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1.
A cost-effective method for producing and using recombinant growth hormone (GH) has been developed. A cell-surface protein display system was constructed to produce recombinant Japanese flounder growth hormone (r-JfGH) in yeast Saccharomyces cerevisiae and the biological activity of expressed r-JfGH was evaluated by oral delivery to juvenile Japanese flounder. The expression vector pYD-E2, containing the JfGH gene, S. cerevisiae GAL1 promoter, and the S.cerevisiae URA3 gene downstream and upstream sequences, was designed and constructed to integrate the GH gene into the yeast genome and secrete r-JfGH stably on the surface of the yeast cell. Expression of r-JfGH was confirmed by flow cytometry until the 50th continuous generation of transgenic yeast strain y-E2, and over 0.98 μg r-JfGH per 1 mg total protein of y-E2 was estimated by an ELISA–RA assay. Growth of Japanese flounder was promoted significantly by the direct supplementation of freeze-dried transgenic yeast into the Japanese flounder juvenile feed at levels of 0.5 and 1%. The weight-specific growth rate and feed conversion ratio of the treated groups were 23.07 and 21.57% (0.5% feeding level) and 28.85 and 41.18% (1% feeding level), respectively, higher than that of the control group in a 7-week bioactivity study.  相似文献   

2.
To elucidate the role of gonadotropin-releasing hormone (GnRH) in gonadal maturation in wild female Japanese flounder Paralichthys olivaceus, we monitored changes in the levels of seabream GnRH (sbGnRH) in the olfactory bulb, telencephalon, hypothalamus, and pituitary during ovarian development together with changes in plasma levels of testosterone (T), estradiol-17β (E2), and 17α, 20β-dihydroxy-4-pregnen-3-one (DHP). Fish were caught offshore of the northern mainland of Japan in the Pacific Ocean at 3- to 4-week intervals between April and September by gill net. The netted fish were categorized into six groups based on ovarian stages: previtellogenic (April–early May), early yolk (April–late May), late yolk (late May–June), early spawning (June–August), late spawning (September), and termination (September) stages. The gonadosomatic index significantly increased from the previtellogenic to early spawning stages and decreased thereafter. In the olfactory bulb, no significant differences were observed in sbGnRH levels among the developmental stages. In contrast, sbGnRH levels in the telencephalon and hypothalamus were very high in the previtellogenic stage, lower in the early spawning stage, and relatively high in latter stages. sbGnRH levels in the pituitary were high in the previtellogenic stage and low in the early spawning stage. In addition, the relatively high levels of pituitary sbGnRH were found together with high plasma T, E2, and DHP levels in fish in the late yolk stage. These results indicate that sbGnRH in the telencephalon, hypothalamus, and pituitary is involved in ovarian maturation and that sbGnRH may play an important role in the initiation of ovarian recrudescence in wild Japanese flounder.  相似文献   

3.
We sought to provide a useful indicator of the presence of endocrine-disrupting contaminants along the marine coast of the South Pacific using Chilean flounder (Paralichthys adspersus). In light of the lack of information on vitellogenin for this species, we induced, purified, and identified the plasma vitellogenin of Chilean flounder inhabiting the Chilean coast. Vitellogenin (Vg) from Chilean flounder was purified by size exclusion and ion-exchange chromatography using plasma from juvenile males induced by injecting 17β-estradiol. The Vg was detected by SDS–PAGE and Western blot analyses using an antibody against turbot (Scophthalmus maximus) vitellogenin. These analyses revealed a protein band of 205 kDa and three minor bands of 120, 90, and 68 kDa. These proteins were identified as Vg by means of mass spectrometry (LCQ Duo ESI-IT-MS), matching sequences of tryptic peptides to known sequences for several other fish species. The matches showed the presence of vitellogenin (VgI, VgII, Vg A and Vg B) in Chilean flounder, similar to species such as mummichog (Fundulus heteroclitus), Japanese medaka (Oryzias latipes), and white perch (Morone americana). These results are discussed in terms of identifying Vg in Paralichthys adspersus with the antibody to turbot Vg. Moreover, we compare the molecular size of Vg from Chilean flounder (large) with that of other flatfish species. Finally, we discuss the potential use of this molecule as a biomarker for the presence of xeno-estrogenic compounds along the Chilean coastline.  相似文献   

4.
Shih  Wang  Tan  & Chen 《Journal of fish diseases》2001,24(3):143-150
Three hybridoma clones secreting monoclonal antibodies (MAbs) were produced from mouse myeloma and spleen cells immunized with white spot syndrome virus (WSSV) isolated and purified from Penaeus monodon (Fabricius), collected from north-eastern Taiwan. By sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE), the protein profile of this isolate contained four major proteins with sizes of approximately 35 (VP35), 28 (VP28), 24 (VP24), and 19 kDa (VP19). Western blot analysis revealed that two MAbs (1D7 and 6E1) recognized epitopes on VP28 and one MAb (3E8) recognized an epitope on VP19. The MAb 6E1 isotyped to the IgG1 class was used in both an indirect immunofluorescence assay (IFA) and in an immunochemical staining protocol for successful identification and localization of WSSV in infected shrimp tissues. Antigenic similarity of isolates from Indonesia and Malaysia to the Taiwan isolate was illustrated by IFA with MAb 6E1. A MAb (2F6) which bound specifically to two shrimp proteins, 75 and 72 kDa, and reacted to the healthy and non-target tissues of WSSV in infected shrimp, such as hepatopancreas, is also described here and shows the necessity for specific identification of antibodies.  相似文献   

5.
This study was conducted to investigate the effect of dietary supplementation of cottonseed and soybean meal (CS) on growth performance, feed utilization, and gossypol accumulation in juvenile (mean body weight 11 g) Japanese flounder Paralichthys olivaceus. Five isonitrogenous and isocaloric experimental diets (designated CS0, CS10, CS20, CS30, and CS40) containing 0, 10, 20, 30, and 40% CS mixture (1∶1 w/w) were formulated. After 9 weeks of feeding trial, fish fed all diets did not show significant differences in growth performances, feed utilization, and whole body compositions. Hemoglobin content gradually decreased (P<0.05) with increments of CS inclusion levels. There was a positive linear relationship between dietary gossypol and total liver gossypol contents. Free radical scavenging activity in the experimental diets gradually increased with increment of dietary CS inclusion. This finding indicates that the mixture of cottonseed and soybean meal with methionine and lysine supplementation could replace up to 40% of fishmeal protein in diets. However, 20% fishmeal protein replacement by dietary inclusion of cottonseed (9.4%) and soybean (8.7%) meal might be the optimum and safe level for the commercial use of CS in juvenile Japanese flounder.  相似文献   

6.
Larvae of jack mackerel Trachurus japonicus were sampled in seven depth layers (0–10 m, 10–20 m, 20–30 m, 30–40 m, 40–50 m, 50–70 m, and 70–100 m), with a closing-type frame net, on 26th–27th February, 2003, at Sta. V (27°50′N, 126°00′E) continental shelf edge in the southern part of East China Sea, to examine their vertical distribution. Most of the larvae (>90%) occurred in the upper 50 m layer during both day and night. In the daytime, larvae of length from 3.0 mm to 4.5 mm were most abundant in the 10–20 m layer, those from 5.0 to 6.0 mm were most abundant in the 20–30 m layer, those from 6.0 to 8.0 mm occurred at greater depth, in the 30–40 m layer, and those more than 9.0 mm were found in the 0–10 m layer and performed ontogenetic vertical migrations. Larvae from 6 mm to 8 mm length performed diurnal vertical migration, for the percentage (16.7%) of larval number in the 0–20 m layer in the daytime increased to 42.1% at night and the percentage (78.1%) in the 20–50 m layer in the daytime decreased to 49.0% at night.  相似文献   

7.
以相同规格的牙鲆Paralichthys olivaceus普通群体为对照,采用静水生物测试法开展了Hg2+对牙鲆克隆群体的急性毒性实验,实验所用幼鱼为130日龄,体长为(12.38±1.13)cm,体质量为(17.97±5.79)g。实验期间,水温为(21±1.2)℃,pH为8.1-8.3,盐度为31.0,硬度6200 mg/L,连续充气保证溶氧充足。研究表明,实验初始阶段,高浓度组克隆牙鲆在容器内躁动不安,上升到水面沿池壁缓慢转圈游动,一段时间后安静伏在池底;而普通牙鲆则仅个别鱼在水面游动,多数静止在池底不动。随中毒程度加深,用玻璃棒触碰普通牙鲆时,其反应剧烈,在水中侧翻、打转、急速游动或上下窜动,而克隆牙鲆则仅沿池底缓慢游动。最终克隆牙鲆和普通牙鲆均身体僵直,失去呼吸能力死亡。Hg2+对克隆牙鲆的24、48、96 h的半致死质量浓度分别为1.50、0.86、0.84 mg/L,而对普通牙鲆的半致死质量浓度则分别为1.93、1.04、0.90 mg/L。结果表明,克隆牙鲆对Hg2+的耐受性弱于普通牙鲆,对Hg2+更加敏感。从死亡过程看,各处理组的克隆牙鲆从出现死亡个体开始,死亡时间集中在8 h内,具有同步性;普通牙鲆则在实验期间陆续出现死亡,持续时间较长,同步性较差。  相似文献   

8.
In the Uranouchi Inlet, which has a commercial fishery of littleneck clam Ruditapes philippinarum, potentially toxigenic species of Dinophysis acuminata showed an annual occurrence except for the summer of 2000–2002. Relatively high abundances of D. acuminata were observed in April 2000 and November 2002. Some samples of D. acuminata cells, which were collected from the inlet in autumn 2002, contained pectenotoxin-2 (0.9–2.8 pg/cell) and okadaic acid (1.9–2.8 pg/cell). Cell length and hypotheca width of D. acuminata found in the Uranouchi Inlet were 33.7–45.0 and 21.0–31.6 μm, respectively. The length-width ratios ranged 1.21–1.71. Sequences of the 5.8S ribosomal RNA gene and flanking internal transcribed spacers, 1 and 2 of cells of D. acuminata from the inlet fell into a D. acuminata complex clade that included, D. acuminata-D. sacculus-D. acuta reported by others. In the inlet, occurrences of D. acuminata were detected within a broad water temperature range between 13.0 and 26.9°C, and those showing more than 25 cells/L were within 16.4–17.3°C for temperature and 27.6 and 34.1 for salinity. This is the first report that cells of Dinophysis acuminata from southern Japanese coastal waters contained okadaic acid and pectenotoxin-2.  相似文献   

9.
The stimulatory effect of fermented vegetable product (FVP) upon the phagocytic and superoxide generation of leukocytes was studied in the Japanese flounder Paralichthys olivaceus. The phagocytic activity of casein-induced, intraperitoneal leukocytes was investigated and quantified, that is the activity significantly increased (P<0.05 or <0.01) by the addition of FVP beyond 3 mg/kg body weight. Further analysis investigated the effect of FVP on superoxide generation in leukocytes. Established in vitro cytochrome c reduction assay was used to measure superoxide generation; reduced levels of FVP in assay samples had a profound effect on superoxide generation. FVP was also incorporated in commercial diets and fed to Japanese flounder for 4 weeks. The phagocytic activities and superoxide generation of peritoneal induced leukocytes were significantly higher (P<0.05, <0.01) in fish fed the FVP supplemented diet than fish fed the control diet. FVP feeding in fish had a significantly higher (P<0.05) activity of lysozyme than in the control fish.  相似文献   

10.
To clarify the feeding strategy of pelagic larvae of stone flounder in Mutsu Bay, the dietary composition and pray size was investigated from February to April during 1989–1999. Diets were compared with the numerical and volumetric compositions and frequency of occurrence of each prey species. Mensuration formulae were applied to estimate individual prey volume in the diet, while the length of planktonic species was measured from net and water samples. Prey shapes were assumed as sphere, cylinder, ellipsoid, pyramid, two elliptical cones, or a combination of ellipsoid and cylinder. Prey-size range increased as the larvae grew. Preflexion larvae fed mainly on copepod nauplii. Flexion and postflexion larvae ingested primarily appendicularians, with a suggestion that these larvae might depend on some parts of the microbial food web. Low frequencies of flexion and postflexion larvae with empty guts (1.7 and 1.4%, respectively) might be derived from feeding on slow-swimming appendicularians. From a size comparison between ‘house’-like organ length and trunk length of the appendicularian Oikopleura sp., almost all house-like organs with trunks in the larval diet were nonexpanded ‘house rudiments’, not expanded ‘houses’. Thus, stone flounder larvae may not chew the houses, but swallow the house rudiments with trunks.  相似文献   

11.
To estimate genetic parameters of growth traits in Japanese flounder Paralichthys olivaceus, full-sib and half-sib families were produced in three consecutive years at the Beidaihe Central Experiment Station in China. Each year 8–28 families were produced. The body weight, body length and body depth at 180, 240, and 360 days of age were measured for 5,224 individuals. Four animal models were used to examine the phenotypic variation of growth traits and were compared using the likelihood ratio test. The results showed that estimates for additive genetic effect heritabilities varied greatly depending on the model, trait and age. The maternal effect had a significant impact on phenotypic variation only for body depth at 180 days of age, which explained 49% of the phenotypic variance. The ratio of full-sib effect to phenotypic variation ranged from 0.09 to 0.22. Growth traits all exhibited low heritability (0.13–0.39), indicating that there is the potential for family selection breeding for these traits in Japanese flounder. Using the full model with the fixed, full-sib family, additive and maternal genetic effects, genetic correlations among the three traits for fish of the same age were estimated to be more than 0.80. Generally, the genetic correlations gradually increased as age increased.  相似文献   

12.
Near simultaneous exposure to selenium and salinity stress is an environmental reality faced by white sturgeon juveniles in the San Francisco Bay-Delta, having the potential for adverse synergistic impacts on juveniles during seaward migration. White sturgeon juveniles (36–132 g) were transferred into higher salinities (15–20 ppth) after exposure to dietary selenium (0.4, 9.6, 20.5, 41.7, 89.8, or 191.1 μg Se/g diet) in the form of l-selenomethionine (SeMet) for 8 weeks in freshwater (0 ppth). White sturgeon fed above 41.7 μg Se/g diet experienced significant (P < 0.05) declines in survival relative to sturgeon fed between 0.4 and 41.7 μg Se/g diet when transferred into brackish water (15 ppth salinity) for 3 days. Plasma osmolality and ionic concentrations in white sturgeon after abrupt transfer correlated positively with dietary Se concentrations. Transfer of white sturgeon fed above 41.7 μg Se/g diet into 20 ppth brackish water for 1 day resulted in significant declines in survival relative to white sturgeon fed 0.4–41.7 μg Se/g diet. Hematocrit and dehydrational weight loss were significantly correlated with dietary Se concentrations while plasma osmolality and ion concentrations were not. The transfer of sturgeon into 20 ppth water for 3 days resulted in severe survival declines in all treatment groups. The results of this study suggest that the previously established threshold for Se toxicity (10–20 μg Se/g) is sufficient to protect white sturgeon from the adverse interactive affects resulting from increases in dietary Se and ambient water salinity.  相似文献   

13.
A combination of eight isolates of Aeromonas hydrophila was used to produce monoclonal antibodies (MAbs). Ten different groups of MAbs specific to Aeromonas were selected. The first five groups of MAbs demonstrated high specificity and bound to only one or two isolates of A. hydrophila. The sixth and the seventh groups of MAbs were A. hydrophila specific. They recognized seven of eight A. hydrophila isolates (AH1, 2, 3, 4, 5, 6, 8); however, the MAb in the seventh group also showed cross‐reactivity to one isolate of Aeromonas caviae (AC3). The eighth MAb group recognized two isolates of A. hydrophila (AH2 and AH5) and demonstrated cross‐reactivity to one isolate of Aeromonas sobria (AS1) and one isolate of A. caviae (AC3). The tenth group of MAbs bound to all isolates of Aeromonas spp. tested (AH1‐8, AS1‐6, AC1‐5, Aeromonas veronii and Aeromonas jandaei) without cross‐reactivity to any of the other bacteria tested. MAbs in the ninth group showed similar specificity to those in the tenth group but did not recognize two isolates of A. sobria (AS4 and AS6) or A. jandaei. All the MAbs could be used to identify Aeromonas by dot blotting with a sensitivity ranging from 105 to 107 CFU mL?1. However, the sensitivity of detection was increased to 102–103 CFU mL?1 after inoculation of the sample in tryptic soy broth for 3–6 h before performing the dot blotting. The dot blot method can be used for the direct detection of A. hydrophila infection in symptomatic and asymptomatic goldfish. This study demonstrated a convenient immunological tool that can be used for the direct detection of A. hydrophila and Aeromonas infections in a complex sample without the requirement for separation of the bacteria or isolation and biochemical tests.  相似文献   

14.
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15.
ABSTRACT:     The acute toxicities of boron were examined for Japanese flounder Paralichthys olivaceus and red sea bream Pagrus major in terms of fish size and water temperature. Japanese flounder of 0.1–70.0 g and red sea bream of 0.6–20.3 g were exposed to different concentrations of boron for 96 h at 20°C under semi-static conditions. In both fish species, the median lethal concentration (LC50) for 96 h of boron increased linearly with increasing fish size, ranging from 108 to 252 mg B/L for the flounder, and from 97 to 172 mg B/L for the sea bream. The effect of water temperature on acute toxicity was examined for Japanese flounder of 0.6 and 1.5 g at 10, 15, 20 and 25°C, and for red sea bream of 0.6 and 2.4 g at 12, 15, 20 and 25°C. The toxicity of boron for the flounder increased linearly with increasing water temperature. The 96 h LC50 values ranged from 299 to 108 mg B/L for the 0.6 g flounder and from 350 to 113 mg B/L for the 1.5 g flounder. A similar trend was shown for the 2.4 g red sea bream; however, the relationship for the 0.6 g red sea bream was not significant.  相似文献   

16.
A feeding trial was conducted to investigate the effect of partial fish meal (FM) replacement by dietary soybean meal (SBM) on the growth and protein and lipid metabolism of the juvenile Japanese flounder. Four isonitrogenous and isolipidic diets (Diets 1–4) were formulated containing 47% protein and 9% lipid with different SBM inclusion (Diet 1, 11%; Diet 2, 16%; Diet 3, 24%; and Diet 4, 41%). The fish were fed to satiation twice daily for a feeding period of 56 days. The weight gain rate (WGR) and protein efficiency ratio of fish fed Diet 4 were significantly lower than those fed Diets 1, 2, and 3, and feed conversion ratio in the group fed Diet 4 was significantly higher than that in the groups fed Diets 1, 2, and 3. Hepatosomatic indices showed the similar trend as WGR, and the value (1.75%) for Diet 1 was significantly lower than that (1.96–2.2%) for other diets. There were no differences in whole body moisture, crude protein, crude lipid, and ash content among all treatments. With increasing dietary SBM level, serum triglyceride (TG), cholesterol (CHO), and low-density lipoprotein cholesterol (LDL-C) concentrations increased, whereas the serum total protein (TP) and high-density lipoprotein cholesterol (HDL-C) concentrations decreased. Compared with Diet 1, the serum TG, CHO, and LDL-C concentrations of fish fed Diet 4 significantly increased by 78, 37, and 36%, respectively, while the TP and HDL-C concentrations decreased by 14 and 33%, respectively. No significant differences in condition factor, blood urea nitrogen concentration, and alanine and aspartate aminotransferase activities were observed among the dietary treatments. These results indicated that dietary SBM inclusion above 24% could adversely affect the growth and protein and lipid metabolism of Japanese flounder.  相似文献   

17.
The effect of stocking density on the survival and growth of pikeperch, Sander lucioperca (L.), larvae was examined in two consecutive experiments. In experiment I, 4-day-old larvae [body wet weight (BW): 0.5 mg; total body length (TL): 5.6 mm] were reared in 200-l cylindro-conical tanks in a closed, recirculating system (20 ± 0.5°C) at three stocking densities (25, 50 and 100 larvae l−1) and fed a mixed feed (Artemia nauplii and Lansy A2 artificial feed) for 14 consecutive days. At densities of 25 and 100 larvae l−1, growth rate and survival ranged from 2.7 to 1.9 mg day−1 and from 79.2 to 72.3%, and fish biomass gain ranged from 0.6 to 2.0 g l−1, respectively. There were two periods of increased larval mortality: the first was at beginning of exogenous feeding and the second during swim bladder inflation. In experiment II, 18-day-old larvae (BW: 35 mg; TL: 15.6 mm) obtained from experiment I were reared under culture conditions similar to those of experiment I, but at lower stocking densities (6, 10 and 15 larvae l−1). The fish were fed exclusively with artificial feed (trout starter) for 21 consecutive days. At densities of 6 and 15 larvae l−1, the growth rate and fish biomass gain ranged from 28.8 to 23.1 mg day−1 and from 2.0 to 3.3 g l−1, respectively. The highest survival (56.5%) was achieved at a density of 6 larvae l−1. Mortality at all densities was mainly caused by cannibalism II type behaviour (27–35% of total). In both experiments, growth and survival were negatively correlated and fish biomass gain positively correlated with stocking densities. The present study suggests that the initial stocking density of pikeperch larvae reared in a recirculating system can be 100 individuals l−1 for the 4- to 18-day period post-hatch and 15 individuals l−1 for the post-19-day period.  相似文献   

18.
采用饱和硫酸铵分步盐析结合Sephacryl S-300凝胶过滤层析及DEAE Sepharose离子层析法,对牙鲆皮肤黏液中的免疫球蛋白(Ig)进行了分离纯化,并通过SDS-PAGE及Western-blotting技术对纯化蛋白的部分特性进行了分析。结果表明,30%、50%饱和硫酸铵分步盐析可以去除牙鲆黏液中除Ig外的很多杂蛋白,得到粗提黏液Ig;再经Sephacryl S-300纯化,Ig纯度较高,其中含有72和26 ku的条带;DEAE Sepharose层析法可进一步纯化Sephacryl S-300柱层析的产物,所提取黏液Ig经SDS-PAGE检测,只含有72和26 ku两个条带,初步认为是牙鲆黏液Ig的重链和轻链。Western-blotting结果显示,抗牙鲆血清Ig单克隆抗体可与黏液Ig重链72 ku条带发生发应。  相似文献   

19.
Habitat utilization, feeding, and growth of a rare pleuronectid flatfish, spotted halibut Verasper variegatus, were examined in a brackish lagoon in northeastern Japan: Matsukawa-ura. The distribution and date–length data of spotted halibut collected mainly from beam-trawl samplings during 1983–2008 indicated that age-0 juveniles [n = 25, 6.0–18.0 cm total length (TL)] and older spotted halibut (n = 71, 13.8–43.0 cm TL) inhabited almost the entire northern part of Matsukawa-ura. Comparative distribution surveys of spotted halibut, stone flounder, and marbled flounder during 1985–1989 revealed similar distribution patterns of these flatfishes; the highest densities were found around the central part of Matsukawa-ura, although significantly lower abundance was detected for spotted halibut. Linear growth equations of age-0 juveniles demonstrated that spotted halibut were able to achieve high growth after June, probably because abundant prey (e.g., mysids and gammarids) and suitable physical conditions (warmer water temperature and lower salinity) were present. The main prey items shifted from various crustaceans including mysids, caridean shrimps, and anomurans to brachyura Hemigrapsus spp. (≥20 cm TL). Our study shows that spotted halibut use the shallow brackish lagoon as an important nursery for juveniles, and also as a feeding ground for young and adults.  相似文献   

20.
Mouse monoclonal antibodies (MAbs) were produced by using yellow grouper nervous necrosis virus (YGNNV) as an immunogen, isolated from infected yellow grouper, Epinephelus awoara (Temminck & Schlegel), and propagated in GB cells. In enzyme linked immunosorbent assay (ELISA), 43 hybridoma clones secreting MAbs strongly reacted with the purified virus. Ten of them showed a higher neutralization index (NI) value between 6.5 and 4.5 (log10 NI) than the other 33 MAbs against YGNNV infection in cell culture. All 10 MAbs belonged to the IgG isotype with a κ light chain and recognized the 42 kDa coat protein of YGNNV by Western blot analysis. Immunohistochemical results demonstrated that the viral signals co-located with pathological lesions observed in retina, brain and spinal cord. These results indicate that the MAbs are useful for confirmative diagnosis of YGNNV infection.  相似文献   

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