The use of luteinizing hormone releasing hormone analogue for ovulation induction in black sea bass (Centropristis striata)

Interest in commercial production of black sea bass has increased in recent years, but reliable spawning methods remain problematic. The objective of this study was to evaluate the effects of oocyte size and luteinizing hormone releasing hormone analogue (LHRHa) dosage and delivery systems on ovulatory success for in vitro fertilization. Vitellogenic females with maximum oocyte diameters of 400–625 μm were implanted with a 95% cholesterol–5% cellulose pellet containing 50 μg of LHRHa. Fish with maximum oocyte diameters < 450 μm failed to ovulate. In contrast, 90% of fish with 500 μm oocytes spawned within 36 h and 40% of this group ovulated a second time. All of the females containing oocytes > 550 μm ovulated. In a second experiment, females with uniformly vitellogenic oocytes (> 500 μm) and implanted with 50 μg LHRHa ovulated substantial numbers of eggs (45,000–192,000 eggs/kg body weight (BW), but fertility was consistently low (0–15%). In a third experiment, 19 of 39 females receiving implants containing 6.3–23.6 μg LHRHa/kg BW during the spawning season ovulated, but fecundity (17,000–339,000 eggs/kg) and fertilization (0–98%) were highly variable. When fish were grouped by developmental index, calculated as the number of oocytes with diameters > 400 μm/total number of oocytes measured, there were no statistical differences among groups with respect to the number of spawns, fecundity or fertilization success. In a fourth experiment, 11 of 13 females with a clutch of fully vitellogenic oocytes that were injected with 20 or 100 μg/kg BW LHRHa ovulated between 1 and 2 times on consecutive days. Five of seven given an implant containing 12.5 μg LHRHa ovulated one or more times. Fish implanted with shams or injected with vehicle alone did not ovulate in any of the experiments. No differences were found in the number of spawns, fecundity or fertilization success from fish receiving different doses of injected or implanted LHRHa. Incubation of pooled eggs produced 155,000 larvae (60% hatch) and 95,000 one-gram juveniles. These results demonstrate that injected or implanted LHRHa is effective for inducing ovulation in black sea bass with maximum oocyte diameters > 500 μm.     

Light intensity effects on early life stages of black sea bass, Centropristis striata (Linnaeus 1758)

The effects of four light intensities on growth and survival of first‐feeding stage black sea bass larvae Centropristis striata were investigated in a controlled‐environment laboratory. Fertilized eggs, obtained from LHRHa‐induced spawning of captive broodstock, were stocked (72 eggs L^?1) into twenty 15 L black tanks under light intensities of 100, 500, 1000 and 1500 lx, with five replicate tanks per treatment. The photoperiod was 12L:12D, the temperature was 20°C and the salinity was 35 g L^?1. Larvae were fed rotifers Brachionus rotundiformis from day 2 post‐hatching (d 2ph) at 5–10 rotifers mL^?1. Microalgae Nannochloropis oculata and Isochrysis sp. were added (1:1) daily to maintain a density of 300 000 cells mL^?1. Hatching success and larval growth and survival from d 2ph through d 15ph were monitored. Hatching success was 28–38% under all light intensities, and notochord length at hatching ranged from 2.8 to 3.0 mm, with no significant differences among treatments. By d 15ph, growth (mg wet weight) was significantly higher in the 1000 lx (0.914) and 1500 lx treatments (0.892) than in 100 lx (0.483), and a highly significant trend (P<0.01) towards increased survival with increasing light intensities was observed, from 1.3% at 100 lx to 13.9% at 1500 lx. Higher light intensities within the range of 100–1500 lx improved growth and survival of early larval black sea bass, suggesting that even higher light intensities may improve culture performance. This is consistent with conditions in shallow, near‐shore locations where eggs and larvae are distributed in nature.     

Production and storage of sperm from the black sea bass Centropristis striata L

Black sea bass Centropristis striata L. are protogynous hermaphrodites that develop and spawn as females before changing sex to male. Since all fish eventually become males, determining the relationship between sperm production, sperm quality and seasonal changes in plasma levels of testosterone (T) and 11‐ketotestosterone (11‐KT) could be useful for identifying appropriate males to maintain as broodstock. Milt and blood samples were collected three times during an 8‐week spawning season. Milt volume (3.5±0.76 mL kg^?1), sperm density (3.2 × 10⁸± 0.31 cells mL^?1), sperm production [11 × 10⁸±3.4 cells kg^?1 body weight (BW)] and sperm motility (80±0.6%) were at their highest during the first sampling interval and coincided with the highest 11‐KT levels (1.0± 0.11 ng mL^?1). All of the sperm indices decreased to their lowest levels during the final 3 weeks of the study. Sperm viability was highly correlated (adjusted R²=0.84) with sperm motility. Sperm cryopreserved in modified Mounib's extender (MME) had the highest post‐thaw motility compared with two other extenders. Post‐thaw motility of sperm cryopreserved in MME was not different from fresh after 90 days of storage. There was no difference in fertilization rates between fresh (69±2.4%) and post‐thaw (67±4.1%) sperm samples taken from the same male or among males. These results demonstrate that the quality of black sea bass spermatozoa is higher earlier in the spawning season and that acceptable post‐thaw fertilization rates can be obtained from cryopreserved sperm.     

Effects of shelter on growth and survival in age-0 black sea bass, Centropristis striata (L.)

Black sea bass Centropristis striata (L.) juveniles were reared in aquaria containing either shelter or no shelter to investigate the effects of shelter on growth and survival. The specific growth rates of juveniles were significantly higher in the sheltered aquaria. With shelter present, the average mortality of juveniles caused by agonistic behaviour was 44% compared with no mortality in the unsheltered aquaria. The results provide evidence that shelter is advantageous for growth, but not survival of black sea bass juveniles under culture conditions.     

Effects of dietary protein and lipid levels on growth performance and body composition of black sea bass Centropristis striata (Linnaeus 1758) during grow-out in a pilot-scale marine recirculating system

The influence of four formulated practical diets, with different protein and lipid levels, on the growth and body composition of black sea bass ( Centropristis striata L.) pre-adults was evaluated in a pilot-scale marine recirculating system. Four test diets were prepared with a combination of two protein levels (44% and 54%) and two lipid levels (10% and 15%). The diets were as follows: low protein and low lipid (LP:LL; 44:10), low protein and high lipid (LP:HL; 44:15), high protein and low lipid (HP:LL; 54:10) and high protein and high lipid (HP:HL; 54:15). Fish (mean weight=75.5 g) were fed the respective diets for 90 days. For fish fed LP:HL, body weight gain was significantly ( P <0.05) higher than fish fed LP:LL. Increasing the protein level from 44% to 54% did not produce a significant effect on weight gain at high lipid level. A significant ( P <0.01) interactive effect between dietary protein and lipid levels on the growth and feed utilization was observed. Total lipid content in the whole body, muscle and liver was significantly affected by the dietary lipid levels. The results suggested that a combination of 44% dietary protein and 15% lipid was optimal for the growth of black sea bass.     

Comparative fatty acid composition of eggs from wild and captive black sea bass, Centropristis striata L.

Lipid content, lipid class and fatty acid compositions were determined in eggs from wild and captive black sea bass, Centropristis striata L., from northern (New England) and southern (South Carolina) regions to determine the effects of diet on egg composition and fertilization success. The formulated diets fed to the northern captive (NC) fish were higher in total lipids (22%) compared with the cut fish and squid diet fed to the southern fish (SC; 3.3% lipid) and had a higher relative amount of linoleic acid (LA) and lower relative amounts of docosahexaenoic acid (DHA) and arachidonic acid (AA). These dietary differences were broadly reflected in the egg lipid composition. The LA levels were higher in all lipid classes in eggs of NC fish while AA levels were lower. The DHA was higher in the major polar lipids (PL) (phosphatidylcholine and phosphatidylethanolamine) of SC eggs compared with NC. Compared with wild fish, both captive groups produced eggs with PLs richer in LA and lower in DHA and AA. Over all fish groups, fertilization success was directly correlated with levels of DHA and AA in the PLs and was inversely correlated with LA levels.     

The effects of temperature on sex differentiation and growth of black sea bass (Centropristis striata L.)

To examine the effects of temperature on sex differentiation in the black sea bass ( Centropristis striata L.), a protogynous hermaphrodite, juveniles (∼0.5 g) were cultured in recirculating systems at 17, 21 or 25 °C. Growth was assessed at 155, 182, 241 and 275 days post hatch and sex differentiation was determined histologically. No differences were found in the sex ratios of fish reared at different temperatures, but only 55–64% developed as females. Growth was significantly greater in males across all temperature treatments. These results suggest that black sea bass exhibit sexually dimorphic growth patterns and that female-specific sex determination can be disrupted in culture.     

Effect of dietary lipid level on juvenile black sea bass Centropristis striata fed two protein sources

A 10-week feeding trial was conducted with juvenile black sea bass Centropristis striata fed isonitrogenous diets (46% CP) with four lipid levels (7, 10, 13 and 16%) to identify dietary lipid levels supporting maximum weight gain and efficiency. Menhaden fish meal (FM) and animal protein concentrate (APC) were used in a 2×4 factorial experiment to evaluate effects of protein quality and lipid level on production. Weight gain, thermal growth coefficient, protein and energy retention, proximate composition and hepatic indices were evaluated. Weight gain was higher (P<0.05) in FM treatments and no protein sparing effect was observed. All growth, efficiency and retention parameters, except apparent net protein utilization (ANPU) among FM diets, were independent of dietary lipid. Based on proximate composition, diets should be restricted to no more than 13% lipid in 46% CP diet, and can be reduced to 7% without loss of growth and efficiency. Second order polynomial regressions on ANPU indicated 10–12% dietary lipid in a 46% CP diet is appropriate depending on protein quality. Reducing dietary lipid in current commercial feeds fed to black sea bass could represent a substantial cost savings in feed production.     

Hormonal induction of ovulation in pikeperch (Sander lucioperca L.) using human chorionic gonadotropin (hCG) and mammalian GnRH analogue

The aim of the present study was to study spawning stimulation in artificial reproduction of females pikeperch (Sander lucioperca L.) using “Chorulon” containing the human chorionic gonadotropin (hCG) and compare with “Supergestran” containing a mammalian GnRH ([D-Ala⁶]GnRHProNhet) analogue. The females were divided into eleven experimental groups and injected with hCG at 250, 500, 750, and 1 000 IU kg^?1 body weight (BW) and mGnRHa at 1, 2.5, 5, 10, 25, and 50 μg kg^?1 BW. In all treatments, a single intramuscular injection of hormone was performed. Control group was injected with 0.9 % NaCl, 0.9 cm³ kg^?1 BW. The average percentages of ovulating females were 88.5 ± 12.3 and 80.8 ± 10.9 % in hCG- and mGnRHa-treated groups, respectively. The average diameter of eggs was 0.95 ± 0.06 and 0.98 ± 0.06 mm in hCG- and mGnRHa-treated groups, respectively. Neither ovulation rate nor diameter of egg was statistically differed among hormonally treated groups. Statistical difference was observed only in hatching rate, where the average were 73.6 ± 14.4 and 50.6 ± 17.7 % in hCG and mGnRHa-treated groups, respectively. Among hormonally treated groups, the best results were observed in groups treated with hCG at 500 and 750 IU kg^?1 and in groups treated with mGnRHa at 25 μg kg^?1. No ovulation was observed in the control group. This study indicated successful ovulation in pikeperch using a single intramuscular injection of hCG or mGnRHa analogue.     

Evaluation of zinc supplementation in European sea bass (Dicentrarchus labrax) juvenile diets

This study aimed to investigate the effect of increased zinc dietary levels on the growth performance, feed utilization, immune status and induced wound healing in European sea bass (Dicentrarchus labrax). Fish weighing 10 g were fed five diets containing organic zinc at 30, 70, 110 and 150 mg kg^?1 diet and one inorganic zinc source (zinc oxide) at 150 mg kg^?1 diet for a period of 12 weeks. No significant (P>0.05) differences were found in the growth performance parameters (final weight, specific growth rate), immunological indices tested (respiratory burst activity in whole blood) and the wound‐healing process. Zinc accumulation in the skin, vertebrae and liver increased significantly (P<0.01) with increased dietary zinc levels but not in muscle. The highest zinc concentrations were obtained in skin tissue, followed by vertebrae, liver and muscle tissue. Using the concentration of zinc in skin as a response criterion, broken‐line analysis showed that the supplementation of 148 mg organic Zn kg^?1 diet seemed to be the optimum dietary zinc supplementation level for sea bass juveniles.     

Nutrient digestibility profile of premium (category III grade) animal protein by-products for temperate marine fish species (European sea bass, gilthead sea bream and turbot)

Three trials, with classical experimental designs for in vivo digestibility studies, were conducted to determine the apparent digestibility coefficient (ADC) of protein (ADCp), lipid (ADCl), energy (ADCe) and amino acids (AA) in selected animal by-products fed to European sea bass, Dicentrarchus labrax (Trial 1), gilthead sea bream, Sparus aurata (Trial 2), and turbot, Psetta maxima (Trial 3). In each trial, five experimental diets [including a reference diet (RD)] where fish meal (FM) was used as the sole protein source were fed ad libitum to the fish for a period of 4 weeks. Test diets were based on the FM RD and obtained by replacing 30% of the RD with a category III designated European animal by-products (fit for human consumption), namely: steam hydrolysed feather meal (HFM), enzyme-treated feather meal (EFM), poultry meat meal (PMM) and spray-dried haemoglobin meal (SDHM). Faecal material was collected using the 'Guelph system', and nutrient and energy digestibility coefficients were related to the measurement of chromic oxide (Cr₂O₃) incorporated into the diet at a rate of 0.5%. Without any exception, FM diets yielded the best digestibility values for all macro-nutrients and by all fish. Among the test ingredients, ADCp was consistently higher for PMM and SDHM in the three species (85.5%, 91.1% in sea bass; 79.2%, 82.8% in sea bream; and 78.4%, 74.8% in turbot). Conversely, ADCp of HFM and EFM were less efficiently digested (67.2%, 68.2% in sea bass; 21.5%, 21.7% in sea bream; and 46.6%, 36.0% in turbot). However, the novel processing method applied to feather meal did not considerably influence the digestibility of most of the nutrients in this feedstuff. The current investigation yielded valuable numerical ADC for EAA considered to be of prime importance in generating balanced diet formulations.     

Effects of dietary docosahexaenoic acid (22:6n‐3) and arachidonic acid (20:4n‐6) on the growth,survival, stress resistance and fatty acid composition in black sea bass Centropristis striata (Linnaeus 1758) larvae

The objectives of this study were to determine the effects of the dietary docosahexaenoic acid (DHA) to arachidonic acid (ARA) ratio on the survival, growth, hypersaline stress resistance and tissue composition of black sea bass larvae raised from first feeding to metamorphic stages. Larvae were fed enriched rotifers Brachionus rotundiformis and Artemia nauplii containing two levels of DHA (0% and 10% total fatty acids=TFA) in conjunction with three levels of ARA (0%, 3% and 6% TFA). On d24ph, larvae fed the 10:6 (DHA:ARA) treatment showed significantly (P<0.05) higher survival (62.3%) than larvae fed 0:0 (DHA:ARA) (27.4%). Notochord length and dry weight were also significantly (P<0.05) greater in the 10:6 (DHA:ARA) treatment (8.65 mm, 2.14 mg) than in the 0:0 (DHA:ARA) (7.7 mm, 1.65 mg) treatment. During hypersaline (65 g L⁻¹) challenge, no significant differences (P>0.05) were observed in the median survival time (ST50) between larvae fed 10% DHA (ST50=25.6 min) and larvae fed 0% DHA (ST50=18.2 min). The results suggested that black sea bass larvae fed prey containing 10% DHA with increasing ARA within the range of 0–6% showed improved growth and survival from first feeding through metamorphic stages.     

Photoperiodic manipulations of the reproductive cycle of sea bass (Dicentrarchus labrax) and their effects on gonadal development, and plasma 17ß-estradiol and vitellogenin levels

The annual profile of plasma vitellogenin (VTG) and 17ß-estradiol (E₂) levels, as well as gonadal development and spawning characteristics were investigated in captive female sea bass (Dicentrarchus labrax). Endocrine and gonadal changes were studied in fish reared under natural conditions or exposed to manipulated photothermal cycles. In natural conditions of photoperiod and temperature, sea bass spawned from February through March (East coast of Spain, 40°N 0°E). One or two months of constant long-days (15L/9D) in a constant short-day photoperiod regime (9L/15D) all-year-round, given early in the year (March and March–April), advanced spawning by 3 months. The same treatment applied later in the year (September–October) delayed spawning by 1 month, compared to controls.In all groups, changes in plasma VTG levels were correlated with E₂ levels, oocyte growth and spawning time. In control females, VTG was low (<100 ng ml^-1) during the summer, until its first surge in plasma 4 months before the beginning of spawning. The VTG (3.1 ± 0.3 mg ml^-1) and E₂ (4.1 ± 0.5 ng ml^-1) levels showed a single annual peak during late vitellogenesis, the time of the highest proportion of vitellogenic oocytes in the ovary. Constant high levels of VTG (1–1.4 mg ml^-1) and E₂ (1.6–1.9ng ml^-1) were maintained during the entire spawning time, together with the presence of vitellogenic oocytes, suggesting the existence of several waves of oocyte growth in the ovary and thus, several spawns per female. Endocrine profiles and oocyte development in fish exposed to constant photoperiods were similar to controls, but were shifted in time in relation to the displacement of the spawning time. In the fish showing advanced spawns, the duration of the gametogenic proces was compressed when compared to controls. The differences observed in the evolution of the reproductive-related factors in the advanced groups, which were exposed to a reduction in temperature to 15°C, suggest an influence of the temperature in the early stages of the reproductive cycle in sea bass.     

Effect of electrical stunning at slaughter on the carcass, flesh and eating quality of farmed sea bass (Dicentrarchus labrax)

Food quality aspects of farmed sea bass (Dicentrarchus labrax) were compared following two methods of slaughter: the normal commercial method of killing, by immersion in an ice slurry, or by first electrically stunning the fish, before immersion in an ice slurry. Quality was assessed for up to 10 days of storage on ice after slaughter. No differences were found between the slaughter methods in terms of an overall sensory evaluation of cooked fillets, or in terms of overall carcass quality: overall appearance, internal and external haemorrhage, fin damage, burst gall bladder, staining of the body cavity by leakage from the gut or damage to the spine. Using objective measurements of colour, no differences between fish from either treatment were found in terms of external colour or colour of the fillets. A chemical analysis of flesh nucleotide breakdown products as well as the freshness indicator K_i value did not differentiate the two treatments nor did the industry standard freshness scoring technique (QIM, quality index method), over 10 days of storage on ice. Flesh pH was marginally lower in electrically stunned fish at 4 h post mortem (6.42 cf 6.56) but by 24 h, pH in fish from both treatments had decreased to a similar level (6.22). Humane electrical stunning of sea bass at slaughter neither measurably improved nor decreased product quality for between 1 and 10 days of storage on ice. Electrical stunning accelerated the pattern of onset and resolution of rigor mortis. If electrical stunning were to be widely adopted, re‐education of buyers would be necessary as rigor mortis is currently used by buyers as a proxy measure of fish freshness.     

Evaluation of electrical stunning of sea bass (Dicentrarchus labrax) in seawater and killing by chilling: welfare aspects, product quality and possibilities for implementation

The objective was to assess neural, behavioural responses and product quality in farmed sea bass (Dicentrarchus labrax) upon electrical stunning in seawater. The electrical sinusoidal 50 Hz or pulse square wave alternating 133 Hz current induced a general epileptiform insult with a current of 3.3±0.2 or 3 A dm^?2, respectively, for 1 s head to tail in seawater. The total duration of the insult was 48±34 and 23±11 s. After stunning, the electro‐cardiogram revealed fibrillation and ceased or showed malfunction. Product quality was assessed in a group electrically stunned, followed by chilling in ice water and the controls were only chilled in ice water. The pH of the fillets was 0.1–0.2 lower (P<0.01) when stunned electrically at days 1, 2, 8 and 10 postmortem, where the colour did not differ. The percentage of sea bass effectively stunned using an electrical sinusoidal or pulse square wave current was above 85% within a confidence level of 95%. A combination of electrical stunning for 10 s, followed by chilling in seawater with ice flakes resulted in the death of all fish. The former method is recommended to be adapted for implementation in practice.     

Improved feed utilization,intestinal mucus production and immune parameters in sea bass (Dicentrarchus labrax) fed mannan oligosaccharides (MOS)

In the present study, the effects of dose dietary administration of mannan oligosaccharides (MOS) (Bio‐Mos; Alltech Inc.) on growth, digestibility, liver morphology, lipid and carbohydrate metabolism enzymes, organoleptic properties, immune parameters and gastrointestinal tract mucus production were assessed. MOS dietary inclusion significantly reduced lipogenic enzymes activity in liver and resulted in a lower lipid vacuolization and more regular‐shaped morphology of hepatocytes around the sinusoidal spaces, which could be associated to a better energy and feed utilization because of the improvement of feed utilization and the reduction of feed intake found in the current experiment. Moreover, MOS inclusion enhanced phagocytic activity of head kidney leukocytes in fish fed 4 and 6 g kg^?1 MOS diets at days 30, 45 and 60. Gut quantitative morphological analyses showed a marked dose‐dependent enhancement in the number of cells secreting acid mucins by unit of area which could be related to the previous improvement in resistance to bacterial infection. No effects on sensorial parameters and biochemical composition of flesh were detected.     

Fructose 1,6 bisphosphatase activity in liver and gonads of sea bass (Dicentrarchus labrax). Influence of diet composition and stage of the reproductive cycle

The fructose 1,6 biphosphatase (FBPase) activity was measured in the liver and gonads of broodstock sea bass that were fed three experimental diets over the reproductive cycle. Enzyme activity was measured at the pre-spawning (November), spawning (February) and post-spawning (May) periods. Additionally, some biometric indices were calculated at these three times. The different diets, one with standard composition (51% protein, 13% lipid, 10% digestible carbohydrates), one with a high-carbohydrate content (32% protein, 13% lipid, 32% digestible carbohydrates) and one deficient in ω3 essential fatty acids (51% protein, 13% lipid, 10% digestible carbohydrates), had similar affects on growth and gonad development, as deduced from the similar weight, condition factor, hepatosomatic index, gonadosomatic index and liposomatic index over the reproductive cycle. FBPase activity was detected at all times in the liver of both sexes, and in the ovaries but not in the testes. In general, females showed greater liver activity than males, especially at spawning time. During post-spawning, FBPase activity fell in tissues of both sexes. Enzyme activity was also affected, mostly in the liver, by the different types of feed; hepatic activity in fish fed the high-carbohydrate diet fell during the spawning and post-spawning periods, though at a different level for each sex. The diet deficient in ω3 EFA presented the lowest FBPase activity, compared with the other diets, in females during the spawning period; there was very little change in males or females over the reproductive cycle. The results showed that the highest gluconeogenic activity in sea bass broodstock occurred at the spawning time and the females had larger glucose requirements than males over the reproductive cycle, as demonstrated by the absence of FBPase activity in testes. This work also confirmed the adaptative capacity of this enzyme to different nutritional and physiological conditions.