Urea and trimethylamine oxide in diets for seawater farmed rainbow trout: effect on fat belching,skin vesicle,winter ulcer and quality grading

Three experiments were conducted in order to evaluate the effects of urea and trimethylamine oxide (TMAO) supplementation in diets for seawater‐farmed rainbow trout Oncorhynchus mykiss. In all experiments, commercial extruded feed was used as basal control diet, whereas experimental diets were the basal diet supplemented with either 10 g kg^–1 TMAO, 10 g kg^–1 urea or 10 g kg^–1 TMAO and 10 g kg^–1 urea. The experiments were conducted both in research units and in a commercial fish farm. Quality was evaluated in three different commercial slaughterhouses, sorting the rainbow trout into superior, ordinary or processing grade. Significantly more TMAO was observed in muscle of rainbow trout given dietary supplementation of TMAO. However, the levels were low compared with levels in marine fish in general. Trimethylamine (TMA) was not found. No increase in muscle urea content was observed in trout fed urea in the diet. Significantly reduced fat belching, seen as reduced number of fat droplets surfacing in the net pen, was observed in groups given TMAO‐supplemented feeds. At least twice as many rainbow trout with skin vesicles were observed in the control group as in the group fed urea‐supplemented diet. In addition, skin vesicles developed into open sores in the winter only in rainbow trout fed control diet. In a large‐scale trial, quality grading of almost 200 000 market‐sized rainbow trout showed significantly more (88 vs. 83%) trout of superior grade in the group fed dietary urea compared with fish fed control diet. The main reason for downgrading from superior to ordinary and processing grade was skin lesions, supporting the observations made in the experimental units.     

Comparison of NADH-dependent cytochrome b5 reductase activity and in vitro methemoglobin induction by sodium nitrite in Oncorhynchus mykiss, Salmo salar, and Salvelinus fontinalis

Methemoglobin is hemoglobin containing ferric iron. Methemoglobin cannot bind to oxygen and at high concentrations causes tissue hypoxia. Brook trout (Salvelinus fontinalis) develop significantly greater methemoglobinemia than Atlantic salmon (Salmo salar) or rainbow trout (Oncorhynchus mykiss) following general anesthesia with benzocaine or tricaine methanesulfonate. The objective of this study was to compare the activity of the major methemoglobin reducing enzyme, NADH-dependent cytochrome b5 reductase (CB5R), in brook trout erythrocytes to the activity of CB5R in Atlantic salmon and rainbow trout erythrocytes. Methemoglobin levels were compared using co-oximetry following in vitro incubation of erythrocytes with sodium nitrite (NaNO₂). The CB5R activity was measured using a ferricyanide assay. There was significantly greater methemoglobin at time 0 in brook trout erythrocytes than in rainbow trout or Atlantic salmon erythrocytes (2.79 ± 0.29 %, 2.19 ± 0.23 %, 2.08 ± 0.14 %), (P < 0.001). There was significantly greater methemoglobin induction by NaNO₂ in brook trout erythrocytes (33.14 ± 3.32 %) than in rainbow trout or Atlantic salmon erythrocytes (28.73 ± 2.92 % and 24.85 ± 1.40 %, respectively), (P < 0.001). The CB5R activity was significantly less in brook trout erythrocytes (median of 3.05 μmol/min/μl) than in rainbow trout erythrocytes (median of 6.73 μmol/min/μl). The CB5R activity in Atlantic salmon erythrocytes (median 4.09 μmol/min/μl) was not significantly different than in brook or rainbow trout erythrocytes. Total methemoglobin at any one time is a balance between induction by oxidants and reduction by antioxidants. Lower CB5R activity in brook trout erythrocytes may contribute to a species-specific sensitivity to methemoglobin induction; however, there are likely additional factors.     

草鱼肝微粒体的提取及CYP酶活性的测定

CYP酶代谢是药物生物转化的主要途径,其数量和活性大小直接影响药物在体内的活化与代谢。我们对草鱼肝微粒体CYP酶含量及其活性进行了初步研究,以差速离心法提取草鱼肝微粒体,以CO还原差示光谱法测得CYP酶及细胞色素b5含量分别为0.619±0.102 nmol/mg、0.264±0.042 nmol/mg。以7-乙氧异吩噁唑酮-O-脱乙基反应、苯胺-4-羟化反应、氨基比林-N-脱甲基反应作为CYP1A、CYP2E、CYP3A的探针反应,测得EROD酶活为0.043±0.004 nmol/mg/min,ANH酶活为0.028±0.002 nmol/mg/min,AMND酶活为0.207±0.035 nmol/mg/m in。结果表明草鱼肝微粒体中CYP酶发育完好,并且具有参与药物代谢的3种主要亚型活性,其含量与活性大小与其它实验动物相差较大。本实验的方法与结果为草鱼CYP酶的系统研究提供可靠手段,最终为指导水产合理用药提供理论依据。     

Seasonal variations of trimethylamine oxide and urea in the blood of a cold-adapted marine teleost,the rainbow smelt

Trimethylamine oxide (TMAO), which has previously not been known to occur in significant amounts in the blood of marine teleosts, rose to concentrations of approximately 50 mM in the blood of winter-acclimatized rainbow smelt, Osmerus mordax. Urea also increased in the blood of cold-acclimatized smelt, and, with TMAO, contributed significantly to the winter freezing point depression. TMAO and urea also varied seasonally in muscle and liver tissues. TMAO and urea appeared to be reabsorbed from the urine. Losses of TMAO and urea from the head region of the fish, where most of the losses appeared to occur, were approximately 9 μmol and 8 μmol 100 g⁻¹ h⁻¹, respectively. Despite the effluxes, TMAO and urea levels in both the blood and muscle either increased or were maintained in starved, cold-acclimated fish, suggesting that they were synthesized in response to cold temperature. TMAO was also found in the blood of some other cold-hardy teleosts.     

Production Characteristics of Rainbow Trout,Oncorhynchus mykiss and Brook Trout,Salvelinus fontinalis Under Seasonal Pond Conditions

Abstract

The production characteristics of juvenile rainbow trout, Oncorhynchus mykiss and brook trout, Salvelinus fontinalis were compared under winter pond conditions. Juvenile rainbow trout (55.1 ±1.5 g) and brook trout (28.9 ±0.4 g) were stocked at a density of 8,750 fish/ha into six 0.04-ha ponds. After 163 days, survival, growth, and feed conversion were similar (P >0.05). The results of this study suggest that brook trout may attain growth rates similar to rainbow trout under winter pond conditions in temperate regions of North America.     

Evaluation of AQUI-STM (efficacy and minimum toxic concentration) as a fish anaesthetic/sedative for public aquaculture in the United States

A preliminary evaluation of efficacy and minimum toxic concentration of AQUI-S^TM, a fish anaesthetic/sedative, was determined in two size classes of six species of fish important to US public aquaculture (bluegill, channel catfish, lake trout, rainbow trout, walleye and yellow perch). In addition, efficacy and minimum toxic concentration were determined in juvenile–young adult (fish aged 1 year or older) rainbow trout acclimated to water at 7°C, 12°C and 17°C. Testing concentrations were based on determinations made with range-finding studies for both efficacy and minimum toxic concentration. Most of the tested juvenile–young adult fish species were induced in 3 min or less at a nominal AQUI-S^TM concentration of 20 mg L^–1. In juvenile–young adult fish, the minimum toxic concentration was at least 2.5 times the selected efficacious concentration. Three out of five species of fry–fingerlings (1.25–12.5 cm in length and < 1 year old) were induced in ≤ 4.1 min at a nominal concentration of 20 mg L^–1 AQUI-S^TM, with the other two species requiring nominal concentrations of 25 and 35 mg L^–1 for similar times of induction. Recovery times were ≤ 7.3 min for all species in the two size classes. In fry–fingerlings, the minimum toxic concentration was at least 1.4 times the selected efficacious concentration. There appeared to be little relationship between size of fish and concentrations or times to induction, recovery times and minimum toxic concentration. The times required for induction and for recovery were increased in rainbow trout as the acclimation temperature was reduced.     

Fertilization rate,motility, lipid peroxidation and pH changes after chilled storage of rainbow trout (Oncorhynchus mykiss) eggs and spermatozoa by a RMPI medium

The aim of this study was to evaluate the effects of Roswell Park Memorial Institute (RPMI) 1,640 medium on chilled storage of eggs and spermatozoa of rainbow trout (Oncorhynchus mykiss). After 3 days of storage, eggs in RPMI 1,640 media (pH 8.2, 9 and 10), Cortland medium and coelomic fluid were inseminated with fresh spermatozoa (Experiment 1). Eggs in RPMI 1,640 medium at pH 8.2 shown the lowest thiobarbituric acid‐reactive substances (TBARS, 0.053 ± 0.003 nmol/ml) and pH changes (from 8.20 ± 0.01 to 8.18 ± 0.01), the highest fertilization rate (82 ± 3%). Undiluted and diluted spermatozoa at ratios of 1:2 and 1:9 with RPMI 1,640 media (pH 8.2, 9 and 10) and Cortland medium were inseminated with fresh eggs (Experiment 2). Spermatozoa in RPMI 1,640 medium at pH 9 (1:9) caused the lowest TBARS content (0.037 ± 0.002 nmol/ml) and pH changes (from 9.00 ± 0.01 to 8.98 ± 0.01), the highest fertilization rate (77 ± 2%) and motility parameters. Based on Experiments 1 and 2, eggs and spermatozoa were stored for another 3 days in RPMI 1,640 medium at pH 8.2 and 9 (1:9) respectively (Experiment 3). Fertilization rate of storage eggs and spermatozoa in Experiment 3 was 79 ± 5%, showing successfully storage of rainbow trout gametes with the same medium.     

Distribution of 5-hydroxytryptamine and related compounds in various brain regions of rainbow trout (Oncorhynchus mykiss)

The levels of tryptophan (Try), 5-hydroxytryptamine (serotonin, 5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) were determined in the brain regions of rainbow trout (Oncorhynchus mykiss) by high performance liquid chromatography with electrochemical detection (HPLC-EC). Brain tryptophan concentrations varied from 3.972 ± 357 ng/g cerebellum) to 8.841 ± 772 ng/g (hypothalamus). The 5-HT concentrations varied from 69 ± 7 ng/g (optic tectum) to 573 ± 34 ng/g (hypothalamus). The concentrations of 5-HIAA varied from 29 ± 3 ng/g (medulla oblongata) to 68 ± 7 ng/g (hypothalamus). Total and free serum tryptophan levels were also determined; in adult rainbow trout 92% of the serum tryptophan was observed to be free i.e., not protein-bound.     

Effects of dietary β-carotene on the immune response of rainbow trout Oncorhynchus mykiss

SUMMARY: We attempt to assess the impact of different levels of dietary β-carotene on immune function in rainbow trout. Semi-purified diets containing 0, 40, 200, and 400 mg β-carotene/kg dry diet were fed for 12 weeks to fish with average weight of 45 g. In addition to the humoral and cellular immune parameters, growth and feed utilization were examined. There were no marked differences in growth and feed utilization showing that β-carotene was not particularly efficient in enhancing growth of rainbow trout. Of the immune parameters measured, total immunoglobulin was significantly highest for the 200 mg β-carotene fed group. Serum complement activity (alternate pathway) at 200 and 400 mg β-carotene supplementation was significantly higher than that of the unsupplemented group. An increasing trend in lysozyme activity was observed, however, the differences among the groups were not significant. Phagocytic activity was similar among diet groups except at the highest level of supplementation where it was the maximum. Oxygen radical production by peripheral blood leukocytes appeared to be lower at higher levels of carotenoid supplementation. Overall, dietary β-carotene clearly enhanced immune response parameters in rainbow trout such as serum complement activity and total plasma immunoglobulin but did not show a definite influence for the other factors examined in the present study.     

NADH-dependent cytochrome b5 reductase and NADPH methemoglobin reductase activity in the erythrocytes of Oncorhynchus mykiss

Methemoglobin is oxidized hemoglobin that cannot bind to or dissociate from oxygen. In fish, it is most commonly caused by exposure to excess nitrites and can lead to abnormal swimming, buoyancy, or death. The methemoglobin concentration in mammals is determined by the balance of oxidizing agents versus reducing enzymes in erythrocytes. The objective of our studies was to characterize the enzymes that reduce methemoglobin in fish erythrocytes. Whole blood was collected from healthy rainbow trout. Methemoglobin was induced in vitro by NaNO₂ exposure. Methemoglobin reduction in controls was compared to reduction in samples with added NADH, NADPH, or NADPH and methylene blue. Rainbow trout whole blood was also fractionated into cytosol, microsomal, and mitochondria/plasma membranes/nuclei fractions. The fractions were compared for NADH-dependent cytochrome b5 reductase (CB5R) activity and for nitrite induction of methemoglobin. The CB5R activity in rainbow trout erythrocytes was compared to the CB5R activity in equine, feline, and canine erythrocytes. Rainbow trout erythrocytes had significant NADPH methemoglobin reductase activity in the presence of methylene blue (P < 0.001). The CB5R activity was greatest (P < 0.001) in the plasma membrane/mitochondria/nuclei fraction. The CB5R activity in rainbow trout erythrocytes was not significantly different than canine or equine activity but was significantly lower than feline CB5R activity (P < 0.0001). Methemoglobin in rainbow trout erythrocytes can be reduced by CB5R or NADPH-dependent methemoglobin reductase. Unlike mammalian anuclear erythrocytes, which are dependent on soluble CB5R, the nucleated RBCs of rainbow trout use membrane-bound CB5R to reduce methemoglobin.     

The non-specific activation of rainbow trout, Salmo gairdneri Richardson, complement by Aeromonas salmonicida extracellular products and the correlation of complement activity with the inactivation of lethal toxicity products

Abstract The possible mechanism of inactivation of the toxicity of Aeromonas salmonicida extracellular products (ECP) by normal rainbow trout serum was investigated using juvenile rainbow trout. ECP was prepared from culture supernatant by an acetone precipitation method. The ECP was incubated with normal rainbow trout serum at 20°C for 2 h, and the interrelationship between ECP proteolytic activity and immune complex-initiating, haemolytic complement activity (CH₅₀) of normal serum against antibody-sensitized goldfish red blood cells was evaluated. When normal serum was incubated with increasing concentrations of ECP, the CH₅₀ activity of serum decreased. The CH₅₀ activity was completely abolished in serum treated with undiluted ECP. ECP treated with serum was administered to trout intraperitoneally to determine mortality. All the fish receiving untreated ECP (0.05 ml = 0.5 mg protein) alone died within 24 h. When ECP was treated with serum at 1:1 to 4:1 (serum: ECP) in volume a similar high mortality was produced. These inocula possessed high protease activity and no or low CH₅₀ activity. However, mortality decreased and finally no mortality was recorded as ECP was treated with large volumes of serum (9:1 to 19:1). These inocula had lower protease activity and considerably higher CH₅₀ activity. Fish receiving ECP treated with heat-inactivated serum at 19:1 showed 100% mortality. A serum: ECP inoculum derived from fish which had been administered lipopolysaccharide from Salmonella enteritidis and which possessed a low CH₅₀ activity also gave a high mortality when used at 19:1. These results suggest that rainbow trout complement is implicated in the inactivation of toxicity of A. salmonicida ECP.     

Preliminary Comparisons of Yield and Profit Achieved from Different Rainbow Trout,Oncorhynchus mykiss,Production Systems in Inland Western Australia

Abstract

Farmers throughout the wheatbelt of Western Australia are interested in farming rainbow trout, Oncorhynchus mykiss, in saline groundwater on salt-affected farmland, to generate an alternative source of income. We compared the relative productivity and profitability of three different production systems: extensive (trout stocked in earthen ponds and totally reliant on natural food); semi-intensive (trout stocked in earthen ponds and provided with supplementary diet); intensive (trout stocked in closed, recirculating tanks). The yield of fish increased with increasing production intensity. The mean wet weight (±SE) of trout after 4 months of grow-out was 61.3±2.7 g in extensive systems, 157.9±5.2 g in semi-intensive systems and 137.9±3.9 g in intensive systems, giving mean yields of 10.8 kg/pond (13.5 kg/ha), 27.9 kg/pond (34.8 kg/ha) and 54.9 kg/tank (21.1 kg/m³), respectively. A preliminary economic analysis of the different production systems showed that the increases in yield were sufficient to balance the extra operating costs involved in semi-intensive systems, but not in intensive systems. We conclude that semi-intensive production systems deserve further study for the commercially viable production of rainbow trout from saline groundwater in Western Australia.     

Effects of skeleton photoperiods on melatonin secretion in the rainbow trout, Oncorhynchus mykiss (Walbaum)

Abstract Plasma melatonin levels were measured at hourly intervals in rainbow trout, Oncorhynchus mykiss (Walbaum), maitained under three different skeleton photoperiods: 8L:2D:2L:12D;8L:7D:2L:7D; and 8L:12D:2L:2D. Blood samples were taken before, during and after the 2-h light pulses. Melatonin levels increased rapidly to mean scotophase (dark period) values of 261 ± 7 pg/ml after the first light-dark transition and had returned to mean photophase (light period) values of 51 ± 2 pg/ml 30min after the end of the scotophase. Light pulses during the early, mid- and late scotophase elicited similar reductions in melatonin levels, followed by rapid increases after the light pulse, reaching previous scotophase values within 90 min. The inability of any of the light pulses to modulate or truncate melatonin secretion under a LD 8:16 photoperiod provides further support for the hypothesis that melatonin secretion in the rainbow trout is a direct response to darkness, and is not under endogenous circadian control as in other vertebrates examined.     

Studies on cryopreservation of eggs from rainbow trout (Salmo gairdneri) and coho salmon (Oncorhynchus kisutch)

The effect of pre-freezing treatments as well as freezing of inseminated, not water-activated eggs from rainbow trout Salmo gairdneri, and coho salmon, Oncorhynchus kisutch, was investigated in relation to survival and further development.Effects above freezing temperatures included: the temperature at insemination, viability of inseminated and unactivated eggs after storage, suitability of an incubation medium and the tolerance of eggs to various levels of the cryoprotectant dimethylsulfoxide (DMSO). Freezing experiments included: investigating the action of DMSO (0, 1, 2 mole) and the tolerance of coho eggs to temperatures between ?4.6 to ?30°C. Insemination temperatures between 0.5°C and 9.8°C (coho eggs) as well as incubation in an artifical medium (1-0°C) for 80 min (rainbow trout eggs) and 170 min (coho eggs) did not influence subsequent fertility. Storage of inseminated and unactivated rainbow trout eggs for 135 min and beyond reduced egg fertility. DMSO at 2 and 4 mole was detrimental to coho eggs (1-0°C). One mole DMSO had no (coho) or reduced (rainbow trout) influence on egg fertility when it was added gradually.In the presence of 1 mole DMSO most eggs remained unfrozen (67–89%) when kept for 10 min in frozen artificial medium (?4.6%) and 27–32% subsequently reached the eyed stage (control = 100). Further cooling (0.3°C/min) to ?10°C was still tolerated (62% unfrozen, 22% eyed eggs) but not to ?20°C (6% unfrozen, no development) and ?30°C (no survival). Use of 2 mole DMSO did not improve the results.     

Characterization of differentially expressed genes in liver in response to the rearing temperature of rainbow trout Oncorhynchus mykiss and their heritable differences

To characterize thermal-responsive genes in fish, firstly, juvenile rainbow trout were reared in four different temperature conditions (average temperatures were 10, 14, 18, and 22 °C, respectively) and differentially expressed genes were identified. Gene expression in the liver was analyzed by the differential display method, followed by validation using real-time PCR. Subsequently, to examine whether the identified genes show heritable differences, the gene expression levels were compared among juveniles of three genetically distinct lines of rainbow trout (a strain and two closed colonies) by rearing at two different temperature conditions (average 14 and 22 °C). By rearing at 22 °C, growth retardation was observed compared with fish reared at 14 and 18 °C, and six genes were identified as differentially expressed genes in response to the rearing temperature in the gene expression analyses. With the increase in rearing temperature, gene expressions of a complement C1q and two ribosomal proteins were significantly up-regulated. On the other hand, three metabolic genes (betaine homocysteine methyltransferase, triosephosphate isomerase, and glucose-6-phosphatase) were down-regulated, indicating a metabolic depression due to high temperature. In the subsequent analyses, in response to the rearing temperature (14 and 22 °C), there was a trend that the complement C1q and glucose-6-phosphatase genes showed different expression patterns among the three rainbow trout lines, suggesting heritable differences in these genes. Our study provides information on thermal-responsive genes in fish, and we anticipate it will facilitate further investigation in the thermal biology of fish.     

新型锌制剂对虹鳟生长及免疫功能的影响

为研究纳米锌及纳米锌多糖复合体对虹鳟生长及免疫功能的影响,选取平均体质量(120.0±3.0) g的虹鳟300尾,随机分成5组,每组3个平行,每个平行20尾,按1μL/g进行尾部肌肉注射纳米锌及纳米锌多糖复合体溶液,含量分别为1000、3000 mg/kg,试验周期为10 d。试验结果显示,纳米锌及纳米锌多糖复合体可不同程度提高虹鳟特定生长率(P>0.05)。1000 mg/kg纳米锌多糖复合体组虹鳟血液NBT阳性细胞数量百分比在第3 d最高,达22.3%(P<0.05);3000 mg/kg纳米锌多糖复合体组白细胞吞噬能力在第6 d最强(P<0.05)。3000 mg/kg纳米锌组虹鳟血清杀菌能力第3 d最高(19.4%),而3000 mg/kg纳米锌多糖复合体组血清杀菌能力第6 d最高,达32.43%(P<0.05)。第3 d,1000 mg/kg纳米锌多糖复合体组过氧化氢酶活性、一氧化氮合成酶活性分别为26.39、43.38 U/mL (P<0.05);第6 d,超氧化物歧化酶、酸性磷酸酶活性最高,分别为29.91 U/mL、201.4 U/L(P<0.05),微量丙二醛浓度最低,为3.19 nmol/mL。试验结束后通过杀鲑气单胞菌攻毒感染,7 d内观察虹鳟累积死亡率,1000 mg/kg纳米锌多糖复合体组累积死亡率仅30%。由试验结果可知,纳米锌和纳米锌多糖复合体均能对虹鳟免疫功能产生促进作用,以1000 mg/kg纳米锌多糖复合体效果最佳。     

Distribution and induction of cytochrome P450 1A1 in the rainbow trout brain

Cytochrome P450 (CYP) 1A1 participates in the activation as well as detoxification of environmental pollutants such as aromatic hydrocarbons. This CYP form is also efficiently induced by aromatic hydrocarbons. The presence of CYP 1A1 in the brain might thus be of physiological and toxicological importance. In the present investigation on rainbow trout, the distribution of 7-ethoxyresorufin-O-deethylase (EROD) activity, a cytochrome CYP 1A1 catalyzed reaction, was measured in whole tissue homogenates from brain parts. In control fish, a relatively high activity was found in the rainbow trout olfactory bulb compared to the other brain parts. Although an EROD induction (3 to 7-fold) by β-naphthoflavone (BNF) was recorded in all brain parts from the rainbow trout, the highest induced activity was measured in the olfactory bulbs. To ascertain the distribution of EROD activity in cells, whole brain tissue was subfractionated by differential centrifugation. The fractionation scheme separated mitochondria (P2 fraction) and microsomes (P3 fraction) as determined by marker enzymes and electron microscopy. In control rainbow trout, a low EROD activity could be measured in the P2 fraction. BNF induced the EROD activity in both P2 and P3 fractions. Western blotting showed the induction by BNF of a protein band in the P2 and P3 fractions with a molecular mass around 58,000 when highly specific anti-cod CYP 1A1 antibodies were used. ELISA measurements confirmed the induction of CYP 1A1 protein in the rainbow trout brain subcellular fractions.     

Hsp60/10 and sHsp families of heat shock protein genes in rainbow trout (Oncorhynchus mykiss) and their expression under heat stress

Heat shock proteins (Hsps) are highly conserved proteins whose expression can be induced by high temperature and play an important role in a variety of biological processes. However, systematic identification of the Hsp60/10 and small Hsp (sHsp) gene family in rainbow trout has not yet been reported, and there is little available information about its roles in evolution in rainbow trout, a typical economical cold-water fish. In this study, we performed a comprehensive analysis of the rainbow trout Hsp60/10 and sHsp gene family and to investigate their expression profiles. A total of one Hsp60 gene, one Hsp10 gene, and ten sHsp genes were identified. According to RNA-seq analysis of rainbow trout liver and head kidney under heat stress, a total of six out of ten sHsp genes were significantly upregulated in liver and head kidney. Real-time RT-PCR (RT-qPCR) was used to quantitatively analyze the expression levels of these genes in different tissues of rainbow trout. Results showed that the expression of hspe1 and hspd1 was lowest in liver and gill, respectively, and highest in brain. In sHsp gene family, all genes are highly expressed in the liver and head kidney, but relatively low in the heart, spleen, brain, gills, and muscles. This systematic analysis provided valuable information about the diverse roles of Hsp60/10 and sHsp in the evolution of teleost fish, which will contribute to the functional characterization of Hsp60/10 and sHsp genes in further research.

     

三倍体雌性虹鳟性腺发育阶段细胞色素相关基因的表达

Cyp19a1b、Cyp11a1、Cyp11b2、Cyp17a1、Hsd3b1等细胞色素相关基因能够调节硬骨鱼类性类固醇的合成,对性腺发育和性别决定产生影响。本研究以全雌三倍体虹鳟(Oncorhynchus mykiss)为研究对象,正常雌性二倍体虹鳟为对照,选取31~68 dpf(days post fertilization)时间段的虹鳟仔鱼脑组织,采用q RT-PCR和酶联免疫的方法研究以上几种基因的表达状况和脑芳香化酶的活性变化,以期探明导致三倍体雌性虹鳟性腺发育异常的关键原因。q RTPCR结果显示,二倍体中Cyp19a1b在30~50 dpf时表达量上调并且维持在较稳定水平,但50~56 dpf时表达量逐渐下调,之后56~68 dpf表达量持续上调;三倍体中Cyp19a1b表达量在30~35 dpf开始上调,35~47 dpf逐渐下调,47~55 dpf开始第二次上调,之后维持在较稳定水平直至68 dpf,但三倍体Cyp19a1b的表达量显著(P0.05)低于同期二倍体的。二倍体Cyp11a1表达量在34 dpf出现峰值,三倍体Cyp11a1在38 dpf时出现峰值。二倍体Hsd3b1表达量在33~42 dpf时维持在较高水平,在38 dpf时出现高峰;三倍体Hsd3b1表达量在47~59 dpf时较高,在49 dpf出现高峰。二倍体中Cyp11b2在37 dpf出现峰值,之后开始下调;三倍体在40 dpf出现峰值,之后逐渐下调,但三倍体Cyp11b2表达量显著低于同期二倍体。二倍体Cyp17a1的表达量在35~46 dpf时逐渐上升,在45 dpf时达到高峰之后直至69 dpf逐渐下降,并且维持在较为平稳的水平上;但是在相同的实验条件下未检测到同一时期三倍体Cyp17a1的表达量。酶联免疫结果显示,在40 dpf时二者的脑芳香化酶活性到达高峰,但在40~60 dpf时期,二倍体虹鳟脑芳香化酶活性显著(P0.05)高于三倍体虹鳟,尤其在45~50 dpf时,该酶活性分别较三倍体的高1.15倍和1.12倍。以上结果表明三倍体虹鳟早期性腺发育迟缓的原因之一是Cyp19a1b、Cyp11a1、Cyp11b2、Cyp17a1、Hsd3b1等基因的表达晚于二倍体,且表达量低于二倍体,造成雌二醇不能正常合成,最终导致性腺发育迟缓。