Hypoxic conditions induce centrum defects in red sea bream Pagrus major (Temminck and Schlegel)

A previous study elucidated that an extreme hypoxia during somitogenesis induced the most frequent skeletal malformation centrum defects in red sea bream (RSB), Pagrus major. In this study, details of the hypoxic conditions to induce them in RSB, dissolved oxygen (DO) concentration and exposure time to hypoxia, were investigated. Fertilized eggs were exposed to seawater of six DO concentrations (0%, 10%, 25%, 50%, 75% and 100% of saturation) for seven different periods (5, 10, 30, 60, 120, 240 and 360 min) during somitogenesis. Somitic disturbances in newly hatched larvae were induced by exposure to 0% and 10% DO concentration for 10 and 120 min and longer respectively. Rearing eggs exposed to hypoxic condition of 10% DO for 240 min for 40 days post‐hatch showed that the location and the frequency of somitic disturbances in larvae and centrum defects in juveniles were significantly correlated (P<0.01). Dissolved oxygen concentration of the interstitial water in the egg high density layer formed at the water surface in a stationary state abruptly decreased to 3.7% within 7 min. Centrum defect induction by exposure of eggs to extreme low DO concentrations for a short period, which is the probable situation in the practical juvenile production, suggests that careful maintenance of DO concentration is important in the incubating water of fertilized eggs during egg sorting and transportation, where eggs are made into a pile and undergo hypoxia, for the prevention of centrum defects.     

Teratogenic effects of isolated and combined short‐term hypercapnia and hypoxia on red sea bream (Pagrus major) embryos

Some vertebral anomalies in cultured fish arise from unusual environmental conditions during embryogenesis. We investigated the individual and combined teratogenic effects of short‐term hypercapnia and hypoxia on embryos of red sea bream (RSB), Pagrus major. Ten‐somite stage embryos were exposed to hypercapnia (60 and 120 mg/L dissolved carbon dioxide: DCD) and hypoxia (0%, 10%, 25%, 50%, and 75% dissolved oxygen: DO) independently and concomitantly for seven different periods (0, 30, 60, 90, 120, 150 and 180 min) to examine somitic disturbances at hatching, which are prodromal symptoms of centrum defects. Another experiment examined vertebral anomalies in juveniles raised from eggs exposed to hypercapnia (120 mg/L DCD) and hypoxia (10% DO) independently and concomitantly. Short‐time exposures (30–180 min) to hypercapnia (60 and 120 mg/L) and hypoxia (10% DO or less) independently and additively caused larval somitic disturbances and juvenile centrum defects. The results indicate that short‐term hypercapnia and hypoxia generally have the same teratogenic effect on embryos, although there were some differences in the locations of the somitic disturbances and centrum defects, with additive teratogenicity when the conditions were combined. These results suggest the necessity of maintaining appropriate DCD and DO concentrations during egg incubation and transport and during reproduction in recirculating aquaculture.     

Vertebral deformities in cultured red sea bream, Pagrus major, Temminck and Schlegel

Vertebral deformities were investigated in cultured red sea bream, Pagrus major. In the field, deformities in seedlings were categorized and their incidence was calculated. In the laboratory, the symptoms of major vertebral deformities were examined morphologically using radiographs and by making transparent skeletal specimens. The internal structure of deformed vertebrae was examined histologically. The shortened body condition had the highest incidence (0.9–8.3%) of all deformities in the seedlings. In individuals with the shortened body condition, the ratio of trunk and caudal part length to body height was smaller. These fish had skeletal anomalies in the vertebrae, mainly centrum defects (64.3%) or undersized centrums (25.2%). The specimens with centrum defects had a characteristic anomaly in the vertebrae, with plural pairs of neural and haemal spines on a single centrum. This anomaly was frequently observed in the posterior abdominal vertebrae. The internal skeletal structure of such abnormal centrums was basically the same as that of normal centrums. In all the specimens with undersized centrums, both the centrum length and diameter were shorter than normal except for the first and second centrum, and urostyle.     

Induction of centrum defects in amberjack Seriola dumerili by exposure of embryos to hypoxia

ABSTRACT:   Artificially hatched Seriola species have the problem of malformation, mainly in their vertebrae, head, and mouth parts. To clarify the cause of vertebral malformation, the effects of hypoxia during embryogenesis on the induction of centrum defects was investigated in artificially hatched amberjack Seriola dumerili . Firstly, 7-somite stage embryos were exposed to waters of 0, 12.5, 25, 50, and 100% dissolved oxygen (DO) for 0.5, 1, 2, 3, 4, and 5 h to confirm the effective dose (DO concentration and duration of exposure) of hypoxia that induces somitic disturbances in newly hatched larvae. Exposure of embryos to 12.5% DO concentration for longer than 0.5 h induced somitic disturbances. Following this result, centrum defects in juveniles were investigated by an induction experiment with embryos exposed to 12.5% DO for 2 h at the gastrula, 1- or 2-somite, 10-somite, 15-somite, or heart beating stage. This experiment revealed that centrum defects were induced only during somitogenesis, and somitic disturbances were the premonitory symptom of centrum defects. These results indicate hypoxia during somitogenesis as a possible cause of centrum defects in amberjack.     

Spawning grounds of red sea bream in the east Seto Inland Sea

We surveyed the distribution of red sea bream eggs between 2006 and 2010 in the east Seto Inland Sea to determine the location of the spawning grounds and the characteristics of preferred spawning habitats. We identified early-development stage red sea bream eggs using a monoclonal antibody assay. The early-development stage eggs were present in waters around Awaji Island (AW area) and around Ieshima Island, Shodo Island, and Bisan Seto (collectively the ISB area). The mean red sea bream egg density in the AW area was not significantly different from that in the ISB area. Based on the locations where we observed early-development stage eggs and the density of eggs, the ISB and AW areas appeared to be the primary spawning grounds in the east Seto Inland Sea during the survey period. A quotient analysis suggested that spawning fish had clear preferences for physical conditions. The areas suitable for spawning appear to be those with sandy substrate at a depth of 30–70 m. The temperature range suitable for spawning was between 16.5 and 21.5 °C.     

Spawning season,spawning grounds,and egg production of red sea bream in Hiuchi-nada,Seto Inland Sea

The spawning season and grounds of red sea bream in Hiuchi-nada, the central part of the Seto Inland Sea, were described using a new method based on monoclonal antibodies for identifying Pagrus major eggs, and the daily egg production (standardized by the incubation time and survival rate) was estimated. At the peak of spawning (May), the ranges of sea temperature, salinity, and chlorophyll a where red sea bream eggs occurred were 14.8–17.4°C, 32.0–33.0, 0.5–4.4 μg/L, respectively. The main spawning grounds of the red sea bream were confirmed as being the areas near the Geiyo Islands, Misaki Peninsula, Saijyo, Niihama. The spatial spread of red sea bream eggs increased with the egg developmental stage. The ranges of daily egg production in 2005 and 2006 were 0.3–19.3 and 0.2–6.7 × 10⁹ eggs/day, respectively. In Hiuchi-nada, aquaculture farms are located close to the spawning grounds, and the potential spawning population from the aquaculture farms equaled or exceeded that of the estimated spawning population obtained by the egg production method. Red sea bream eggs in Hiuchi-nada might be produced by both wild and aquaculture-based spawning populations.     

Effects of Fish Size and Water Temperature on the Acute Toxicity of Copper for Japanese Flounder,Paralichthys olivaceus,and Red Sea Bream,Pagrus major

The acute toxicities of copper were examined for Japanese flounder, Paralichthys olivaceus, and red sea bream, Pagrus major, in terms of fish size and water temperature. Artificial seawater of low pH of 5.4–6.7 was used as testing water to keep dissolved copper concentration at 0.04–41 mg Cu/L. Japanese flounder of 0.3–17 g and red sea bream of 0.5–13 g were exposed to different concentrations of copper for 96 h at 20 C under semistatic condition. Median‐lethal concentration for 96 h of Japanese flounder and red sea bream were 8.7–12.2 and 2.0–5.2 mg Cu/L, respectively. No significant relationships were observed between median‐lethal concentrations for 96 h and fish size for Japanese flounder, while the value decreased significantly with increasing fish size for red sea bream. Effect of water temperature on the acute toxicity was examined for Japanese flounder of 0.3 and 0.4 g at 10, 15, 20, and 25 C and red sea bream of 0.5 and 1.0 g at 12, 15, 20, and 25 C. Ninety‐six‐hour median‐lethal concentrations for Japanese flounder and red sea bream were 5.1–11.2 and 1.0–5.3 mg Cu/L, respectively. No significant relationships were observed between median‐lethal concentrations for 96 h and water temperature for both fish species.     

Vertebral deformity in cultured Atlantic cod larvae: ontogeny and effects on mortality

The effects of different egg incubation densities on the incidences of vertebral deformities in Atlantic cod larvae were investigated. Cod eggs were incubated at four different densities, 3, 6, 12 and 48 mL eggs L⁻¹, of water.  When all the eggs hatched, larvae were reared in 30 L glass aquaria. Larval samples were taken at 0, 14, 42 and 56 days post hatch (dph) for deformity analysis. Larval samples were stained using bone and cartilage staining methods to determine vertebral deformity. Incubation densities did not have any significant effects on vertebral deformities in Atlantic cod larvae. However, the incidence of larval vertebral deformity was high at hatch and decreased as the larvae grew older until 42 dph, indicating selective mortality of deformed larvae during this period. Larvae at 56 dph, however, showed an increase in the incidence of vertebral deformity, indicating a possible nutritional or prey-type effect. To our knowledge, no studies have documented the occurrence of variable patterns in vertebral deformities in cod at various developmental stages. Overall, our results suggest that broodstock husbandry, genetics and/or nutrition could play a major role in causing vertebral deformities in Atlantic cod at hatch; however, nutrition and prey type may play a major role during metamorphosis.     

Skeletal development and deformities in cultured larval and juvenile seven-band grouper, Epinephelus septemfasciatus (Thunberg)

The seven‐band grouper, Epinephelus septemfasciatus (Thunberg), is currently recognized as a potential new species for aquaculture in Japan. This study describes normal and abnormal skeletal development of the jaw and vertebrae in cultured larvae and early juveniles of E. septemfasciatus. The ontogenetic stages at which skeletal deformities of jaw and vertebra developed were also described for this species. Osteological observations were made using a clearing and staining method for larvae and soft X‐ray photographs for juveniles. A high incidence of skeletal deformities was observed in the jaws and vertebral column during the larval and juvenile stages. Most of the jaw deformities were explained by an abnormal maxilla curvature. Jaw deformities were visually evident from flexion stage after ossification of the deformed elements. Deformities in the vertebral column (mostly lordosis) were observed from the post‐larval stage and became more evident as growth proceeded. The lordosis generally occurred on the positions of the 8–11th vertebra. These types of deformities are compared with those of other species, and possible causative factors of the skeletal deformities are discussed.
     

Efficacy of taurine supplementation for preventing green liver syndrome and improving growth performance in yearling red sea bream Pagrus major fed low-fishmeal diet

ABSTRACT:   This study was performed to evaluate the efficacy of taurine supplementation for preventing green liver syndrome and improving growth performance in red sea bream Pagrus major fed a low-fishmeal (FM) diet. Yearling red sea bream were fed for 34 weeks on low-FM diets either supplemented with taurine, or without taurine, and the tissue taurine and bile pigment concentrations were measured. Compared to the fish fed the FM diet, fish fed the low-FM diet without taurine supplementation resulted in inferior feed performances and higher incidence of green liver related to the morphological transformation of the erythrocytes. In these fish, the hepatopancreatic taurine concentration was significantly lower and hepatopancreatic biliverdin concentration was high compared to the fish fed the FM diet. These parameters were markedly improved by taurine supplementation of the low-FM diet and were similar in levels to the fish fed the FM diet. These results indicate that green liver appearance and inferior feed performances of red sea bream fed the low-FM diet without taurine supplementation were caused by dietary taurine deficiency, and indicate the requirement of taurine supplementation to low-FM diets for red sea bream.     

A new type of lordosis and vertebral body compression in Gilthead sea bream,Sparus aurata L.: aetiology,anatomy and consequences for survival

A new type of vertebral malformation is described, consisting of deformed cartilaginous neural and haemal processes and the compression and fusion of vertebral bodies. The malformation is designated as haemal vertebral compression and fusion (haemal VCF). We studied the aetiology of the malformations and described microanatomical histopathological alterations. The malformations were detected during routine quality control in one of six monitored Gilthead sea bream populations. Haemal VCF affected the posterior part of the vertebral column (haemal vertebrae). In 20% of the deformed specimens, haemal VCF was combined with lordosis. At 35 dph (days post‐hatching), early anatomical signs of the haemal VCF consisted of abnormal centrum mineralization, malformed cartilaginous neural and haemal processes and developing lordotic alterations. The histological examination of the deformed individuals revealed that haemal VCF is preceded by notochord abnormalities. The frequency of deformed individuals was three times higher at 35 than at 61 dph (50.3% vs. 17.2%, n = 157 and n = 250, respectively). No signs of repair or reversion of malformations have been observed. Thus, the steep decrease in deformities in older animals suggests that haemal VCF is linked to high mortality rates. The results are discussed in respect of the possible causative factors of haemal VCF.     

Etapes du developpement embryonnaire et larvaire du sar, Diplodus sargus L., en elevageStages of the embryonic and larval development of the white sea bream, Diplodus sargus L., in rearing conditions

Eggs of white sea bream (Diplodus sargus), taken from wild parents, were incubated and, after hatching, produced larvae which were then reared up to the fry stage. The different stages of development during the embryonic and larval periods are described. Some characteristics of the eggs (buoyancy, duration of incubation, critical periods) and of the larvae (metrical characters) are reported.     

Diluted rotifer, Brachionus plicatilis (Muller), extract as culture medium stimulates hatching of red sea bream, Pagrus major (Temminck & Schlegel)

Hatching rates and hatching times (50% hatch) of red sea bream, Pagrus major (Temminck & Schlegel), eggs in culture media that were prepared to dilute rotifer Brachionus plicatilis Muller extract at 4000-, 6000-. 8000- and 10 000-fold with artificial sea water (σ₁₅, = 24.0) were investigated. Although differences in hatching rate between the culture media and control artificial sea water were negligible, hatching time was significantly shorter in media at 4000- and 6000-fold dilutions than in media at 8000- and 10 000-fold dilutions and in the control artificial sea water, when the eggs immediately post spawning were incubated. The hatching rates of eggs at three developmental stages, immediately post-spawning, gastrula stage and heart-beat initiation stage, incubated respectively in the medium at 6000-fold dilution and the control, did not differ either, yet the hatching times at each egg stage in the medium were significantly shorter than that in the control. These results suggest that hatching enzyme secretion from the hatching glands is promoted to perceive chemical stimuli in the water-soluble fraction of rotifer extract via the embryo's chemoreceptors, from the heart-beat initiation stage to immediately before hatching, thus shortening the hatching time.     

Stress assessment,quality indicators and shelf life of three aquaculture important marine fish,in relation to harvest practices,water temperature and slaughter method

The objective of this study was to evaluate the effect of harvest practices and slaughter method on stress, quality and shelf life of whole fish (gilthead sea bream, European sea bass and red sea bream) towards the development of a humane slaughter practice. The use of hook and line resulted in significantly lower plasma cortisol and glucose in European sea bass and gilthead sea bream. Water temperature at harvest affected significantly the concentrations of stress indicators (plasma cortisol and glucose), mainly in European sea bass and gilthead sea bream. No significant differences were observed between the harvesting methods, regarding fish appearance, microbial spoilage and sensory scoring and subsequently shelf life. However, significantly lower hardness was recorded in fish harvested using the electric stunner, compared to the fish killed with slurry ice of the same harvesting period. Shelf life ranged between 13–18 days for gilthead seabream, 10–16 days for European sea bass and 10–14 days for red sea bream, depending on harvesting period.     

Necessity of dietary taurine supplementation for preventing green liver symptom and improving growth performance in yearling red sea bream Pagrus major fed nonfishmeal diets based on soy protein concentrate

The necessity of dietary taurine supplementation for preventing green liver symptom and improving growth performance of red sea bream Pagrus major fed nonfishmeal (non-FM) diets was investigated. Yearling red sea bream (initial body weight, 580 g) were fed for 36 weeks on non-FM diets based on soy protein concentrate (SPC) supplemented with taurine at levels of 0%, 0.5%, 1.0%, 1.5%, and 2.0%. Specific growth rate (SGR) and feed conversion ratio (FCR) of fish fed the taurine-unsupplemented SPC diet were markedly inferior. In these fish, incidence of green liver was markedly higher and was accompanied by a decrease of tissue taurine concentration and an increase of hepatopancreatic bile pigment content. The green liver symptom was mainly caused by an increase of hemolysis since the erythrocytes became osmotically fragile due to taurine deficiency. Physiological abnormality and growth performance (SGR and FCR) were markedly improved by taurine supplementation to the SPC diets. These results indicate that dietary taurine supplementation is necessary for yearling red sea bream fed non-FM diet based on SPC to maintain normal physiological condition and growth performance.     

Ontogeny of tolerance to and avoidance of ultraviolet radiation in red sea bream Pagrus major and black sea bream Acanthopagrus schlegeli

ABSTRACT:   Ontogenetic changes of tolerance to, and avoidance of, ultraviolet-B radiation (UV-B) were examined in red sea bream Pagrus major and black sea bream Acanthopagrus schlegeli . In the tolerance experiment, larvae and juveniles (age 13–46 days) were put in beakers, and were exposed to one of five different levels of UV-B radiation (1.8, 1.1, 0.2, 0.1, and 0 W/m²) for one hour. Their survival rates were calculated either 12 or 24 h later. In the avoidance experiment, fish (age 3–49 days) were put in a long experimental tank, half of which was covered with UV-blocking film and placed under two levels of UV-B radiation (1.1 and 0.2 W/m²), and their avoidance indices were calculated. Black sea bream had significantly better survival compared to red sea bream for most ages. Only black sea bream of ages 37 and 49 days showed significant avoidance of UV radiation under the higher level of UV-B, whereas both species did not show avoidance on any days at the lower level. The present results suggest that black sea bream are significantly better adapted to habitats with high UV-B radiation, than red sea bream, reflecting that back sea bream live in shallower waters through their early life stages.     

Dietary inorganic phosphorus or microbial phytase supplementation improves growth,nutrient utilization and phosphorus mineralization of juvenile red sea bream,Pagrus major,fed soybean‐based diets

A 2 × 3 factorial design with triplicates examined the interaction between dietary inorganic phosphorus (IP) and phytase on growth, mineral utilization and phosphorus (P) mineralization in juvenile red sea bream. The treatments were three levels of dietary IP supplementation at 0, 2.5 and 5 g kg^?1, either without or with phytase supplementation [2000 FTU kg^?1; phytase unit is defined as the amount of enzyme activity which liberates 1 micromol of inorganic phosphorus per minute at pH 5.5 and 37 °C at a substrate concentration (sodium phytate) of 5.1 mmol L^?1]. Juvenile red sea bream (IBW = 1.3 g ± 0.1) were stocked twelve fish per tank and fed for 50 days. Growth and feed efficiency were significantly (P < 0.05) enhanced by both dietary P and phytase supplementation. Feed intake and survival rate were not significantly affected by the dietary treatments. Both dietary IP and phytase supplementation significantly increased plasma IP and Mg levels. Concentration of vertebral mineral and scale P was significantly increased by both dietary treatments. A skeletal malformation syndrome of scoliosis occurred in fish fed both non‐IP and non‐phytase supplemented diet. Interaction between main dietary effects was detected for vertebral Zn, scale P and whole‐body ash and Mg content. With regard to growth and other examined productivity traits, phosphorus requirement of juvenile red sea bream can be met if supplemented with 2000 FTU phytase kg^?1 or in the absence of phytase, by dietary inclusion of 2.5–5 g kg^?1 of IP.     

Assessment of reference dietary amino acid pattern for juvenile red sea bream, <Emphasis Type="Italic">Pagrus major</Emphasis>

Four semi-purified diets, containing crystalline amino acids (CAAs), were fed to juvenile red sea bream, Pagrus major in order to ascertain the ideal dietary amino acid pattern for this species. A control diet containing 50% casein–gelatin as protein sources, but no CAAs were fed to the fish. The other diets contained 30% casein–gelatin and 20% CAAs. CAAs were added to diets to simulate with amino acid pattern of the red sea bream eggs protein (REP), red sea bream larvae whole body protein (RLP), red sea bream juvenile whole body protein (RJP), and brown fishmeal protein (BFP). The juveniles (average initial body weight, 1.58 ± 0.01 g) were maintained in triplicate tanks and fed twice daily for 30 days. The highest weight gain was observed in juveniles fed the RJP diet. No significant difference was observed in juveniles fed the RLP and BFP diet. Feed efficiency ratio, protein efficiency ratio and amino acid retention in the whole body were significantly (p < 0.05) affected by the simulated dietary amino acid patterns. The essential amino acid profile and A/E ratios of the whole body after the growth trial showed little difference among the dietary treatments. The results suggest that red sea bream juveniles are able to utilize high amounts of CAA in coated form. The amino acid pattern of RJP could be used as an appropriate of reference dietary amino acid for this species.     

Subtropical water influences temporal fluctuations of early life stages of Vinciguerria lucetia (Osteichthyes: Phosichthyidae) in the Humboldt Current System (1998–2004)

In the last decades, the fish Vinciguerria lucetia (Garman) has been of important interest to the fisheries sector; nonetheless, the spawning and nursery zones in the Humboldt Current System (HCS) have not yet been defined. By using a temporal series of 23 oceanographic surveys from austral spring of 1998 to autumn 2004 off northern Chile, the spatial and temporal distribution and abundance of eggs and larvae of V. lucetia were studied. The relationships with environmental conditions (sea surface temperature, water column stratification, salinity, dissolved oxygen) were modeled using generalized additive models (GAMs). Seasonal variations in eggs and larval abundances were recorded, and higher abundances were observed in spring and summer, respectively. The main spawning areas were located at approximately between 40 and 80 nautical miles offshore. The largest abundances of V. lucetia eggs were found during spring 2003; however, larval abundances reached the highest values following the strongest ENSO event 1997–98. GAMs predicted that offshore location, sea surface temperature, and the deepening of the oxygen minimum zone, characteristics of the subtropical waters (22–24°C, >34.9, 3–6 ml/L) drove eggs and larval distributions of V. lucetia in the HCS during 1998–2004, toward areas with scarce food availability for larvae. These results suggest that spawning and larval development of this oceanic species occur in oligotrophic waters as a loophole strategy, in order to reduce predation risk during early life stages.     

Transgenic medaka with brilliant fluorescence in skeletal muscle under normal light

ABSTRACT:   Green fluorescent protein ( GFP ) and red fluorescent protein ( RFP ) genes regulated by the medaka skeletal muscle actin promoter were microinjected into fertilized d-rR medaka eggs to establish transgenic medaka lines. Intense fluorescence was detected in skeletal muscle. During development, GFP and RFP became detectable in anterior somites at the 12- and 30-somete stages, respectively. After hatching, intense fluorescence in skeletal muscle enabled individual fish to be identified under normal lighting without fluorescent microscopy. Fluorescence was also observed in the gills and esophagus of the adult fish. These data indicated that medaka lines are convenient not only for the study of skeletal muscle but also for the identification of cells or individuals in various studies.