Physiological roles of FSH and LH in red seabream, Pagrus major

The duality of gonadotropins (GTHs), follicle-stimulating hormone (FSH) and luteinizing hormone (LH), has been confirmed in most teleost species, but very little is known about their biological functions. To elucidate the physiological roles of FSH and LH in fish reproduction, the expression profiles of GTH subunit genes during gonadal development were analyzed in both male and female red seabream. Furthermore, in vitro studies were carried out to examine the effects of GTHs on steroid hormone production and cytochrome P450 aromatase (P450arom) expression in red seabream gonads. In both sexes, LHβ mRNA was maintained at high levels from the early gametogenesis until spawning season, and declined with gonadal regression. Interestingly, FSHβ mRNA levels in males increased in parallel with testicular development, whereas those in female were remained low throughout oocyte development. From in vitro studies using purified red seabream FSH and LH, both GTHs had a similar potency in stimulating 11-ketotestosterone production by testicular slices, while the biological activity of FSH was much lower than that of LH in stimulating production of estradiol-17β by vitellogenic follicles. Moreover, expression of P450arom mRNA was induced by LH, but not FSH, in ovarian follicles in vitro. FSH was also ineffective in inducing maturational competence and final oocyte maturation. These results suggest that, unlike salmonids, FSH may play an important role during gametogenesis in male, but not female, red seabream, whereas LH may be involved in regulation of both early and late gametogenesis in both sexes.     

Effects of gonadotropin-releasing hormone agonist and testosterone on gonadotropin subunit gene expression in female red seabream (Pagrus major)

We examined the effects of gonadotropin-releasing hormone agonist (GnRHa) and testosterone (T) on the level of gonadotropin subunit mRNAs in the pituitary of ovariectomized or intact female red seabream. Ovariectomy induced increase of seabream (sb) GnRH, glycoprotein (GP) α and luteinizing hormone (LH) β mRNA levels. GnRHa treatment also stimulated GPα and LHβ mRNA levels. T treatment reduced GPα and LHβ mRNA expression probably via negative feedback action on sbGnRH. Both GnRHa and T treatment had no effect on follicle-stimulating hormone (FSH) β mRNA levels. These results suggest that the regulatory mechanisms of GPα and LHβ gene expression differ from those of FSHβ gene.     

Regulation of gonadotropin subunit genes expression by 11-ketotestosterone during early spermatogenesis in male red seabream, Pagrus major

To examine the roles of gonadal steroids in the regulation of expression of gonadotropin subunit genes, male red seabream were gonadectomized and a sub-group was treated with 11-ketotestosterone (11-KT). Castration of males during the early stage of spermatogenesis elicited a significant increase in FSHβ mRNA levels, which was prevented by 11-KT replacement. By contrast, LHβ mRNA levels were not changed by castration or 11-KT replacement. In addition, administration of 11-KT to sham-operated males suppressed the steady-state FSHβ and LHβ mRNA levels. These results indicate that 11-KT may function as a negative feedback regulator of FSHβ gene expression, and may act through the testis to down-regulate LHβ mRNA levels in male red seabream during this period.     

The isolation of intestinal microflora of farmed red seabream (Pagrus major) and black seabream (Acanthopagrus schlegeli) at larval and juvenile stages

The aerobic bacterial flora in the intestine of farmed red seabream and black seabream was investigated at their larval and juvenile stages in relation to the microflora of ambient water and diets. The total bacterial number in the intestine was 7.4×10⁴ and 3.4×10⁴ colony-forming units (CFU)/fish for red seabream and black seabream, respectively. Among the intestinal flora, Vibrio (ca. 45%) and Pseudomonas (30%) were the most dominant bacteria.The total bacterial counts were 3.1×10⁴ CFU/ml for rearing water, 2.1×10⁷ CFU/g for live diets (rotifer and brine shrimp) and 1.2×10⁴ CFU/g for artificial diets. Among the water bacteria, Pseudomonas (22%) and Moraxella (29%) constituted dominant groups, and in the live diets Pseudomonas (48%) was predominant. The incidence of Vibrio in waters and live diets was 7% and 11%, respectively.     

Evaluation of canola oils as alternative lipid resources in diets for juvenile red seabream, Pagrus auratus

This study examined three potential oil resources, crude and refined canola oil and refined soybean oil as replacements for added dietary fish oil in diets for juvenile red seabream. These oil resources were evaluated for their potential to replace added fish oil (40 g kg^?1) in fishmeal based (600 g kg^?1) diets, with 100 g kg^?1 of total lipids. Each of the three plant oils was used to replace 25%, 50%, 75% or 100% of the added dietary fish oil. Each of the three plant oils showed potential as a replacement for dietary fish oil, although a significant reduction in growth and feed utilisation was observed with the complete (100%) replacement of added fish oil by crude canola oil. No other significant effects of oil type or inclusion level on growth were apparent. A negative control (no added fish oil or plant oil, 60 g kg^?1 of total lipid) yielded poorer growth than all treatments except the diet containing 40 g kg^?1 of added crude canola oil (100% replacement). This observation confirmed that the added oils were utilized by the fish. A positive control diet containing 80 g kg^?1 of added fish oil (140 g kg^?1 total dietary lipid) sustained the best growth in the study, confirming that the 13 experimental diets were energy limiting as planned. Notably, few effects of the alternative oils were seen on the proximate composition of the fish. However, the influence of the alternative oils on the tissue fatty acid composition was considerable, irrespective of plant oil type or processing grade. Particularly notable was the overall increase in the level of polyunsaturated fatty acids in the tissues of the fish fed the plant oil diets, with these trends becoming more apparent with the greater levels of fish oil replacement. Minimal reductions in the levels of the long‐chain polyunsaturated fatty acids of eicosapentaenoic (20:5n‐3) and docosahexaenoic (22:6n‐3) acid were observed from any of the plant oil treatments. Sensory assessment, by an Australian taste panel, of the fish fed the fish oil reference, or the 100% replacement by refined canola or refined soybean diets showed a preference in order of canola oil > soybean oil > fish oil fed fish. Clearly, both canola and soybean oils have considerable potential as replacements of fish oils in diets for this species.     

真鲷仔稚鱼对实验微粒饲料中卵磷脂适宜需要量的研究

制备了4种n-3HUFA含量基本相等、卵磷脂含量分别为0、2．5％、5．0％、7．5％的实验微粒饲料，探讨了卵磷脂的含量对真鲷仔稚鱼生长、存活、体内相关成分以及对外部压力的耐受性的影响。25d的养殖试验结果表明，当真鲷仔稚鱼的实验微粒饲料中卵磷脂含量不超过5．0％时，随着实验微粒饲料中卵磷脂含量的增加，真鲷仔稚鱼的生长速度和成活率也相应增加。养殖试验结束后，对仔稚鱼体内相关成分的分析结果表明，仔稚鱼体内n-3HUFA和DHA的含量随着实验微粒饲料中卵磷脂含量的增加而增加。但当卵磷脂含量从5．0%增加到7．5％时，真鲷仔稚鱼的生长速度、成活率以及体内n-3HUFA和DHA的含量却没有相应增加，因此，真鲷仔稚鱼对实验微粒饲料中卵磷腊的适宜需要量应为5．0％左右。活力测试结果表明，当真鲷仔稚鱼的实验微粒饲料中卵磷脂的含量不超过5．0％时，随着实验微粒饲料中卵磷脂含量的增加，真鲷仔稚鱼对外部压力的耐受性增强，但是当实验微粒饲料中卵磷脂的含量从5．0％增加到7．5％时，真鲷仔稚鱼对外部压力的耐受性并没有继续增强，从而进一步说明真鲷仔稚鱼对实验微粒饲料中卵磷脂的适宜需要量应为5．0％左右。     

Effects of dietary inactivated Pediococcus pentosaceus on growth performance,feed utilization and blood characteristics of red sea bream,Pagrus major juvenile

A feeding trial was carried out to determine effects of dietary supplements of inactivated Pediococcus pentosaceus (PP) on growth, feed utilization and blood characteristics of red sea bream. Five diets containing different concentrations [0 (PP0), 1.6 × 10¹⁰ (PPI), 1.6 × 10¹¹ (PPII), 1.6 × 10¹² (PPIII) and 3.2 × 10¹² (PPIV) cells g^?1] of PP were fed to fish for 56 days. At the end of the feeding trial, the obtained results revealed a significant increase (P < 0.05) in weight gain, specific growth rate and skin mucus secretion in all PP‐supplemented groups when compared with control group. Moreover, fish fed PPIII and PPIV diets resulted in significantly higher final body weight, feed efficiency ratio and bactericidal activity than those of fish fed PP0 diet (P < 0.05). Further, fish fed the PPIII diet also had a higher feed intake, protein efficiency retention, haematocrit, mucus lysozyme activity (LA), plasma and serum proteins than those of PP0 fed group (P < 0.05). Similarly, fish fed PPIV diet resulted in a significant increase in serum LA, peroxidase activity and tolerance against low‐salinity stress test (P < 0.05). Considering these promising results, we suggest that the supplementation of PP Prep improves growth and health conditions of Pagrus major.     

Biosynthesis of estradiol-17β in the ovarian follicles of the red seabream Pagrus major during vitellogenesis

ABSTRACT: The red seabream Pagrus major is a useful experimental fish for studying the endocrine control of oogenesis in teleosts. This study investigated the steroidogenic pathway for estradiol-17β (E2) biosynthesis in the ovarian follicles of red seabream. Intact follicles were isolated during vitellogenesis and incubated in vitro with different radiolabeled steroid precursors. When 17-hydroxyprogesterone (17-P), dehydroepiandrosterone (DHEA), or androstenedione (AD) were used as precursors, both testosterone (T) and estrone (E1) were synthesized by follicles, leading to estradiol-17β (E2) production. Serum steroid levels measured by enzyme-linked immunosorbent assay showed that T, E1, and E2 were present in the circulation at levels ranging from 1 ng/mL to 2 ng/mL throughout the day during the spawning season. In vitro conversion of E1 into E2, however, was 15.8-fold greater than T conversion into E2, suggesting that E2 is synthesized mainly via E1 rather than T. The results showed that E2 was synthesized from pregnenolone via 17-hydroxypregnenolone, DHEA, AD, and E1. Thus, the study demonstrated the complete steroidogenic E2 synthesis pathway in the ovarian follicles of red seabream, and revealed that E1 is the major precursor of E2.     

Identification of a heparin-binding,mesoderm-inducing peptide in the swim-bladder of the red seabream,Pagrus major: a probable fish fibroblast growth factor

The brain, pituitary, liver, ovary and swim-bladder of the red seabream (Pagrus major) were screened for the presence of a growth factor capable of stimulating the proliferation of BHK-21 cells, and the swimbladder extract was found to exert the most powerful activity. Swim-bladder derived growth factor (SBDGF) with heparin-binding ability was isolated by heparin-affinity and size-exclusion chromatography using heparin-Sepharose CL-6B and Superose 12, respectively. The relative molecular mass of SBDGF was 22,500 and the isoelectric point was 9.4. The growth of mammalian fibroblasts (BALB/3c 3T3, BHK-21) and endothelial cells (CPAE) was markedly stimulated by SBDGF. Its growth-promoting activity was also effective in fish fibroblasts (BF-2 and RTG-2), and was heat- and acid-labile. In the mesoderm-induction assay, explants of presumptive ectoderm dissected from Xenopus laevis blastulae (stage 9) were incubated with SBDGF. SBDGF induced ventral mesoderm in the explants, including hemocyte-like cells, mesenchyme and coelomic epithelium. These properties suggest that SBDGF is a fish fibroblast growth factor and that the swim-bladder is the source.     

水温对施氏鲟幼鱼消化酶活力的影响

分别在14℃、21℃、28℃水温条件下饲养施氏鲟(Acipenser schrenckiiBrandt)幼鱼10 d,初始鱼体质量(124.20±15.12)g,体长(26.95±2.89)cm,检测不同水温条件下施氏鲟幼鱼消化器官(胃、幽门盲囊、十二指肠、瓣肠和肝脏)蛋白酶、淀粉酶和脂肪酶的活力。结果表明:(1)幼鱼幽门盲囊、瓣肠和肝脏中蛋白酶活力均以水温21℃时最高(P<0.05),十二指肠和胃则在28℃最高(P<0.05),各消化器官蛋白酶活力均在14℃最低(P<0.05);(2)各消化器官的淀粉酶活力均在水温21℃时最高(P<0.05),幽门盲囊、瓣肠和十二指肠以水温28℃时最低(P<0.05),胃和肝脏在水温14℃时最低(P<0.05);(3)各消化器官脂肪酶活力均在水温14℃时最高,28℃时最低(P<0.05)。在水温21℃时,各消化器官蛋白酶和淀粉酶活力由大到小依次为:幽门盲囊、瓣肠、十二指肠、胃、肝脏,水温14℃时瓣肠消化蛋白质和碳水化合物能力要高于十二指肠,28℃时低于十二指肠(P<0.05);在水温14℃和21℃时各消化器官脂肪酶活力由大到小依次为:瓣肠、十二指肠、胃、肝脏、幽门盲囊,28℃时瓣肠消化脂肪能力低于十二指肠(P<0.05)。[中国水产科学,2007,14(1):126-131]     

Polymorphisms and haplotypes of the myostatin gene associated with growth in juvenile red sea bream Pagrus major

Polymorphisms of the myostatin gene (MSTN) have been studied in vertebrates including several aquaculture species, revealing their role in growth. We attempted to identify polymorphisms in MSTN associated with growth traits of juvenile farmed red sea bream Pagrus major, an important cultured fish species in Japan. Polymorphisms in the coding region of MSTN were screened, and six polymorphisms were found among three exons: a deletion in exon 1, a single nucleotide polymorphism (SNP) in exon 2 and four SNPs in exon 3. The deletion in exon 1 eliminated a codon (glutamine) from the region comprising the polyglutamine structure, but all SNPs were synonymous. We analysed the polymorphisms for association with growth phenotype in large and small P. major specimens obtained from commercial production at 50 days post hatching. Two SNPs located in exon 3 (c.+846T>C and c.+1140T>C) were significantly more frequent in the large group, while no SNPs were significantly associated with the small phenotype. Haplotypes were reconstructed using genotype information of the two phenotype groups, and nine haplotypes (Hap_1 to Hap_9) were successfully reconstructed. Hap_6 and Hap_7 were significantly more often observed in small and large groups respectively. Analysis of diplotypes (haplotype combinations) of individual specimens revealed 19 diplotypes. The Hap_7/Hap_7 diplotype was found in 46.7% of large phenotype specimens, significantly more frequently than in the small group (13.5%). These results will be useful for marker‐assisted selection of red sea bream to improve production with respect to growth.     

水温对细鳞鱼幼鱼消化酶活性的影响

1在水温6℃、14℃、22℃下和180×60×50cm控温水族箱中,饲养初始鱼体质量11.3±0.94g的细鳞鱼Brachymystax lenok幼鱼21d,测定了幼鱼消化器官（胃、幽门盲囊、肝脏和肠）中的蛋白酶、淀粉酶和脂肪酶的活力。结果表明：水温14℃下胃蛋白酶活性最高,其次为水温6℃时,22℃时消化酶活性最低,...     

水温对细鳞鱼幼鱼消化酶活性的影响

1在水温6℃、14℃、22℃下和180×60×50cm控温水族箱中,饲养初始鱼体质量11.3±0.94g的细鳞鱼Brachymystax lenok幼鱼21d,测定了幼鱼消化器官（胃、幽门盲囊、肝脏和肠）中的蛋白酶、淀粉酶和脂肪酶的活力。结果表明：水温14℃下胃蛋白酶活性最高,其次为水温6℃时,22℃时消化酶活性最低,显著低于前2个处理组（P〈0.05）。肠中脂肪酶活性最高,显著高于胃和肝中（P〈0.05）,其次为幽门盲囊。各消化器官脂肪酶活力均在14℃时最高,其中肠和幽门盲囊中脂肪酶活力显著高于其他2个处理组。各水温条件下,幽门盲囊中淀粉酶活性最高,其次为肠。6℃时,幽门盲囊中淀粉酶活性显著高于胃和肝脏中（P〈0.05）;14℃和22℃时,幽门盲囊和肠中淀粉酶活性显著高于胃和肝脏中（P〈0.05）,胃和肝脏中差异不显著（P〉0.05）。14℃时各消化器官淀粉酶活力最高,6℃时肝脏和肠中淀粉酶活性显著低于14℃时,其它组织中各处理组间淀粉酶活力差异不显著（P〉0.05）。     

摄食促进物质对真鲷摄食和生长的影响

将甘氨酸 丙氨酸 组氨酸 精氨酸、甘氨酸 丙氨酸 组氨酸 精氨酸 甜菜碱、甘氨酸 丙氨酸 缬氨酸、甘氨酸 丙氨酸 缬氨酸 甜菜碱作为摄食促进物质，分别添加到真鲷配合饲料中饲喂真鲷，探讨摄食促进物质对真鲷摄食和生长的影响。结果表明，添加摄食促进物质不仅可促进真鲷的的摄食性，增加其摄食量，还可以提高真鲷的增重率、饲料效率、蛋白效率。添加甜菜碱对促进真鲷的摄食性无作用，但可促进真鲷的生长。     

温度对点篮子鱼幼鱼生长、摄食和消化酶活性的影响

研究了不同养殖温度(19℃、23℃、27℃、31℃)对点篮子鱼(Siganus guttatus)幼鱼生长、摄食和肠道消化酶活性的影响。结果表明,实验期间各温度组幼鱼存活率(SR)均达到95%以上,19℃组存活率显著低于其余各组(P0.05);在19~31℃范围内,幼鱼的特定生长率(SGR)和相对增重率(WGR)随温度的升高而显著增加(P0.05),在31℃时达到最高值;不同温度组的体质量增长速度由高到低依次为31℃27℃24℃19℃,31℃和27℃温度组体质量呈二项式增长,23℃和19℃组呈线性增长;随着温度升高,其饵料系数(FCR)逐渐降低(P0.05),回归分析显示在29.89℃时达到最小值;摄食率(FR)随着温度的升高而显著升高(P0.05),曲线分析得其在31.74℃时达到最大值。温度对幼鱼肠道胰蛋白酶活性影响显著,随温度降低呈逐渐升高的趋势,19℃组胰蛋白酶活性显著高于27℃和31℃组(P0.05);温度对幼鱼肠道脂肪酶活性无显著性影响(P0.05);温度对幼鱼肠道淀粉酶和麦芽糖酶活性影响显著(P0.05),在23℃时达到最大值。综合以上结果认为,点篮子鱼幼鱼快速生长的适宜温度范围为29.89~31.74℃,在此温度范围内点篮子鱼幼鱼可获得较大的生长率。     

温度对施氏鲟幼鱼摄食、生长和肠道消化酶活性的影响

研究不同养殖温度(15℃、18℃、21℃、24℃、27℃)对施氏鲟(Acipenser schrenckii)幼鱼摄食、生长和肠道消化酶活性的影响。结果表明,在15～24℃范围内,幼鱼的特定生长率(SGR)和相对增重率(RWG)随温度的升高而显著增加(P<0.05),均在24℃时达到最高值;随着温度升高,其饵料系数(FC)先降低后升高(P<0.05),且在24℃时达到最小值;此外,摄食率(FR)随着温度的升高而显著升高(P<0.05),其不同温度组的生长速度由高到低依次为24℃、21℃、27℃、18℃、15℃组。温度对幼鱼前肠蛋白酶活性影响显著,24℃组和27℃组前肠的蛋白酶活性显著低于15℃组(P<0.05),而温度对中肠和后肠蛋白酶活性则无显著性影响(P>0.05);温度对幼鱼肠道脂肪酶活性无显著性影响(P>0.05),各温度组前肠脂肪酶活性均高于中肠和后肠;温度对幼鱼肠道淀粉酶活性无显著性影响(P>0.05)。综合以上结果认为,施氏鲟幼鱼快速生长的适宜温度范围为21～24℃,在此温度范围内施氏鲟幼鱼可获得最大生长率和存活率。     

Survival, growth and gonad development of triploid red sea bream, Pagrus major (Temminck et Schlegel): use of allozyme markers for ploidy and family identification

Abstract. Triploidy was induced in three full-sib families of sea bream, Pagrus major (Temminck & Schlegel), by subjecting the fertilized eggs to cold-shock treatment at 0°C for a 12 min duration, starting 3 min after insemination. Results showed that this treatment gave 100% triploid fish with reasonable survival rates of more than 61% up to first feeding, Triploidy induction was proved by the use of allozyme markers at three ( ADH *, EST * and 6PGD *) loci and red blood cell size analysis.
At 2 months of age, the survival of the triploid group was lower than that of the diploids but it was comparable at 10 months. No differences in growth rate between diploid and triploid full-sibs were observed up to the age of 10 months. However, differences in body weight between families were noticed both in diploid and triploid groups when cultured in the same net cage. This possibly suggests a high heritability for the trait examined in the broodstock. The gonads of the triploid fish were significantly smaller than those of the diploids, Histological analysis of the gonads revealed sterility in triploid fish.     

Effect of dietary taurine enhancement on growth and development in red sea bream Pagrus major larvae

This study investigated the effect of feedings taurine‐enriched rotifers on the growth and development of larval red sea bream (RSB). Rotifers incubated in taurine‐enriched water at a taurine concentration of 800 mg L^?1 (T‐800) and 0 mg L^?1 (T‐0) were fed to larvae from 3 to 20 days after hatching (DAH). Notochord length, body weight and specific growth rate of T‐800 group were significantly greater than those of T‐0 at 14, 17, 9–11 and 18–20 DAH. Taurine content of larvae in the T‐800 group increased rapidly from 11 DAH and thereafter remained significantly higher than T‐0. Flexion larvae firstly appeared in both groups at 8 DAH, however, at 20 DAH post‐flexion larvae were significantly more abundant in T‐800 than T‐0. While nucleic acid and protein contents (μg mg^?1 wet fish) showed remarkable changes, ontogenetic growth in RSB larvae stage was observed to switch from hyperplastic growth to hypertrophic growth with the start of the flexion stage. Although a similar change in nucleic acid contents was observed between the two groups, the protein content (μg fish^?1) and protein/DNA ratio of T‐800 remained higher than that of T‐0 during the hypertrophic growth period. These results suggest that dietary taurine accelerates the growth and development in RSB larvae especially during hypertrophic growth (flexion stage) after the early hyperplastic growth.     

Purification of gonadotropins (PmGTH I and II) from red seabream (Pagrus major) and development of a homologous radioimmunoassay for PmGTH II

Two gonadotropic glycoproteins (PmGTH I and II) were purified by ion-exchange chromatography, gel filtration and preparative SDS-PAGE, from pituitaries of red seabream, a marine teleost which has an asynchronous-type ovary and spawns almost daily during the spawning season. The glycoproteins were composed of distinct subunits and the molecular weights were estimated to be 32 and 38 kDa for PmGTH I and PmGTH II, respectively. Both PmGTH I and II were active in two homologous bioassays: in vitro oocyte maturation and/or in vitro estradiol-17 production assays. These two GTHs were distinct in electrostatic properties, molecular weight, stability and yields from pituitaries during the spawning season. These properties suggest that PmGTH I and II correspond to salmon GTH I and II, respectively.A homologous radioimmunoassay with which to measure PmGTH II was developed using a rabbit antiserum against the subunit of PmGTH II and intact PmGTH II as standards and radioactive competitors. Competition curves for red seabream plasma and pituitary extract were parallel to the standard curve, while PmGTH I had low cross-reactivity (3.1 %) with the antibody. This specific RIA system showed an in vivo LHRHa induced GTH surge in the plasma of female red seabream.