Cryopreservation of turbot Scophthalmus maximus (L.) sperm: fertilization and hatching rates

The present paper assesses the fertilization and hatching rates of an artificial fertilization series (n=1153) using fresh and cryopreserved sperm from 49 specimens of turbot Scophthalmus maximus (L.), carried out to confirm the results of a previous study, with the ultimate aim of transferring these cryo‐preservation techniques to commercial hatcheries. No significant differences were found between the fertilization rates of the two groups (fresh and cryopreserved sperm) when their respective fertility rates were >69.2%, which was the case in 75% of all fertilizations. Likewise, no significant differences in hatching rates were found. In order to use the cryopreserved sperm more efficiently, a key concern for commercial use of this technique, we also experimented with a lower sperm:diluent ratio (1:1) than used previously at our centre. We also compared the traditional 0.5‐mL straws with 2‐mL cryotubes able to contain a higher volume of sperm, finding no significant differences in the resulting fertilization and hatching rates. In conclusion, the use of cryopreservation for turbot sperm presents major advantages for broodstock management in commercial hatcheries.     

Handling time and predation behaviour by the crab, Cancer pagurus, preying on cultured scallop, Pecten maximus

We investigated handling time, persistence time and shell‐breaking techniques by crab, Cancer pagurus (L.) (13–15 cm carapace width), offered cultured scallops, Pecten maximus(L.), within the recommended release size for bottom culture. Three shell height groups were used: 50–55, 60–65 and 70–75 mm. The results showed that the crabs managed to open scallops from all the three size groups. The median handling time in the 50–55 mm group (788 s) was significantly different from the median handling time of the 60–65 mm (2482 s) and 70–75 mm (2980 s) groups. The median persistence time increased significantly with each scallop size, from 89 s in the 50–55 mm group to 97 s in the 60–65 mm and 125 s in the 70–75 mm group. We observed a change in the shell‐breaking techniques from a dominance of smashed scallops in the 50–55 mm group to more punched and chipped scallops in the 60–65 and 70–75 mm groups. The shift in predation behaviour when crabs were offered 50–55 mm scallops compared with the larger groups is discussed in relation to strategies in the release of scallops to seabed cultures.     

Comparative composition and shelf-life of fillets of wild and cultured turbot (Scophthalmus maximus) and Atlantic halibut (Hippoglossus hippoglossus)

Turbot and Atlantic halibut are highly valued fish species. However,very little is known about fillet shelf-life characteristics associated withboth species. Thus, fillet -tocopherol content and proximate compositionof wild turbot (1.5 kg) and Atlantic halibut (1.1 kg)caught off the south coast of Ireland and the north-west coast of Iceland,respectively, were investigated. In addition, the susceptibility of fillets, storedunder retail conditions, to lipid oxidation and colour change was studied.Proximate composition analysis showed that turbot had significantly highermoisture (P < 0.001) and lower protein (P < 0.001) contents compared toAtlantic halibut. Atlantic halibut incorporated significantly higher (P <0.001) levels of -tocopherol into fillets than turbot. Over 14 days ofstorage on ice, fillets from Atlantic halibut exhibited significantly lower (P =0.020) levels of lipid oxidation than those of turbot. However, malondialdehyde(MDA) concentrations were generally very low, never exceeding 0.6 gg^–1 fillet. Turbot maintained a significantly higher (P< 0.001) pH over the storage period. The lightness (L^* values) offillets from both species increased over 14 days of storage, but wassignificantly higher (P < 0.001) in Atlantic halibut than in turbot. Turbotdeveloped a relatively intense yellow colour during storage (decrease in hueangle and increase in b^* values), whereas this was not the case forAtlantic halibut. The results of this study demonstrate that fillets of wildAtlantic halibut stored on ice, were less prone to lipid oxidation anddiscolouration than those of wild turbot. However, quality changes in turbotwere very small showing that both fish have tremendous shelf-life capacities interms of lipid oxidation. These findings are considered in the context of knownmaterial for farmed fish.     

大菱鲆胚胎玻璃化方法研究

用大菱鲆(Scophthalmus maximus)神经期胚对6种抗冻剂的毒性进行检测,发现其毒性排列为1,2-丙二醇(PG)<甲醇(MeOH)<甘油(Gly)<二甲基甲酰胺(DMF)<乙二醇(EG)<二甲亚砜(DMSO).以DMF为主因子配制和筛选出11种玻璃化程度较好的玻璃化液,并用大菱鲆肌节期胚对其中5种玻璃化程度最好的玻璃化液进行检测,结果显示,A4(DMSO 20%+DMF 25%)、A7(DMSO 25%+DMF 20%)较适合于大菱鲆胚胎的平衡处理,利用A4和A7平衡处理神经期胚、肌节期胚、心跳期胚、出膜前期胚,结果显示大菱鲆神经胚和肌节胚对玻璃化液的适应能力较强.实验测试出不同时期胚胎在两种玻璃化液中的成活率,为大菱鲆各期胚胎在玻璃化液中的平衡处理提供了依据.利用大菱鲆肌节胚对不同的平衡步骤进行了筛选,发现五步法的平衡效果较好.     

大菱鲆正常代谢水平及节律性的研究

对工厂化养殖的不同规格大菱鲆(Scophamus maximus Linnaeus)的日常代谢进行了研究。结果表明,4种规格的大菱鲆表现出相同的代谢规律性,摄食对大菱鲆的代谢有明显的影响,摄食后代谢率呈迅速上升的趋势。在摄食后3～9 h内代谢率出现第1高峰值,在凌晨3:00～6:00(摄食后21-24 h)出现第2高峰值,代谢率昼、夜之间的差异不显著(P>0.05)。体重对大菱鲆的代谢有着显著的影响,4种规格9.69±3.35g、33.34±8.34g、130.18±26.75g和264.11±35.79g的大菱鲆日平均耗氧率分别为0.2702±0.0225、0.1880±0.0189、0.1592±0.0436、0.1033±0.0184mgO2/g·h,日平均氨氮排泄率分别是0.6496±0.0303、0.4560±0.0669、0.3744±0.0320、0.2831±0.0662mgN/100g·h,随着体重的增加,单位体重的代谢率呈下降的趋势(P<0.05)。     

大菱鲆早期变态发育和体表黑色素细胞形态学观察

对大菱鲆早期变态发育及体表黑色素细胞的形成过程进行了研究。绘制了1－60日龄大菱鲆鱼苗形态发育图，同时观察拍摄不同发育时期体表黑色素细胞的形态变化。14－17℃水温时，大菱鲆早期变态发育约需60d。1－14日龄主要为器官发育；14－40日龄主要为形态变化，25日龄左右开始变态；40－60日龄变态基本完成。变态前，鱼苗体表的幼体黑色素细胞对称分布，以素细胞密度先增加后减少。变态开始后，鱼体两侧体色出现不对称。有眼侧体表幼体黑色素细胞减少，逐渐由成体黑色素细胞替代，体色变深；无眼侧幼体黑色素细胞逐渐消褪，未出现成体黑色素细胞，体色逐渐变为白色。若有眼侧体表成体黑色素细胞的出现受阻，则形成白化。     

The effect of dietary vitamin E and C level on market-size turbot (Scophthalmus maximus) fillet quality

Fish fillet quality has been shown to be influenced by the level of antioxidants in preslaughter diet. Thus, an experiment was conducted to study the effect of different levels of vitamin E and C on the fillet quality of market‐size reared turbot (Scophthalmus maximus). Turbot of a mean initial weight of 347 ± 20 g were divided into four groups and fed commercial turbot diets (60% protein, 12% fat), supplemented with α‐tocopheryl acetate (mg kg^?1) and ascorbyl‐2 monophosphate (mg kg^?1) at the following dietary levels: 500/100, 1000/100, 100/1000, 100/100 respectively. Over a dietary supplementation period of 15 weeks, fish were fed to satiation and reached a final mean weight of 916 ± 29 g. α‐Tocopherol levels increased significantly (P < 0.001) in tissue (i.e. muscle, liver, heart and kidney) of fish fed diets containing elevated levels of α‐tocopheryl acetate. In ice storage, fillets of these fish exhibited significantly lower (P < 0.001) levels of lipid oxidation, and showed significantly less (P < 0.001) colour deterioration (higher hue angle and lower chroma). Elevated dietary α‐tocopheryl acetate levels had a negative effect (P ≤ 0.001) on the concentration of ascorbic acid in muscle tissue. An increase in dietary vitamin C did not have any detectable effect on fillet quality. Prolonged feeding times had a negative effect on lipid oxidation (P < 0.001) and colour deterioration (P < 0.01). These results suggest that increased dietary α‐tocopheryl acetate could prevent colour deterioration and lipid oxidation of turbot fillets in retail storage on ice.     

中国大菱鲆虹彩病毒主要衣壳蛋白基因的PCR扩增及序列分析

应用同源PCR技术，从被一种球状病毒感染的患病大菱鲆（Scophthalmus maximus）脾脏和肾脏组织中扩增出了一段长度为620bp的DNA片断。序列测定和Blast分析表明，该DNA片断与鱼类虹彩病毒主要衣壳蛋白（MCP）C末端编码区的DNA序列高度相似，由此证实感染养殖大菱鲆的这种球状病毒为一种鱼类虹彩病毒，暂命名为大菱鲆红体病虹彩病毒（TRBIV）。多序列比对和分析发现，TRBIV MCP C末端的205个氨基酸序列与GenBank中20种虹彩病毒相应序列的相似性分别为99．47％（韩国大菱鲆虹彩病毒）、97％～98％（待指定病毒属的7种病毒），以及50％以下（蛙病毒属、淋巴囊肿病毒属、虹彩病毒属的12种病毒），由此绘制出了包含TRBIV在内的21种虹彩病毒的系统发育树。研究结果表明，感染中国养殖大菱鲆的TRBIV属于虹彩病毒科待指定病毒属，位于该属ISKNV亚群和RSIV亚群之间，是该病毒属的一个新成员。     

Characterization of nodavirus and viral encephalopathy and retinopathy in farmed turbot, Scophthalmus maximus (L.)

An outbreak of nodavirus infection in turbot larvae is described with respect to histopathology, immunohistochemistry, cell culture cultivation, RT-PCR amplification and sequence analysis of the capsid protein gene RNA2. Affected turbot developed classical signs of viral encephalopathy and retinopathy (VER) with abnormal swimming behaviour and high mortality levels. In the acute stage of infection, light microscopy revealed vacuolation of the central nervous system (CNS), with positive immunohistochemical staining for nodavirus. Later in the infection, CNS lesions appeared more chronic and contained clusters of cells immunopositive for nodavirus. Bacterial overgrowth in the intestines of the fish may have provoked or influenced the course of the nodavirus infection. We were unable to propagate the virus in cell culture. While RT-PCR using primers designed to detect Atlantic halibut nodavirus gave negative results, further testing with primers complementary to a more conserved region of RNA2 resulted in amplification of a product of the expected size. The entire RNA2 segment was cloned and sequenced. Sequence alignment showed that the turbot nodavirus (TNV) was different from previously described fish nodaviruses. In addition, phylogenetic analysis based on an 823 nt region of the sequence indicated that TNV clustered outside the four established fish nodavirus genotypes, suggesting a fifth genotype within the betanodaviruses.     

Stress responses of larval turbot, Scophthalmus maximus L., exposed to sub-lethal concentrations of petroleum hydrocarbons

Turbot larvae were exposed to dilutions of the water soluble fraction (WSF) of crude oil (1–25%), in a laboratory flow-through system. Whole body immunoreactive cortisol (IRC) content of premetamorphic 345 degree C day and early metamorphosing 450 degree C day larvae (23 and 30 days post hatch, respectively) was elevated when exposed to 25% WSF for 6 h, but the lower WSF concentrations did not induce IRC elevations. Larvae of 450 degree C days exposed to WSF for 6 h and then left in clean sea water for 24 h showed recovery of IRC, whilst the IRC content of 450 degree C day larvae exposed to WSF for 30 h remained elevated. Whole body thyroxine content of 345 degree C day larvae exposed to 25% WSF for 6 h was significantly elevated, which may have implications for larval development/metamorphosis. However, whole body triiodothyronine content of these larvae was not altered by WSF-exposure. Furthermore, larvae of 450 degree C days, WSF-exposed for 6 h or 30 h showed no thyroidal disturbance. Larvae of 345 degree C days exhibited significantly elevated whole body adrenaline and noradrenaline content after exposure for 6 h to 25% WSF; however, more diluted WSF did not influence adrenaline or noradrenaline content. These studies have demonstrated that larvae of 345 and 450 degree C days can successfully mount endocrine stress responses but that environmentally realistic concentrations of petroleum hydrocarbons do not induce these responses.