大菱鲆白化相关基因(DEN-1、DEN-2)的分离鉴定及特征分析

应用mRNA差异显示技术,对比研究正常与白化大菱鲆有眼侧皮肤组织,克隆差异表达基因,经测序,RT-PCR验证,差异表达基因片段DEN-1(GenBank登录号:DQ886390)与DEN-2(GenBank登录号:DQ886391)均在白化大菱鲆有眼侧皮肤组织中下调表达;通过BLAST检索发现,DEN-1与GenBank中的斑马鱼和牛的类似尿苷二磷酸-葡萄糖:糖蛋白葡糖基转移酶(UGGT)基因、与挪威鼠类似尿苷二磷酸-葡萄糖:神经酰氨葡糖基转移酶1(Ugcgl1)基因有较高同源性;DEN-2与原鸡、斑马鱼、人、挪威鼠、家鼠和狗的眼缺乏同源框4(eya4)基因的同源性均较高。采用相对定量RT-PCR对正常鱼有眼侧皮肤(N1)、白化鱼有眼侧皮肤(A1)、正常鱼无眼侧皮肤(N2)、白化鱼无眼侧皮肤(A2)进行表达谱分析,以内参β-actin基因分别对DEN-1和DEN-2表达量进行标准化的光密度扫描显示:DEN-1和DEN-2在4种皮肤组织中有相似的表达模式,即表达量:N1>N2≈A2>A1。电子延伸将DEN-2延伸为497bp的片段,该延伸片段与多种动物的eya4基因在核酸和蛋白水平均有很高同源性。本研究对DEN-1和DEN-2的不同组织器官和不同发育阶段的电子表达谱分析,将对其进一步研究提供有价值参考。     

Fish may fight rather than feed in a novel environment: metabolic rate and feeding motivation in juvenile Atlantic salmon

This study tested the hypothesis that juvenile Atlantic salmon Salmo salar with a high resting metabolic rate and probability of dominance will also have a correspondingly higher feeding motivation in a novel environment to offset their greater costs of maintenance. The opposite was found to be the case: Atlantic salmon with a high standard metabolic rate had a slightly but significantly lower feeding motivation. It is hypothesized that Atlantic salmon with higher maintenance costs opt to be more aggressive at the expense of the costly activities associated with feeding, since elevated aggression in a new habitat is a more successful strategy for acquiring a feeding territory (and hence sustaining food intake in the long-term) than a high feeding motivation alone.     

Utilization of metabolic scope in relation to feeding and activity by individual and grouped zebrafish, Brachydanio rerio (Hamilton-Buchanan)

Metabolic scope and its utilization in relation to feeding and activity were measured in individual and grouped zebrafish (weight range, 430–551 mg) at 24° C by respirometry. Mean maximum metabolic rate, induced by swimming to exhaustion, R_max(i), was 1223 (s.d. , 157) mg O₂, kg^?1 h^?1 for individuals. Standard metabolic rate, R_s. was 364 mg O₂ kg^?1 h^?1, as estimated by extrapolating to zero activity from measurements of unfed, spontaneously active individuals. Mean routine metabolic rate, R_rout, of individuals was 421 (s.d. , 58) mg O₂, kg_-1 h_-1. The mean voluntary maximum metabolic rate, R_max(v), following transfer of minimally exercised fish to the respirometer, was 1110 (s.d. , 83) mg O₂ kg ^?1 h^?1 for groups of six fish, and was not significantly different from the value measured for individuals, 1066 (s.d. , 122) mg O₂, kg^?1 h^?1. Grouped fish acclimated to the respirometer more slowly than individual fish and exhibited significantly higher R_rout, apparently a result of greater social interaction and activity in groups. Mean R_rout for groups was 560 (s.d. , 78) mg O₂, kg^?1 h^?1. While groups of zebrafish fed a ration of 5% wet body weight day^?1 exhibited consistently higher metabolic rates than fish fed rations of 2.5% wet body weight day^?1 the high ration group still used only a maximum of 77% of the metabolic scope. Zebrafish of the size studied do not appear to demonstrate a high degree of conflict in utilization of metabolic scope by different respiratory components. The metabolic rates measured for zebrafish are among the highest yet measured for fish of similar size and at similar temperatures.     

溶氧水平对鲫鱼代谢模式的影响

为了探讨水体溶氧水平对鲫幼鱼(Carassius carassius)运动、消化能力及其交互作用的影响,在(25.0±0.5)℃温度条件下,测定了8(饱和溶氧水平)、2和1mg/L溶氧水平下摄食(饱足摄食)和空腹组(空腹2 d)鲫鱼的临界游泳速度(Ucrit)、运动前耗氧率(MO2pre-exercise)、活跃耗氧率(MO2active)和代谢范围(MS)。摄食诱导的耗氧率上升在各溶氧水平下无显著差异。在饱和溶氧水平下,摄食组和空腹组的Ucrit没有显著差异,但在1和2 mg/L条件下,摄食组的Ucrit显著低于空腹组(P<0.05)。在饱和溶氧水平条件下,消化和运动诱导的耗氧率上升在各个游泳水平均能完全叠加,且摄食组鱼类与空腹组鱼类具有相似的MS和Ucrit和更高的MO2active,提示鲫鱼在常氧下为添加代谢模式。随着溶氧水平下降至2和1mg/L,呼吸能力(摄食组的MO2active)对溶氧水平下降较运动耗氧率更为敏感,消化诱导的耗氧率增加只能在较低游泳速度叠加,与空腹组鱼类比较,摄食组鱼类的MS和Ucrit显著下降,MO2active无显著差异,提示低氧下消化和运动对氧气需求竞争的加剧使其代谢模式转化为消化优先。     

The interaction of temperature and fish size on growth of juvenile turbot

Growth rate of tagged juvenile turbot was significantly influenced by the interaction of temperature and fish size. The results suggest the optimum temperature for growth of juvenile turbot in the size range 25–75 g is between 16 and 19°C. Optimal temperature for growth decreased rapidly with increasing size, and is between 13 and 16°C for 100 g turbot. Although individual growth rates varied highly at all times within the temperature treatments, significant size rank correlations were maintained during the experimental period. The study confirms that turbot exhibit ontogenetic variation in temperature optimum, which might partly explain different spatial distribution of juvenile and adult turbot in ocean waters.     

Effects of growth hormone transgenesis on metabolic rate, exercise performance and hypoxia tolerance in tilapia hybrids

Swimming respirometry was employed to compare inactive metabolic rate ( R _r), maximum metabolic rate ( R _max), resultant aerobic scope and maximum sustainable (critical) swimming speed ( U _crit), in growth hormone transgenic (GHT) and wild-type (W) tilapia Oreochromis sp. hybrids. Although the R _r of GHT tilapia was significantly (58%) higher than their W conspecifics, there were no significant differences in their net aerobic scope because GHT tilapia exhibited a compensatory increase in R _max that was equal to their net increase in R _r. As a consequence, the two groups had the same U _crit. The GHT and W tilapia also exhibited the same capacity to regulate oxygen uptake during progressive hypoxia, despite the fact that the GHT fish were defending a higher demand for O₂. The results indicate that ectopic expression of GH raises metabolic rate in tilapia, but the fish compensate for this metabolic load and preserve such physiological determinants of fitness as aerobic scope, swimming performance and tolerance of hypoxia.     

摄食水平和投喂频率对大菱鲆幼鱼生长及生化成分的影响

以大菱鲆幼鱼(7.56±0.03 g)为试验对象,在水温17～19℃下,设计了4个摄食水平梯度(1.0％、1.5％、2.0％和饱食)和3个投喂频率梯度(1、2和3次·d-1),探讨了不同摄食水平和投喂频率对其生长及生化成分的影响.结果表明:随着摄食水平的升高,大菱鲆幼鱼的终末体重、相对增重率和摄食率显著升高(P＜0.05);特定生长率随摄食水平的增加呈线性升高;湿重、干重、蛋白质和能量转化效率出现先升高后降低的趋势;摄食水平对鱼体的体成分和组间表观消化率无显著影响(P＞0.05);不同投喂频率下,终末体重、相对增重率、特定生长率及转化效率呈先升高再降低的趋势,在2次·d-1时取得最大值;投喂频率对鱼体的体成分和组间表观消化率无显著影响(P＞O.05).     

Effect of temperature on the drinking rates of two species of flatfish, flounder and turbot

Flounders responded to a temperature change of 10° C by increasing drinking rate with a Q ₁₀ value of about 1.5 while turbot showed a limited capacity to respond to a similar temperature change. Drinking as an indicator of osmoregulatory performance is well developed in the euryhaline flounder compared to the stenohaline turbot indicating fundamental differences in the physiological response to temperature.     

Effects of temperature on metabolic rate and lower dissolved oxygen tolerance of juvenile speckled peacock bass Cichla temensis

The speckled peacock bass Cichla temensis is a popular sport and food fish that generates substantial angling tourism and utilitarian harvest within its range. Its popularity and value make this species important for management and a potential aquaculture candidate for both fisheries enhancement and food fish production. However, little is known of optimal physiochemical conditions in natural habitats, which also are important for the development of hatchery protocols for handling, spawning and grow-out. Speckled peacock bass have been documented to have high sensitivity to extreme temperatures, but the metabolic underpinnings have not been evaluated. In this study, the effects of temperature (25, 30 and 35°C) on the standard metabolic rate (SMR) and lower dissolved oxygen tolerance (LDOT) of juvenile speckled peacock bass (mean ± standard error total length 153 ± 2 mm and wet weight 39.09 ± 1.37 g) were evaluated using intermittent respirometers after an acclimation period of 2 weeks. Speckled peacock bass had the highest SMR at 35°C (345.56 ± 19.89 mgO₂ kg⁻¹ h⁻¹), followed by 30°C (208.16 ± 12.45 mgO₂ kg⁻¹ h⁻¹) and 25°C (144.09 ± 10.43 mgO₂ kg⁻¹ h⁻¹). Correspondingly, the Q₁₀, or rate of increase in aerobic metabolic rate (MO₂) relative to 10°C, for 30–35°C was also greater (2.76) than from 25 to 30°C (2.08). Similarly, speckled peacock bass were the most sensitive to hypoxia at the warmest temperature, with an LDOT at pO₂ of 90 mmHg (4.13 mg l⁻¹) at 35°C compared to pO₂ values of 45 mmHg (2.22 mg l⁻¹) and 30 mmHg (1.61 mg l⁻¹) at 30 and 25°C, respectively. These results indicate that speckled peacock bass are sensitive to temperatures near 35°C, therefore we recommend managing and rearing this species at 25–30°C.     

The effect of dissolved oxygen tension and the utility of oxygen uptake rate in insect cell culture

Dissolved oxygen tension and oxygen uptake rate are critical parameters in animal cell culture. However, only scarce information of such variables is available for insect cell culture. In this work, the effect of dissolved oxygen tension (DOT) and the utility of on-line oxygen uptake rate (OUR) measurements in monitoring Spodoptera frugiperda (Sf9) cultures were determined. Sf9 cells were grown at constant dissolved oxygen tensions in the range of 0 to 30%. Sf9 metabolism was affected only at DOT below 10%, as no significant differences on specific growth rate, cell concentration, amino acid consumption/production nor carbohydrates consumption rates were found at DOT between 10 and 30%. The specific growth rate and specific oxygen uptake rate followed typical Monod kinetics with respect to DOT. The calculated _max and max were 0.033 h^-1 and 3.82×10^-10 mole cell^-1h^-1, respectively, and the corresponding saturation constants were 1.91 and 1.57%, respectively. In all aerated cultures, lactate was consumed only after glucose and fructose had been exhausted. The yield of lactate increased with decreasing DOT. It is proposed, that an apparent DOT in non-instrumented cultures can be inferred from the lactate yield of bioreactors as a function of DOT. Such a concept, can be a useful and important tool for determining the average dissolved oxygen tension in non-instrumented cultures. It was shown that the dynamic behavior of OUR can be correlated with monosaccharide (fructose and glucose) depletion and viable cell concentration. Accordingly, OUR can have two important applications in insect cell culture: for on-line estimation of viable cells, and as a possible feed-back control variable in automatic strategies of nutrient addition.Abbreviations DOT Dissolved oxygen tension - OUR Oxygen uptake rate - specific oxygen uptake rate - specific growth rate - X_v viable cell concentration - C_L, C^*, and oxygen concentrations in liquid phase, in equilibrium with gas phase, and medium molar concentration, respectively - H Henry's constant - K_La volumetric oxygen transfer coefficient - P_T total pressure - oxygen partial pressure - oxygen molar fraction - i discrete element     

The ageing phenomenon of turbot spermatozoa: effects on morphology, motility and concentration, intracellular ATP content, fertilization, and storage capacities

The deleterious effect of the ageing phenomenon of turbot spermatozoa was investigated in relation to the sampling date. Spermatozoa with a low or highly condensed chromatin and a middle piece containing numerous or a few vesicles were observed simultaneously 80 and 47 days before the beginning of the spawning period of the females. The middle piece of spermatozoa contained few vesicles, 39 days after the end of the reproductive period. At the same date, some spermatozoa appeared in which the plasma membrane was broken. Sperm motility, assessed just after collection in terms of arbitrary motility scores from 0 to 5, was significantly increased both at 10 and 60 s post-activation, for samples collected 18 days after, 25 days before and 9 days after the beginning of the spawning period of the females, respectively compared to samples collected 6 days before, 55 days after and 88 days after the end of this period. A lower short-term storage capacity was recorded at 10 and 60 s post-activation for sperm samples collected 6 days before and 88 days after the end of the reproductive period, respectively compared to 18 days and 9 days after the beginning of the spawning period. At 60 s post-activation, a higher motility of thawed spermatozoa was observed for samples collected 5 days before the beginning of the spawning period (motility recovery index: 86.4 ± 19.4%) compared to 71 days after the end of this period (55.0 ± 12.0%). The fertilizing capacity of sperm samples collected 61 days after the end of the spawning season (66.1 ± 14.6%) was significantly lower than that recorded for samples collected 34 days after the beginning of the spawning period (75.2±9.6%). On the contrary, there was no significant decrease in endogenous ATP content (31 days after the beginning of the spawning period, 14.53 ± 0.84; 48 days after the end of this period, 10.75 ± 5.26 nmol 10^{? 8} spermatozoa). Furthermore, sperm concentration significantly increased between the same dates (respectively 3.3 ± 0.8–9.4±4.8×10⁹ spermatozoa ml^?1).     

Effects of salinity on the ionic balance and growth of juvenile turbot

The effects of salinity changes (27, 19 and 10‰) on seawater-adapted juvenile turbot were studied on their plasma osmolarity and ion concentrations, on oxygen consumption, on gill Na⁺,K⁺-ATPase activity after 3 months and on growth parameters. All plasma concentrations (except chloride) were unchanged, suggesting that fish were well adapted to their environment. Oxygen consumption was significantly decreased in the 19 and 10‰ groups, where fish weighed significantly more 105 days after transfer than fish maintained in sea water. These results, and the fact that apparent food conversion rates were lower in a diluted environment, suggest that on a long term schedule growth conditions could be improved by adaptation to brackish waters (salinities between 10 and 19‰). The effects of transfer from sea water to 27, 19, 10 and 5‰ were also followed during the first 3 weeks. With salinity 10‰ a steady state was reached on day 21 with all plasma values within the same range. The significant differences observed in osmolarity, plasma ion concentrations and Na⁺,K⁺-ATPase activity 3 weeks after transfer of juveniles to 5‰ salinity, compared with transfers in higher salinities, suggest that there is a threshold of acclimation of turbot to a hypotonic environment.     

The combined effect of body size and temperature on oxygen consumption rates and the size-dependency of preferred temperature in European perch Perca fluviatilis

The present study determined the effect of body mass and acclimation temperature (15–28°C) on oxygen consumption rate (ṀO₂) and the size dependency of preferred temperature in European perch Perca fluviatilis. Standard metabolic rate (SMR) scaled allometrically with body mass by an exponent of 0.86, and temperature influenced SMR with a Q₁₀ of 1.9 regardless of size. Maximum metabolic rate (MMR) and aerobic scope (MMR-SMR) scaled allometrically with body mass by exponents of 0.75–0.88. The mass scaling exponents of MMR and aerobic scope changed with temperature and were lowest at the highest temperature. Consequently, the optimal temperature for aerobic scope decreased with increasing body mass. Notably, fish <40 g did not show a decrease aerobic scope with increasing temperature. Factorial aerobic scope (MMR × SMR⁻¹) generally decreased with increasing temperatures, was unaffected by size at the lower temperatures, and scaled negatively with body mass at the highest temperature. Similar to the optimal temperature for aerobic scope, preferred temperature declined with increasing body mass, unaffectedly by acclimation temperature. The present study indicates a limitation in the capacity for oxygen uptake in larger fish at high temperatures. A constraint in oxygen uptake at high temperature may restrict the growth of larger fish with environmental warming, at least if food availability is not limited. Furthermore, behavioural thermoregulation may be contributing to regional changes in the size distribution of fish in the wild caused by global warming as larger individuals will prefer colder water at higher latitudes and at larger depths than smaller conspecifics with increasing environmental temperatures.     

Relation between dissolved oxygen concentration and ajmalicine production rate in high-density cultures of Catharanthus roseus

The relation between dissolved oxygen (DO) and the ajmalicine production rate of Catharanthus roseus was investigated in 15-L tank reactors at constant stirrer speed and gas flow rate. Below a DO concentration of 29% of air saturation the ajmalicine production rate was less than 0.06 mumol/g/d. Above a DO of 43% the ajmalicine production rate was constant at 0.21 mumol/g/d. Between a DO of 29% and 43% there was a strong relation between the ajmalicine production rate and the DO concentration. After a period of at least 12 days at DO /=57%. A kinetic equation is proposed for the relation between DO and the specific ajmalicine production rate. (c) 1995 John Wiley & Sons, Inc.     

The effects of body mass and feeding on metabolic rate in small juvenile Atlantic cod

Apparent specific dynamic action (SDA) amplitude in young juvenile Atlantic cod Gadus morhua (1 to 8 g wet mass), fed a standardized meal and then exercised in a circular swimming respirometer at a constant swimming speed of 0·5 ± 0·3 body lengths s^-1, occurred within l h after feeding in all juveniles. SDA amplitude were 1·4 to 1·8 times higher in fed fish compared to unfed fish, and rates of oxygen consumption decreased as body mass increased. SDA duration had a tendency to decrease with increasing body mass and was shortest, at 6 h, in the smallest fish (1–1·5 g), but increased to 10–11 h in the largest fish. Apparent SDA in fed fish ( R _r) scaled with a mass exponent of 0·89, while maximum metabolic rate ( R _max) determined by chasing fish to exhaustion and then measuring oxygen consumption for 12 h, and unfed routine metabolic rate (R_r) scaled with a mass exponent of 0·79 and 0·76 respectively. Relative aerobic scope ( R _max– unfed R _r) did not appear to vary over the 1 to 8 g increase in wet mass. These results show that as body mass increased in young juvenile Atlantic cod: (1) apparent SDA ( R _f) increased more rapidly than R _max, and (2) apparent SDA took up >98% of the relative aerobic scope and that young Atlantic cod allocated most of the energy to growth, and left little for other metabolic activities.     

Valid oxygen uptake measurements: using high r2 values with good intentions can bias upward the determination of standard metabolic rate

This analysis shows good intentions in the selection of valid and precise oxygen uptake (O₂) measurements by retaining only slopes of declining dissolved oxygen level in a respirometer that have very high values of the coefficient of determination, r², are not always successful at excluding nonlinear slopes. Much worse, by potentially removing linear slopes that have low r² only because of a low signal-to-noise ratio, this procedure can overestimate the calculation of standard metabolic rate (SMR) of the fish. To remedy this possibility, a few simple diagnostic tools are demonstrated to assess the appropriateness of a given minimum acceptable r², such as calculating the proportion of rejected O₂ determinations, producing a histogram of the r² values and a plot of r² as a function of O₂. The authors offer solutions for cases when many linear slopes have low r². The least satisfactory but easiest to implement is lowering the minimum acceptable r². More satisfactory solutions involve processing (smoothing) the raw signal of dissolved oxygen as a function of time to improve the signal-to-noise ratio and the r²s.     

Effect of different cryoprotectants and vitrificant solutions on the hatching rate of turbot embryos (Scophthalmus maximus)

Vitrification could provide a promising tool for the cryopreservation of fish embryos. However, in order to achieve a vitrifiable medium, a high concentration of permeable cryoprotectants must be employed, and the incorporation of high molecular weight compounds should also be considered. The toxicity of these permeable and non-permeable agents has to be assessed, particularly when high concentrations are required. In the present study, permeable and non-permeable cryoprotectant toxicity was determined in turbot embryos at two development stages (F stage-tail bud and G stage-tail bud free). Embryos treated with pronase (2mg/ml, 10 min at 22 degrees C) were incubated in dimethyl sulfoxide (Me2SO), methanol (Meth.) or ethylene glycol (EG) in concentrations ranging from 0.5 to 6M for periods of 10 or 30 min, and in 5, 10, and 15% polyvinylpyrrolidone (PVP), 10, 15, and 20% sucrose or 0.1, 1, and 2% X-1000 for 2 min. The embryos were then washed well and incubated in seawater until hatching. The toxicity of permeable cryoprotectants increased with concentration and exposure time. There were no significant differences between permeable cryoprotectants. However, embryos tolerated higher concentrations of Me2SO than other cryoprotectants. Exposure to permeable cryoprotectants did not affect the hatching rate except at G stage with X-1000 treatment and 20% sucrose. Taking into account the cryoprotectant toxicity and the vitrification ability of cryoprotectant mixtures, three vitrification solutions (V1, V2, and V3), and one protocol for stepwise incorporation were designed. The tested solutions contained 5M Me2SO+2M Meth+1M EG plus 5% PVP, 10% sucrose or 2% X-1000. The hatching rate of embryos that had been exposed to the the vitrification solutions was analyzed and no significant differences were noticed compared with the controls. Our results demonstrate that turbot embryos can be subject to this cryoprotectant protocol without deleterious effect on the hatching rate.     

The influence of feeding regime on sexual maturation, fecundity and atresia in first-time spawning turbot

Exposure of adolescent turbot Scophthalmus maximus to low rations during vitellogenesis, covering the 4 months immediately prior to spawning, led to a drop of 70% in mean ovary weight, and was associated with poor growth of the vitellogenic oocytes or, in a third of cases, the absence of vitellogenic oocytes. Exposure to low rations during the recruitment of vitellogenic oocytes, 4–8 months prior to spawning, produced a more variable response. A few of these females produced well-developed vitellogenic oocytes, but in most, vitellogenic oocytes were absent or only poorly developed and in some cases there was a high incidence of atresia. Exposure to intermediate rations throughout oocyte recritment and vitellogenesis also depressed oocyte development. In maturing females the number of non-atretic vitellogenic oocytes (relative potential fecundity) averaged 998 per g of somatic body weight of fish at the start of the spawning season in mid-June. Maturation of the males was not significantly influenced by the dietary regime. The small size of the testes and the low levels of milt production, even in fish fed high rations immediately prior to spawning, showed that reproductive investment in males was much lower than in females.     

Effects of dilution on dissolved oxygen depletion and microbial populations in the biochemical oxygen demand determination

The biochemical oxygen demand (BOD) value is still a key parameter that can determine the level of organics, particularly the content of biodegradable organics in water. In this work, the effects of sample dilution, which should be done inevitably to get appropriate dissolved oxygen (DO) depletion, on the measurement of 5-day BOD (BOD₅), was investigated with and without seeding using natural and synthetic water. The dilution effects were also evaluated for water samples taken in different seasons such as summer and winter because water temperature can cause a change in the types of microbial species, thus leading to different oxygen depletion profiles during BOD testing. The predation phenomenon between microbial cells was found to be dependent on the inorganic nutrients and carbon sources, showing a change in cell populations according to cell size after 5-day incubation. The dilution of water samples for BOD determination was linked to changes in the environment for microbial growth such as nutrition. The predation phenomenon between microbial cells was more important with less dilution. BOD₅ increased with the specific amount of inorganic nutrient per microbial mass when the natural water was diluted. When seeding was done for synthetic water samples, the seed volume also affected BOD due to the rate of organic uptake by microbes. BOD₅ increased with the specific bacterial population per organic source supplied at the beginning of BOD measurement. For more accurate BOD measurements, specific guidelines on dilution should be established.     

The relationship between otolith accretion and resting metabolic rate in juvenile Atlantic salmon during a change in temperature

This study confirmed earlier evidence for a correlation between otolith increment width and resting metabolic rate in the absence of somatic growth in Atlantic salmon parr. However, increment width responded more conservatively to a change in temperature than did resting metabolic rate. Consequently, the study demonstrated that although increment width varied with temperature the process governing accretion rate was not simply related to resting metabolic rate.