In vivo and in vitro treatments against Sparicotyle chrysophrii (Monogenea: Microcotylidae) parasitizing the gills of gilthead sea bream (Sparus aurata L.)

The effect of in vitro and in vivo treatments against Sparicotyle chrysophrii, a microcotylid parasite of gilthead sea bream (Sparus aurata L.), was studied. In vitro chemical treatments were targeted to eggs, oncomiracidia and adults, and were tested both as disinfectants and therapeutics for infected animals. The compounds were: distilled water, formalin, limoseptic ®, hydrogen peroxide, chlorine, and praziquantel (PZQ). Larvae were sensitive to all the treatments, but adults were more resistant, as chlorine (60 ppm - 1 h), hydrogen peroxide (100 ppm - 30 min) and PZQ (50 ppm - 30 min) produced only 10% mortality. All adults were killed only with distilled water, limoseptic (0.1% - 5 min), formalin (300 ppm - 30 min), or hydrogen peroxide (200 ppm - 30 min). Eggs were the most resistant stage, as only 30 min in limoseptic (0.1% in distilled water) or in formalin (300 ppm) prevented hatching. PZQ was used in vivo either as a curative or preventive treatment. The highest dose tested (400 mg kg^− 1 BW; effective dose 116.3 mg kg^− 1 BW due to palatability problems leading to 45% reduction in host food intake) did not significantly decrease prevalence of infection when given for 6 consecutive days. A lower dose (200 mg kg^− 1 BW) (effective dose 158.1 mg kg^− 1 BW) was rejected to a lower degree and decreased the prevalence of infection from 90% to 40%. When a lower dose (40 mg kg^− 1 BW) was administered for longer periods (20 days), food intake was reduced slightly, but the infection did not decrease significantly. The oral intubation with PZQ (200 mg kg^− 1 BW) once a week for 4 weeks did not prevent the infection of fish by cohabitation. However, a significant reduction in the abundance of the parasite was registered. In view of the results, recommendations for fish treatment and disinfection of aquaculture facilities are discussed.     

Pharmacokinetic–pharmacodynamic modelling for the determination of optimal dosing regimen of florfenicol in Nile tilapia (Oreochromis niloticus) at different water temperatures and antimicrobial susceptibility levels

Optimized dosing regimen is key to the effective use of antibacterials and to minimizing drug‐related side effects. The current study established a pharmacokinetic–pharmacodynamic (PK‐PD) model for the determination of optimal antibacterial dosing regimen in fish taken into consideration the temperature‐dependent PK and the pathogen‐dependent antimicrobial susceptibility, using florfenicol (FF) in Nile tilapia as an example. The calculated optimal dosages significantly varied by temperature and target MIC levels, ranging from 2.23 (MIC 1 µg/ml at 24°C) to 34.88 mg kg^?1 day^?1 (MIC 4 µg/ml at 32°C). The appropriateness of the calculated dosages was successfully verified by the in vivo studies. After 5 days of oral administration of the calculated optimal dosage at 24°C, the predicted plasma drug values were in line with the mean observed C_min(ss) while at 28 and 32°C underestimation of the C_min(ss) in a dose‐dependent manner was observed and likely due to the occurrence of non‐linear PK at high dosages. The averaged serum protein binding of FF was 19.1%. Our results demonstrated the appropriateness and clinical applicability of the developed PK‐PD approach for the determination of optimal dosing regimens at given temperatures and MICs. Saturation metabolism and PK non‐linearity of FF in tilapia warrant further study.     

A comparison of the palatability of racemic praziquantel and its two enantioseparated isomers in yellowtail kingfish Seriola lalandi (Valenciennes, 1833)

The bitterness of racemic praziquantel (Rac‐PZQ) constrains its use as an in‐feed treatment against monogenean flukes in finfish aquaculture. Evidence exists in mammals that the R‐(?) enantiomer of PZQ is less bitter than the S‐(+) enantiomer. If fish exhibit this same response, then the recently described techniques for the large‐scale resolution of R‐(?)‐PZQ from Rac‐PZQ could facilitate the wide‐spread application of this effective anthelmintic compound via feed. The hypothesis that yellowtail kingfish Seriola lalandi would find R‐(?)‐PZQ more palatable than Rac‐PZQ and S‐(+)‐PZQ was tested in four trials. During the first three trials, the palatability of diets top‐coated with 10 g kg^?1 of Rac‐PZQ or its two enantioseparated isomers were compared in small (85–160 g) and large (1.2 kg) yellowtail kingfish. A fourth trial compared the palatability of R‐(?)‐PZQ and Rac‐PZQ at dietary inclusion levels of 2.5, 5.0 and 10.0 g kg^?1 in small yellowtail kingfish (170 g). Ingestion data showed that R‐(?)‐PZQ to be no more palatable than either Rac‐PZQ or S‐(+)‐PZQ to yellowtail kingfish, regardless of size. Indeed, evidence suggested that the S‐(+)‐PZQ to be slightly more palatable than both R‐(?)‐PZQ and Rac‐PZQ. From these data, we hypothesize that the strong smell of R‐(?)‐PZQ (which was not present in S‐(+)‐PZQ) is an equally important determinant to palatability as taste in yellowtail kingfish. Results demonstrate that dietary inclusion level is a more important determinant to palatability than PZQ chirality; however, administration of R‐(?)‐PZQ may still be advantageous if it is demonstrated to be the only enantiomer efficacious against monogeneans.     

Praziquantel treatment for yellowtail kingfish (<Emphasis Type="Italic">Seriola lalandi</Emphasis>): dose and duration safety study

Regulatory approval is being sought to use praziquantel (PZQ) to treat flukes infecting yellowtail kingfish (YTK), but accurate safety data were not available. We investigated the effect of increased doses or prolonged exposure of orally administered PZQ on YTK by assessing changes in haematological and biochemical characteristics, and mortality. Fish were intubated daily for 3 days with 0, 100, 300 and 500 mg PZQ kg^?1 BW day^?1 or once daily for 9 days at 0 and 100 mg PZQ kg^?1 BW day^?1. Blood was taken 24 h after the cessation of treatment. There was no significant difference between any of the haematological or biochemical indices in YTK treated with PZQ and controls, indicating that PZQ is safe for use at 100 mg PZQ kg^?1 BW day^?1 in YTK and that exposure to high doses or prolonged duration does not have negative effects on the YTK haematological or biochemical parameters we measured.     

Pharmacokinetic model of florfenicol in turbot (Scophthalmus maximus): establishment of optimal dosage and administration in medicated feed

The pharmacokinetics of florfenicol (FF) in turbot (Scophthalmus maximus) was studied after single intravenous (10 mg kg⁻¹) and oral (100 mg kg⁻¹) administration. The plasma concentration–time data of florfenicol were described by an open one‐compartment model. The elimination half‐life (t_1/2) was estimated to be 21.0 h, and the total body clearance, Cl, was determined as 0.028 L kg h⁻¹. The apparent volume distribution (V_d) was calculated to be 0.86 L kg⁻¹ and the mean residence time (MRT_iv) was 30.2 h. Following oral administration, the maximum plasma concentration (C_max) of 55.4 μg mL⁻¹ was reached at 12 h (T_max). The absorption constant (k_a) was 0.158 h⁻¹. The bioavailability was estimated to be 57.1%. The low bioavailability observed at higher doses was explained by the saturation of the mechanisms of absorption. The drug absorption process was limited by its inherent low solubility, which limited the amount of available FF absorbed in the gastrointestinal tract. Based on the pharmacokinetic data, an optimal dosing schedule for FF administration is hereby provided. Based on the minimum inhibitory concentration found for susceptible strains of Aeromonas salmonicida, oral FF administration of first, an initial dose of 30 mg FF kg⁻¹, followed by 6 maintenance doses at 18 mg kg⁻¹/daily could be effective against furunculosis in turbot.     

Effects of Echerichia coli lipopolysaccharides and dissolved ammonia on immune response in southern white shrimp Litopenaeus schmitti

The success of penaeid shrimp aquaculture has been limited by endemic and epidemic infectious diseases around the world and shrimp defense mechanisms are a priority for control, prevention, and diagnosis. The lethal dose (LD₅₀) of Escherichia coli lipopolysaccharides was calculated and pathogen injection and dissolved ammonia concentration on immune response were evaluated in southern white shrimp Litopenaeus schmitti. The lethal dose of lipopolysacharides was 3.78 mg kg^− 1 body weight. Injection caused changes in phenoloxidase activity in plasma and nitric oxide and total haemocyte counts within the first 24 h. High concentration of dissolved ammonia caused a decrease in haemocytes by 66% within the first 72 h, when compared to the control. Hemagglutination in plasma was not affected by injection or high concentrations of dissolved ammonia. The results showed that white shrimp recognized the lipopolysacharides and responded to this microbial elicitor, as indicated by a variety of immunological indicators and that increased dissolved ammonia affected the number of circulating haemocytes.     

Pharmacokinetic disposition of enrofloxacin in brown trout (Salmo trutta fario) after oral and intravenous administrations

In this study, the pharmacokinetic profile of enrofloxacin (EF) and its major metabolite, ciprofloxacin (CF), were investigated in brown trout (Salmo trutta fario) (n = 150) after intravenous (i.v.) and oral (p.o.) administrations of a single dose of 10 mg kg^− 1 body weight (b.w.) at 10 °C. The plasma concentrations of the drugs were determined by high-performance liquid chromatography (HPLC-UV) from 0.08 to 120 h. Pharmacokinetic parameters were described by the two-compartment open model for intravenous and oral administrations, respectively. After intravenous administration, the elimination half-life (t_1/2β), apparent volume of distribution at steady-state (V_ss) and total body clearance (Cl_tot) of enrofloxacin were 19.14 ± 1.51 h, 3.40 ± 0.18 L kg^− 1 and 0.14 ± 0.01 L kg h^− 1, respectively. After oral administration, the maximum plasma concentration (C_max), time of maximum concentration (t_max) and bioavailability (F%) were 2.30 ± 0.08 µg mL^− 1, 8 h and 78 ± 4%, respectively. Ciprofloxacin was not detected in the present study. The elimination half-life for enrofloxacin following oral administration was longer than values calculated for other animals. After oral administration, the mean plasma concentration was well above the minimum inhibitory concentrations (MICs)—that is, > 0.5 µg mL^− 1 at 36 h—for most gram-negative fish pathogens. It is possible and practical to obtain therapeutic blood concentrations of enrofloxacin in brown trout (S. trutta fario) using oral administration of 10 mg kg^− 1 body weight; therefore, it may be effective in the therapy for brown trout diseases.     

Effects of variation in dietary contents of selenium and vitamin E on growth and physiological and haematological responses of yellowtail kingfish,Seriola lalandi

The effects of dietary selenium (Se) and vitamin E and their interaction in the nutrition of yellowtail kingfish, Seriola lalandi, were investigated. Six dietary treatments were prepared in a 3 × 2 factorial arrangement (not supplemented or supplemented with Se at 1 or 2 mg kg^?1 × supplemented with vitamin E at 40 or 180 mg kg^?1). A group of fish in triplicate were fed one of the six experimental diets for 6 weeks, and their growth performance, haematological and immune responses were measured. The results revealed positively interactive effects between dietary Se and vitamin E in yellowtail kingfish. Se significantly increased weight gain of fish fed diets low in vitamin E, but not high in vitamin E. Simultaneous supplementation of both micronutrients resulted in significant increase in serum bactericidal activity. There was no significant effect of Se or vitamin E on survival, feed intake, feed conversion ratio, haematocrit, white blood cell counts and fillet proximate composition. However, Se and vitamin E contents in fillets were significantly responsive to dietary Se and vitamin E, respectively. The supplemental level of Se at 2 mg kg^?1 significantly increased red blood cell counts and haemoglobin concentrations, while lysozyme activity in skin mucus was significantly stimulated by vitamin E. The findings of Se and vitamin E supplementation in this study can be applied to improve growth and health indices of yellowtail kingfish.     

Effect of saponin on hematological and immunological parameters of the giant freshwater prawn, Macrobrachium rosenbergii

Giant freshwater prawns, Macrobrachium rosenbergii (17.9 ± 2.7 g), exposed to different concentrations of saponin at 0, 0.3, 0.6, 0.9 and 1.2 mg l^− 1 for 168 h were examined for osmolality, electrolyte levels, oxyhemocyanin, protein levels, acid-base balance status, total hemocyte count (THC), phenoloxidase activity, and respiratory bursts. Hemolymph oxyhemocyanin, protein, and pO₂ were inversely related to the saponin concentration. Hemolymph oxyhemocyanin, protein, pO₂, pCO₂, and pH of prawns exposed to 1.2 mg l^− 1 saponin were significantly lower than those of prawns exposed to 0.3 mg l^− 1 and control solutions. However, no significant difference was observed in osmolality or electrolyte levels of prawns exposed to different concentrations of saponin for 168 h. The THC of prawns following 168 h of exposure to 0.9 and 1.2 mg l^− 1 saponin increased, but the phenoloxidase activity decreased suggesting that the decrease in phenoloxidase activity under saponin stress was not a consequence of the increase in THC. We concluded that saponin at as low as 0.9 mg l^− 1 decreases the respiratory protein level and acid-base balance, and modulates the immune system of M. rosenbergii.     

Effect of temperature on growth and energy budget of juvenile cobia (Rachycentron canadum)

Growth and energy budget of juvenile cobia (initial body weight ∼ 22 g) at various temperatures (23, 27, 31 and 35 °C) were investigated in this study. Maximal ration level (RL_max, %/day) increased as temperature (T, °C) increased from 23 °C to 31 °C but decreased at 35 °C, described as a quadratic equation: RL_max = −0.023T² + 1.495T − 17.52. Faecal production (f, mg g^− 1 day^− 1) increased with increased temperature (T, °C), described as a power function: lnf = 0.738lnT − 0.806. As temperature increased, feed absorption efficiency in dry weight (FAE_d, %), protein (FAE_p, %) and energy (FAE_e, %) all increased first and then decreased, but the variation of feed absorption efficiency was small, with ranges of 89.59-91.08%, 92.91-94.71%, 93.92-95.32%, respectively. Specific growth rate in wet weight (SGR_w, %/day), dry weight (SGR_d, %/day), protein (SGR_p, %/day) and energy (SGR_e, %/day) showed a domed curve relative to temperature (T, °C), described as quadratic equations: SGR_w = − 0.068T² + 3.878T − 50.53, SGR_d = − 0.079T² + 4.536T −59.64, SGR_p = − 0.084T² + 4.783T − 63.08 and SGR_e = − 0.082T² + 4.654T − 60.99, and SGR_w, SGR_d, SGR_p and SGR_e maximized at 28.5 °C, 28.6 °C, 28.4 °C, 28.5 °C, respectively, as calculated from the regression equations. The relationships between feed conversion efficiency in wet weight (FCE_w, %), dry weight (FCE_d, %), protein (FCE_p, %), energy (FCE_e, %) and temperature (T, °C) also took on a domed curve described as quadratic equations: FCE_w = − 0.726T² + 39.71T − 473.8, FCE_d = − 0.276T² + 15.31T − 190.6, FCE_p = − 0.397T² + 22.05T − 277.9 and FCE_e = − 0.350T² + 19.39T − 239.9, and FCE_w, FCE_d, FCE_p and FCE_e maximized at 27.4 °C, 27.8 °C, 27.7 °C and 27.7 °C, respectively, as calculated from the regression equations. Energy budget of juvenile cobia fed satiation was 100C = 5F + 67(U + R) + 28G at water temperature 27 °C and 100C = 5F + 70(U + R) + 25G at water temperature 31 °C, where C is food energy, F is faeces energy, (U + R) is excretion energy and metabolism energy, and G is growth energy.     

A single‐dose pharmacokinetic study of emamectin benzoate in cod,Gadus morhua L., held in sea water at 9 °C

The pharmacokinetic profile of the antiparasitic agent emamectin benzoate was studied in plasma after intravenous (i.v.) injection and in plasma, muscle and skin following oral (p.o.) administration to cod, Gadus morhua, held in sea water at 9 °C and weighing 100–200 g. Following i.v. injection, the plasma drug concentration‐time profile showed two distinct phases. The plasma distribution half‐life (t_1/2α) was estimated as 2.5 h, the elimination half‐life (t_1/2β) as 216 h, the total body clearance (Cl_T) as 0.0059 L kg^?1 h^?1 and mean residence time (MRT) as 385 h. The volume of distribution at steady state, V_d(ss), was calculated to be 1.839 L kg^?1. Following p.o. administration the peak plasma concentration (C_max) was 15 ng mL^?1, the time to peak plasma concentration (T_max) was 89 h and t_1/2β was 180 h. The highest concentration in muscle (21 ng g^?1) was measured after 7 days and t_1/2β was calculated to be 247 h. For skin, a peak concentration of 28 ng g^?1 at 3 days was observed and a t_1/2β of 235 h was determined. The bioavailability following p.o. administration was calculated to be 38%.     

Effects of moderately oxidized dietary lipid and the role of vitamin E on the development of skeletal abnormalities in juvenile Atlantic halibut (Hippoglossus hippoglossus)

A study was conducted to characterize the effects of oxidized marine fish oil (MFO) on skeletal development in juvenile Atlantic halibut (Hippoglossus hippoglossus) and to determine the role of vitamin E on their bone health and antioxidant defense mechanisms. Juvenile halibut (4.5 ± 0.1 g) were fed six experimental diets containing untreated (peroxide value (POV)  =0.6 meq kg^− 1), mod`rately oxidized (POV = 7.5 meq kg^− 1) and highly oxidized (POV = 15 meq kg^− 1) MFO either with or without α-tocopherol acetate (0 or 300 IU kg^− 1) supplementation for 14 weeks. No significant effects on growth, survival, hepatosomatic indices, or hematocrit were observed among the dietary treatments. Fish fed diets without vitamin E and highly oxidized dietary lipids showed increased hepatic malonaldehyde concentrations indicating a response to oxidative stress. Both muscle and liver α-tocopherol concentrations were significantly lower in fish fed diets without vitamin E supplementation. Alkaline phosphatase levels in serum and bone were increased when vitamin E was present within the diet indicating higher bone formation activity by osteoblasts. Oxidized lipids and lack of dietary vitamin E significantly increased saturated and decreased polyunsaturated hepatic fatty acids. Liver lipids of fish fed diets without vitamin E also exhibited a lower ratio of 22:6n-3 to 22:5n-3 and n-3 fatty acids. The most frequent skeletal deformity observed was scoliosis, spanning the cephalic/prehemal regions, as well as the anterior hemal region of the vertebral column, which increased the frequency according to elevated levels of oxidized dietary lipid. Lordosis was also observed, with no specific pattern along the vertebral column. The pattern and type of abnormalities observed were similar to those reported in an earlier study in halibut from a commercial hatchery.     

Effects of dietary vitamin C on survival, growth, and immunity of large yellow croaker, Pseudosciaena crocea

An 8 weeks growth experiment was conducted to determine the effects of dietary vitamin C on the survival, growth, tissue ascorbic acid contents and immunity of large yellow croaker (Pseudosciaena crocea) with initial weight of 17.82 ± 0.68 g. Seven practical diets were formulated to contain 0.1, 12.2, 23.8, 47.6, 89.7, 188.5 and 489.0 mg ascorbic acid equivalent kg^− 1 diet, supplied as l-ascorbyl-2-polyphosphate (LAPP). Each diet was fed to triplicate groups of fish in seawater floating cages (1.5 × 1.5 × 2.0 m), and each cage was stocked with 120 fish. Fish were fed twice daily (05:00 and 17:00) to apparent satiation for 8 weeks. The water temperature fluctuated from 19.5 to 25.5 °C, the salinity from 25 to 28‰ and dissolved oxygen content was approximately 7 mg l^− 1 during the experimental period. Results showed that the specific growth rate (SGR) (from 1.80 to 1.96% d^− 1) had an increasing trend with the increase of dietary vitamin C, but no significant difference was observed among dietary treatments. No gross deficiency signs were observed in any of the experimental fish. Survival rate, however, significantly increased with increasing dietary vitamin C (P < 0.05). The vitamin C contents in liver and muscle correlated positively with the vitamin C in diets. The vitamin C requirement was estimated to be 28.2 mg kg^− 1 based on survival, and 87.0 mg kg^− 1 on liver content of vitamin C. The activities of serum lysozyme and alternative complement pathway (ACP), phagocytosis percentage (PP) and respiratory burst activity of head kidney significantly increased with increasing dietary vitamin C. The challenge experiment with Vibrio harveyi showed that fish fed the diets with supplementation of vitamin C had significantly lower cumulative mortality compared to the control group (66.7%), and the cumulative mortality (16.7%) in fish with 489.0 mg kg^− 1 ascorbic acid was significantly lower than that (41.7%) in fish with 23.8 mg kg^− 1 ascorbic acid. These results suggested that vitamin C significantly influenced the immune response and disease resistance of large yellow croaker.     

Pharmacokinetics and physio‐metabolic response of single and multiple dose of fenbendazole in Labeo rohita (Hamilton, 1822) fingerlings

An experiment was carried to determine the plasma fenbendazole (FBZ) concentration and physio‐metabolic responses in juveniles of Labeo rohita (90 ± 4 g) after oral administration of single doses at 10, 20 and 50 mg, 20 mg FBZ/kg b.wt. in multiple times on 1st, 3rd and 7th day. The blood samples were collected at 0.5, 1, 2, 4, 8, 12, 24, 30, 48, 72, 96 and 120 hr, after single‐dose administration, and regularly (upto 15 day) in multiple dose. Plasma FBZ concentration was determined up to the limit of detection (LoD) of 0.09 µg/ml by HPLC. There was no parent drug detected in plasma for administration of 10 mg FBZ/kg b.wt. The drug attained the peak concentration (C_max) 1.85 and 3.09 µg/ml in plasma at 4 hr (T_max) after administration of 20 and 50mg FBZ/kg b.wt. respectively. Plasma FBZ was detectable up to 96 and 120 hr with concentration 0.09 ± 0.007 and 0.098 ± 0.006 µg/ml, respectively, after single‐dose administration of 20 and 50mg/kg b.wt. In case of multiple‐dose administration, the maximum concentration of FBZ was 1.01 ± 0.03 µg/ml on 7th day that was less than to the single dose at 50 mg/kg b.wt. However, FBZ was detected up to 11 day after multiple doses. The study revealed that the hepatic antioxidant enzymes activities like superoxide dismutase, catalase and glutathione‐S‐transferase were significantly affected by increasing FBZ in single and multiple doses. The results of the present study could reveal that single‐ or multiple‐oral administration of FBZ at 20 mg/kg b.wt. in feed as antihelminthic drug in L. rohita could be considered as the safe dose.     

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.     

Assessment of nitrogen maintenance requirement and potential for protein deposition in juvenile Tilapia genotypes by application of an exponential nitrogen utilization model

Assessment of quantitative amino acid requirement data in fish nutrition is predominantly based on dose response studies with supplementation of the dietary limiting amino acid. Studies with modeling amino acid requirements are scarce. Three experiments were conducted with different genotypes of juvenile Tilapia [A: all male O. niloticus; B: red Tilpia (O. mossambicus × O. hornorum); C: hybrid Tilapia (A × B)] using fishmeal-wheat gluten diets with five graded levels of dietary protein supply (17.2-48.5% CP). The dietary amino acid ratio was similar, dietary energy contents were adapted to avoid limiting energy supply. Growth response was studied in a semi-closed in-door water recirculation system with 38 tanks (320 l/tank; water temperature 28 ± 0.5 °C; regulated photoperiod 12 h light/12 h dark). Four replicate groups (25 fish per tank) were utilized in a 70 d growth experiment with three times a day feeding until apparent satiation. Ten fish at the beginning and three fish per tank at the end of the growth study were analyzed for body composition to calculate N-deposition and N-excretion. The nitrogen maintenance requirement (NMR) was established based on nonlinear fitting of N-excretion depending on N-intake. The intersection of the applied exponential function with the y-axis was used as NMR, simulating protein free feeding. The theoretical potential for daily N-deposition of the genotypes under study (ND_maxT) was estimated as threshold value due to exponential fitting of N-deposition depending on N-intake. The observed average NMR (70 mg N/BW_kg^0.67/d) and threshold values for genotype A (ND_maxT = 388 mg/BW_kg^0.67/d), genotype B (ND_maxT = 389 mg/BW_kg^0.67/d) and genotype C (ND_maxT = 319 mg/BW_kg^0.67/d), respectively, are the starting point for the studied exponential N-utilization model and its subsequent application for assessing amino acid requirements depending on growth performance and dietary amino acid efficiency in fish feed.     

The effect of density on sea bass (Dicentrarchus labrax) performance in a tank-based recirculating system

Sea bass (Dicentrarchus labrax) (135 ± 4 g) were reared under tank-based recirculating aquaculture system for a 63-day period at four densities: 10, 40, 70, 100 kg m^?3. Fish performance, stress indicators (plasma cortisol, proteonemia plus other blood parameters—Na⁺, K⁺, glucose, pH, total CO₂^?) and water quality were monitored. At the end of the 63-day period, resistance to infection was also studied by a nodavirus challenge. A 25-day test was performed on fish from two extreme densities (10 and 100 kg m³) and one intermediate density (40 kg m³).With regards to the different density treatments, there was no significant difference between the daily feed intake (DFI) and the specific growth rate (SGR) up to a density of 70 kg m^?3. No significant difference was found between treatments concerning the feed conversion ratio (FCR) and the mortality rate. No density effect was observed on the fish stress level (plasma cortisol) or on sensitivity to the nodavirus challenge. Under these experimental rearing conditions, the density above 70 kg m^?3 has an impact on growth performance (DFI and SGR) indicators and also some blood parameters (CO₂) at the highest density tested (100 kg m^?3).This study suggests that a density up to 70 kg m^?3 has no influence on sea bass performance and welfare. At 100 kg m^?3, average specific growth rate was decreased by 14% without welfare deterioration according to the welfare indicators monitored.     

Development of lipid microbeads for delivery of lipid and water-soluble materials to Artemia

Lipid spray beads (LSB) containing high concentrations of phospholipids were produced in order to improve their dispersion in both fresh and saltwater. The beads were developed to deliver both fat-soluble and water-soluble micronutrients to Artemia and other suspension feeders. LSB were prepared by spraying molted lipid into a chamber that was cooled with liquid nitrogen in order to solidify the lipid beads. Addition of soy lecithin to LSB did not affect retention of glycine when the beads were suspended in distilled water. There was an initial loss of 80% incorporated glycine after LSB were suspended in water for 20min. Artemia readily ingested riboflavin-containing LSB and their full guts were evident within 30min of feeding. The riboflavin content of Artemia could be increased from 55 ±  0.6 mg kg^− 1 (dw) to 329 ± 62 mg kg^− 1 (dw) after 1h enrichment. LSB prepared with phospholipids are promising vehicles for enrichment of suspension-feeding organisms used as feed for larval marine fish and crustaceans as well as other suspension feeders.     

Water flow and diurnal variation in metabolite production rates of Atlantic salmon (Salmo salar L.) post‐smolt

Diurnal variation of carbon dioxide (CO₂) and total ammonia production in Atlantic salmon post‐smolt were studied at different water flow rates. The experiment comprised four groups each with two replicates representing specific flow levels of 0.5, 0.4, 0.3 and 0.2 L kg^?1 min^?1. During the first diurnal cycle, the seawater samples were collected eight times during 21 hr. In the second diurnal cycle, six samplings were performed during a prolonged sampling period of 35 hr. The highest CO₂ concentrations were observed in the lowest water flow group (0.2 L kg^?1 min^?1) between 4 and 10 p.m. for the first sampling period and at about 2 p.m. for the second sampling period. The overall real CO₂ production rates were in the range 1.7–5.5 mg kg^?1 min^?1 including diurnal variation in all groups for both sampling periods. In general, a second‐order polynomial model describes the relationship between specific water flow and real mean CO₂ production rate (p < 0.001). Maximum concentrations of total ammonia nitrogen (TAN) occurred around 7 and 8 p.m. in all groups and was highest for the 0.2 L kg^?1 min^?1 group for both sampling periods. Note that maximum concentration of TAN and CO₂ both occurred in the afternoon. The TAN production rates were in the range 0.06–0.44 mg kg^?1 min^?1 including both sampling cycles.     

Oxytetracycline (OTC) accumulation and elimination in hemolymph, muscle and hepatopancreas of white shrimp Litopenaeus vannamei following an OTC-feed therapeutic treatment

The white shrimp, Litopenaeus vannamei, has become a very important species for the development of shrimp aquaculture in Northwest Mexico. However, viral and bacterial diseases are considered a major threat to the development of this industry. In the present study a trial was conducted to evaluate the tissue distribution, maximum concentration, and elimination of the widely used antibiotic oxytetracycline (OTC) in L. vannamei using indoor tanks under laboratory-controlled conditions. OTC was given to shrimp simulating a therapeutic treatment through medicated feed for 14 days followed by a period of feeding without antibiotic for another 14 days to evaluate the elimination pattern. Samples of hemolymph, muscle, and hepatopancreas were taken from medicated animals every two days for 28 days. All tissues were removed and frozen immediately in liquid nitrogen. OTC levels were analyzed by High Performance Liquid Chromatography (HPLC). Results showed an important OTC increase during consumption of medicated feed in all examined tissues. OTC maximum concentrations were 33.54 ± 11.19, 194.37 ± 16.11, and 18.79 ± 5.87 µg g^− 1 for muscle, hepatopancreas and hemolymph, respectively. Although the highest OTC level was found in the hepatopancreas, it required only two days after the start of dosing to reach this value, whereas the maximum OTC for muscle and hemolymph was detected after eight days of dosing. Ten days after the cessation of medicated feeding, the drug content in the shrimp tail muscle was under the detectable limit for the method (0.01 µg g^− 1 of OTC).