in the first 12 h after exposure. The bacteria then cleared more gradually, with total clearance being achieved by 40 h. Treatment of fish with 100 p.p.m. H₂O₂ did not influence bacterial clearance compared to untreated controls. Exposure for 1 h to H₂O₂ concentrations ≤ 100 p.p.m. caused no detectable clinical signs or evidence of ultrastructural damage to the respiratory epithelium. However, levels in excess of this caused significant gill damage.