Purification and biochemical characterization of a cellulase from the digestive organs of the short-spined sea urchin Strongylocentrotus intermedius

We isolated a cellulase from the digestive organs of the short-spined sea urchin Strogylocentrotus intermedius using a combination of ion-exchange chromatography and gel filtration together with an assay for carboxymethylcellulase activity. The isolated cellulase was stained as a single band by Congo red. The molecular weight of the isolated cellulase, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions, was 59?kDa. The isolated cellulase exhibited hydrolytic activity toward carboxymethyl cellulose, with an optimum temperature and pH of 30?°C and pH 8.0, respectively. The thermal stability of the enzyme was characterized by determining the temperature at which activity decreased by 50?% with treatment for 30?min at pH 7.0 and found to be 32?°C. Cellulase activity remained at a high level at 5?C20?°C, which is the growth temperature of the short-spined sea urchin. These results confirm that the short-spined sea urchin should preferably be reared at a water temperature of <20?°C.     

温度对中间球海胆幼胆生存、生长和行为的影响

底播增殖是一种提高中间球海胆(Strongylocentrotus intermedius)产量的有效方法.底播增殖受到多种内在和外在因素的影响,其中温度发挥着十分重要的作用,因此确定中间球海胆适宜增殖温度范围至关重要.本实验将大(23.29±0.27)mm、小(18.78±0.19)mm两种规格的中间球海胆放置在3个...     

虾夷马粪海胆体腔细胞的类型及功能

取平均壳径分别为1．9cm和4．4cm虾夷马粪海胆(Strongylocentrotus intermedius)进行实验观察。结果表明，海胆体腔有2种类型的细胞，即变形吞噬细胞和色素细胞。变形吞噬细胞形状不定，能伸出伪足，核较大，线粒体、溶酶体等细胞器丰富。色素细胞具突起，内有紫红色颗粒，颗粒溶于酒精等多种溶剂中，使得电镜下细胞内含有大量空泡，细胞核很少见，细胞器较少。变形细胞离体后可凝集，具吞噬酵母的能力，吞噬能力与温度成正相关。色素细胞具有辅助的免疫功能。     

中间球海胆生长分化相关的AFLP标记

从同一遗传背景、同环境下培养的1龄中间球海胆(Strongylocentrotus intermedius)中,分别选择大个体50只、小个体30只,分别组成大小2组海胆,两组之间壳径、壳高2个指标均存在显著差异。共选用6对AFLP引物组合对2组海胆进行PCR扩增,发现43个标记在大小2组海胆间存在显著的频率差异(P0.05),其中29个位点差异极显著(P0.01)。所有存在显著频率差异的位点中,2个位点在大海胆组中缺失,4个位点在小海胆组中缺失,14个位点在大海胆组中出现的频率显著高于小海胆组(P0.05),而23个位点在小海胆组中出现的频率高于大海胆组(P0.05)。这一系列谱带可能与中间球海胆生长性状之间存在一定的相关关系。遗传多样性分析结果表明,大海胆组的平均Shannon氏指数、Nei氏杂合度2个指标均显著高于小海胆组(P0.05),提示杂种优势可能与中间球海胆的快速生长存在一定联系。     

Estimates of genetic parameters for growth traits of the sea urchin,Strongylocentrotus intermedius

Heritabilities and genetic and phenotypic correlations were estimated for body weight, test diameter, and test height of the sea urchin from measurements on progeny resulting from 11 sires and 33 dams by artificial fertilization of 3 females by single males, and measurements at 8, 10, and 12 months after metamorphism. Point estimate for heritabilities based on the sire components of variance were moderate to high for body weight (0.21–0.49), test diameter (0.21–0.47), and test height (0.22–0.37). Genetic correlations were significant for body weight with test diameter (0.30∼0.65) and test height (0.30∼0.54) and test diameter with test height (0.31∼0.65). Genetic correlation estimates, derived the nested design and half-sib correlation analysis used in this study, appear to provide reliable estimates. Significant phenotypic correlations were found for body weight with test diameter (0.82∼0.86) and test height (0.49∼0.83), and test diameter with test height (0.47∼0.84). The phenotypic correlations for test height with body weight (0.491) and test diameter (0.467) at 12 months' of age were smaller than those earlier sampling periods.     

Heritability and phenotypic correlations of gonad sweetness in the sea urchin Strongylocentrotus intermedius

Gonad is the only edible part of sea urchins. Thus, a number of studies have been focusing on how to improve both quantity and quality of their gonads. However, as far as our knowledge, the genetic basis of gonad flavor remains totally unknown in sea urchins. In the present study, we found that the heritability of gonad sweetness was at a high level of 0.56, clearly indicating that it is, to a large extent, under genetic control. Gonad sweetness was significantly positively correlated with gonad weight (P < 0.05), a* (P < 0.01) and b* (P < 0.01), while significantly negative correlated with L* (P < 0.01), ΔE ₁ (P < 0.01) and ΔE ₂ (P < 0.01). The present study provides valuable information into the genetic basis of gonad sweetness and evidences that gonad sweetness is potential to be improved in sea urchin genetic breeding programs.     

温度和盐度对虾夷马粪海胆耗氧率和排氨率的影响↑（*）

采用封闭式呼吸器研究了虾夷马粪海胆（Ｓｔｒｏｎｇｙｌｏｃｅｎｔｒｏｔｕｓｉｎｔｅｒｍｅｄｉｕｓ）的耗氧率和排氨率与温度和盐度的关系。结果表明：在温度１５～２５℃的范围内,耗氧率和排氨率随温度的升高而增加;相同温度下,随个体重量的增加耗氧率和排氨率下降,呈负指数关系;虾夷马粪海胆的适宜盐度为３０左右,此时的耗氧率最大,为０．０９１ｍｇ／ｇ·ｈ,排氨率最低,为２３．６２μｇ／ｇ·ｈ。     

温度和盐度对中间球海胆胚胎发育早期进程联合效应

为探讨高温和低盐对中间球海胆(Strongylocentrotus intermedius)早期胚胎发育进程的影响, 本研究利用中心复合设计(CCD)和响应曲面分析法(RSM), 开展不同温度(12~26 ℃)和盐度(22~34)对中间球海胆胚胎发育早期进程的联合效应研究, 旨在建立温度和盐度对中间球海胆胚胎发育进程的定量关系模型, 并通过统计优化方法得出温度和盐度的最佳组合。结果显示, 在实验设定的温度和盐度范围内, 随着温度的升高, 中间球海胆早期胚胎发育时间呈现出先缩短后延长的趋势; 随着盐度的降低, 中间球海胆胚胎发育早期时间延长。温度的一次效应、二次效应和盐度的一次效应均显著影响(P<0.05)中间球海胆胚胎发育早期进程; 温度一次项系数的绝对值均大于盐度的一次项系数; 温盐的联合效应对中间球海胆胚胎发育早期进程的影响不显著(P>0.05)。实验建立的 2 细胞期、8 细胞期、16 细胞期、囊胚期、上浮期和四腕幼虫期发育进程模型方程决定系数分别为 0.9576、0.9508、0.9689、0.9932、 0.9681 和 0.9763。模型优化和验证试验得出, 温度 20.47 ℃和盐度 31.46 时, 中间球海胆 2 细胞期、8 细胞期、16 细胞期、囊胚期、上浮期、四腕幼虫期的发育时间最短, 分别为 1.28 h、2.07 h、3.31 h、4.14 h、11.28 h 和 47.31 h。 研究结果表明高温和低盐会延长中间球海胆早期胚胎发育时间。     

Effect of feed deprivation and refeeding on the MYP gene expression of the sea urchin (Strongylocentrotus intermedius)

To determine the main expression site of major yolk protein (MYP) gene and the mechanisms for adaptation to starvation and refeeding in Strongylocentrotus intermedius, MYP mRNA expression amounts were analysed using a real‐time RT‐PCR. The results showed that MYP could be transcribed in the intestine, stomach, gonad and coelomocytes, and that the intestine was the main expression site of MYP gene in non‐starved urchins. The MYP synthesis in the intestine decreased during 15 days of starvation (67.70%, 52.58% and 71.35% of the control at 5, 10 and 15 days of fasting respectively) and then increased dramatically by different amounts (the peaks were 2.71‐, 12.16‐ and 7.89‐fold that of the control respectively) during the refeeding stages. Nevertheless, the expression amounts in the gonads did not decline, but increased continuously during all periods of fasting (2.66‐, 3.72‐ and 13.19‐fold that of the control at 5, 10 and 15 days of starvation respectively) and during the refeeding stages. At the end of the recovery feeding experiment, the levels reached 9.58‐, 17.48‐ and 100.69‐fold that of the control. These data suggested that the ‘priority’ strategy for the sea urchin is to reduce MYP expression amounts in the intestine if food is limited and to increase MYP gene expression in the gonad to protect reproductive function.     

虾夷马粪海胆不同家系和性别间性腺性状的比较

对来自虾夷马粪海胆20个家系的600枚海胆的性腺湿重、指数、颜色、水分和脂肪酸组成进行了测量和分析,结果显示性腺湿重和性腺指数在家系间均有极显著差异,但在性别间差异不显著,两个性腺产量性状在整个群体内均具有较大的变异;虾夷马粪海胆性腺颜色介于黄色和橙红色之间,且色泽鲜艳,经CIEL*a*b*标准量化和方差分析,L*和a*在家系间无显著差异,但b*、亮黄色差(ΔE1)和亮橙黄色差(ΔE2)在家系间均具有极显著差异,在性别间,雌性海胆的L*、a*和b*值以及两个色差均极显著优于雄性;性腺水分在家系和性别间有极显著和显著差异;性腺总脂中含有较高的EPA(12.11±2.65)%和AA(10.27±3.40)%,而DHA含量较低(0.49±0.64)%,但其在群体内变异极大(130.77%),高不饱和脂肪酸在家系间具有极显著差异,而在性别间仅AA有显著差异(雄性较高)。研究结果表明,对性腺产量和品质进行家系或性别选择均可达到提升性腺产量和品质的目的。     

中间球海胆野生和养殖群体遗传结构的微卫星分析

利用28对微卫星DNA分子标记对中间球海胆的1个野生和1个养殖群体进行了遗传多样性分析。结果表明：在28个基因座中,共检测到91个等位基因,每个基因座位检测到的等位基因数为2～6个,2个群体的平均等位基因数均为3.071 4,平均有效等位基因数分别为2.231 9、2.227 1,平均观察杂合度分别为0.523 4、0.536 5,平均期望杂合度为0.486 8、0.499 3,平均多态信息含量为0.447 7、0.439 6,群体间的多态性差异不显著。2个群体间的遗传相似系数为0.758 7,遗传距离为0.276 2。经HardyWeinberg平衡的卡方检验,有50％的位点显著偏离HardyWeinberg平衡。通过F检验发现,两个群体均处于不同程度的杂合子过剩状态。群体间发生分化程度很弱,遗传变异主要来自群体内个体之间。     

Interactive effects of family and stocking density on survival and growth of the sea urchin Strongylocentrotus intermedius

To investigate the effects of family, stocking density, and their interaction on mortality, growth rate, and size dispersal in the sea urchin Strongylocentrotus intermedius, 1,440 juveniles from six full‐sib families were exposed to three stocking densities, that is, low stocking density (LD), middle stocking density (MD), and high stocking density (HD), and reared for 10 months in sea‐based suspended cages. The results demonstrated significant family effects on cumulative mortality rate (CMR); specific growth rate for test diameter (SGR for TD); specific growth rate for body weight (SGR for BW); coefficient of variation for body weight at the 10th month (CV for BW₁₀); and significant stocking density effects on SGR for TD, SGR for BW, and CV for BW₁₀. Statistically significant family by stocking density interaction was only found in SGR for TD. A certain degree of family‐ranking inversions occurred in SGR for TD. The present study provides evidence for the existence of family by stocking density interaction on the growth rate of test diameter in the family selection for S. intermedius. More attention should be paid to this interaction effect to select correct parents in S. intermedius.     

Banana peel provides a new insight into improving gonad flavor in the sea urchin Strongylocentrotus intermedius

To our knowledge, little information is available on approaches to improving gonad flavor of sea urchins. Although sea urchins fed high content of glutamate and glycine perceived sweeter gonads than those fed high content of valine and methionine (Phillips et al. in Aquaculture 288:205–215, 2009), the problem of improving gonad quality has not been completely solved. Because of the high cost of glutamate and glycine, it is hard to move this finding from the laboratory to gonad production. In the present study, we found that gonad flavor of Strongylocentrotus intermedius fed banana peel was significantly better than that of individuals fed kelp or pumpkin (P < 0.001). However, banana peel did not significantly support gonad yield of sea urchins compared with kelp (P < 0.05). This novel finding provides a new insight into the gonad quality improvement in sea urchins. Further studies should be carried out to test our two suggested methods of banana peel application in sea urchin aquaculture.     

Estimates of heritabilities and genetic correlations for growth and gonad traits in the sea urchin Strongylocentrotus intermedius

The present study is the first estimate for the sea urchin, Strongylocentrotus intermedius, of genetic parameters for growth and gonad traits during the entire reproductive period. 2165–5764 sea urchins were sampled from 74 to 120 families at six different growth stages. Six hundred individuals were sampled from 20 randomly chosen families at harvest. Coefficients of variation for most growth and gonad traits were high (18.08–78.66%) suggesting a good variation foundation for breeding. Point estimate for heritabilities based on the sire components of variance were moderate to high for test height (0.24–0.39), test diameter (0.21–0.48) and body weight (0.16–0.49). Heritabilities of gonad wet weight, gonad index and gonad moisture at harvest were 0.17, 0.41 and 0.50 respectively (moderate or high). Gonad colour parameters, L^* (lightness) and a^* (redness) have a heritability of 0, while b^* (yellowness) and colour difference ΔE₁ and ΔE₂ have very similar heritabilities of 0.2, 0.18 and 0.21 respectively. Heritabilities of β‐carotene concentration and content were 0.89 and 0.44 respectively. The genetic correlations between the two measured traits ranged from ?0.984 to 0.999. Gonad wet weight, L^*, b^*, β‐carotene concentration and β‐carotene content had significant positive genetic correlations with all the growth traits. However, gonad index, gonad moisture and colour differences ΔE₁ and ΔE₂ had significant negative genetic correlations with growth traits. The present study indicates that growth, gonad yield and most gonad quality traits could be improved by selection with moderate to high potential. Indirect selection should be applied on gonad traits using growth traits as secondary traits. The present study provides important theoretical guidance for future selective breeding programmes for S. intermedius.     

Effects of diet and temperature on post‐settlement growth and survival of the short‐spined sea urchin Strongylocentrotus intermedius

Effects of diet and temperature on post‐settlement growth and survival of a sea urchin Strongylocentrotus intermedius, which fed on one of six mono‐cultured benthic diatoms, gametophytes of a brown alga Undaria pinnatifida, a green alga Ulvella lens plus mixed benthic diatoms, or no food at 6, 9, 12 and 15°C, were examined. Digestion efficiency (DE), ingestion rate (IR), excretion rate (ER) and digested cell content index (DCCI) of each diatom species were measured. Growth rates largely differed among the dietary treatments from 7 days post‐settlement. Sea urchins fed on a diatom Cocconeis sublittoralis showed considerably higher growth (7.85‐34.67 μm day^?1) than those fed on the other diets and high survival rates (100%). DCCI was also highest in C. sublittoralis. These results suggest that the quantity of diatom cell contents digested is an important factor affecting growth and survival of the sea urchins. Growth rates of sea urchins were higher in higher water temperatures, and significant differences in growth rate were observed between 6‐9 and 12‐15°C. ERs of C. sublittoralis were generally higher at higher temperatures, but DEs did not differ among the temperature treatments. Variations in growth rate among different water temperature appear to be caused by different food intakes of the urchins.     

Isolation and characterization of bacteria associated with a syndrome disease of sea urchin Strongylocentrotus intermedius in North China

Sea urchin, Strongylocentrotus intermedius, transplanted from Japan in 1989, has been widely cultured along the coasts of Liaoning and Shandong Provinces and has become the dominant and most economically important maricultured species in North China. However, a lesion syndrome symptom of S. intermedius broke out frequently these years, showing lethargy in activities, blackish peristomial membrane and body well lesions, and brought about high mortality eventually. Six representative prominent bacterial strains were isolated from diseased sea urchin from September 2009 to January 2010. By means of API 20NE and 16S rRNA gene sequences analysis, isolates were identified as Shewanella aquimarina, Pseudoalteromonas tetraodonis, Vibrio shilonii, V. harveyi, V. fortis and V. splendidus. Bacterial challenge tests showed that their representative isolates were virulent to S. intermedius with LD₅₀ values ranging from 9.2 × 10⁴ to 3.4 × 10⁶ CFU/g body weight, among which S. aquimarina , V. fortis and P. tetraodonis were highly virulent, and the other three isolates showed moderate virulence. The results indicated that a variety of bacteria including Shewanella, Pseudoalteromonas and Vibrio were involved in the mortality of S. intermedius, and the six isolates were opportunistic pathogens of sea urchins. All isolates reported herein were sensitive to ampicillin, enrofloxacin, ofloxacin, doxycycline and florfenicol.