墨西哥湾扇贝初级卵母细胞发育及卵黄发生超微结构的初步研究

采用电子显微镜术,研究了墨西哥湾扇贝Argopecten irradians concentricus的初级卵母细胞发育和卵黄发生。结果表明,墨西哥湾扇贝初级卵母细胞的发育可分为卵黄合成前期、卵黄合成期和成熟期;5种细胞器,即线粒体、高尔基液泡、糙面内质网、多泡小体和微吞饮泡参与了初级卵母细胞卵黄颗粒的形成,其中,线粒体和高尔基液泡是形成卵黄颗粒的主要胞器。卵黄物质的来源有内源性和外源性两种方式,外源性卵黄物质主要来自消化腺中贮存的营养。另外,研究发现核膜泡可能参与了多泡小体的形成。     

Effect of increasing salinity on physiological response in juvenile scallops Argopecten purpuratus at two rearing temperatures

Argopecten purpuratus can be cultivated using Recirculating Aquaculture Systems (RAS) as a method to increase production. In order to determine physiological response of A. purpuratus under different salinities and temperature conditions, two groups of juvenile scallops (small: h = 6.5 mm and large: h = 25.5 mm) were acclimated and close-cultured at salinities of 34, 38, and 42 g/l, at 16 and 22 °C and fed on Isochrysis galbana and Chaetoceros calcitrans. Survival, shell growth and scope for growth were determined at the end of the trials. Survival showed an inverse relationship with temperature and ammonia levels. In small scallops an increase in salinity at 16 °C increased survival. However, this relationship was not evident at 22 °C. On the other hand, salinity did not affect survival of large juveniles. Small juveniles had a lower survival (approximately 40%) than larger scallops (up to 85%) throughout the trials. Oxygen consumption was not affected by salinity. Small scallops showed similar oxygen consumption at 16 and 22 °C but in large juveniles higher values were registered at 22 °C. In large juveniles routine consumption at 16 °C was higher (up to 35%) than standard consumption. This pattern was not evident at 22 °C, suggesting that oxygen demand is higher regardless of feeding condition. NH₄⁺–N excretion rate is inversely related to salinity. Only small juveniles showed a higher NH₄⁺–N excretion rate at 22 °C. Scope for growth was positive in all treatments, although the upper limit of salinity should not be based only in this index. Higher scope for growth values at 38 and 42 g/l was related with a reduction in ammonia excretion and high absorption efficiency. In addition, an increase in salinity produced a reduction in NH₃–N proportion and under hypersaline conditions scallops tended to decrease excretion as a way of osmoconformation. This explains our findings of higher survival rates at higher salinities. Even though the scope for growth is positive at 42 g/l, the osmotic stress reduces the survival chances. The data obtained can be considered useful information for A. purpuratus culture under controlled conditions.     

Larval rearing of calico scallops, Argopecten gibbus, in a flow-through system

A flow-through (FT) culture system is described for calico scallop, Argopecten gibbus, larvae. Its performance was assessed by larval survival rate, shell growth, settlement rate and post-larval shell growth for the duration of larval life (13 days). Comparisons were made with larvae reared in standard static system (S). Effect of increased larval density on FT performance was also investigated. With comparable larval densities, survival rate of Day 2 larvae to pediveliger stage was similar in both larval rearing systems. Shell growth for FT-reared larvae was comparable or significantly higher than in the static system (P < 0.01). Settlement rate of pediveligers was comparable for both systems, averaging 30.7%, and no significant difference was seen in shell growth of FT- and static-reared pediveligers. Increased initial larval density did not affect survival rate in FT, but did negatively affect larval shell growth, settlement rate and post-larval shell growth yielding lower growth and minimal settlement rate (10.9 ± 2.8%) compared to the static system. This FT system was successful as larval rearing system, optimising space allocation in the hatchery, reducing labour, and eliminating the use of antibiotics. Optimising initial larval density within the system needs to be investigated in association with food ration.     

人工养殖栉孔扇贝和海湾扇贝血淋巴中部分免疫因子的比较研究

采用分光光度技术对人工养殖的栉孔扇贝和海湾扇贝血淋巴中部分免疫因子进行了比较研究。结果表明,海湾扇贝血清和血细胞中溶菌酶、非特异性酯酶(NSE)、过氧化氢酶(CAT)、髓性过氧化物酶(MPO)、一氧化氮(NO)、一氧化氮合酶(NOS)、活性氧和H2O2的活力或含量均高于栉孔扇贝,两种扇贝除血清中MPO活力差异不显著外,其他各组均差异极显著或显著。表明人工养殖海湾扇贝较栉孔扇贝的免疫力强。     

A method to eliminate self-fertilization in a simultaneous hermaphrodite scallop. 1. Effects on growth and survival of larvae and juveniles

Argopecten purpuratus, as several other scallops of commercial importance is a functional hermaphrodite. The usual methods for breeding cause an unavoidable degree of self-fertilization with the subsequent deleterious effects on survival and growth rates. Some researchers have unsuccessfully attempted to eliminate self-fertilization in these bivalves. Applying our knowledge of the cellular mechanisms of spawning in this species we have developed a method by which self-fertilization was eliminated. By this in vitro fertilization method, oocytes were obtained by scraping the female portion of the gonad, meiosis resumption was induced by serotonin, and then the oocytes were fertilized with sperm from different individuals. The progeny obtained by this method were compared with others obtained by cross-fertilization and self-fertilization. In all these procedures, the oocytes were fertilized at a ratio of 60 sperm per oocyte. The fertilization rates and D larvae survival were determined at 2 and 48 h, respectively. After the D larval stage, the hatchery culture of larvae continued as usual with sampling for survival and growth rates every three days. When larvae looked ready to settle and metamorphose, netlon collectors were introduced into the tanks. After 12 days the collectors were placed in 1 mm mesh collection bags and transferred to the sea. Juveniles were brought back to the hatchery every 40-70 days, to be counted, measured, and placed in mesh pearl nets. The assay of the three procedures for fertilization showed similar fertilization rates but low survival for D larvae obtained from the in vitro method. However, this procedure resulted in the highest size and survival rates of larvae. Survival of juveniles coming from this fertilization procedure was also the highest. The growth of juveniles obtained by in vitro fertilization was also higher than that of those obtained by self-fertilization. We conclude that this in vitro fertilization procedure provides a methodology that results in higher survival and growth rates of juveniles than obtained by procedures involving any degree of inbreeding. This in vitro procedure should be applicable to other simultaneous hermaphroditic scallops.     

间接免疫荧光法检测栉孔扇贝急性病毒性坏死症病毒

用纯化栉孔扇贝(Chlamys farrei)急性病毒性坏死症病毒(AVNV)免疫兔子,以兔抗血清为一抗,荧光标记的羊抗兔抗体为二抗,采用冰冻切片技术,建立了栉孔扇贝AVNV的间接免疫荧光检测方法。用该方法分析栉孔扇贝和海湾扇贝(Argopecten irradians)体内AVNV感染强度,并对感染率进行统计。结果表明,栉孔扇贝的肾脏、肝胰腺中,AVNV阳性信号最强,呈现中度到重度感染,其AVNV感染率100％。鳃丝、性腺及闭壳肌中未检测到阳性信号。在海湾扇贝的肝胰腺及肾中AVNV的感染率也较高。提示AVNV感染扇贝的靶器官主要是肾脏、肝胰腺。对与栉孔扇贝同一海区养殖的海湾扇贝在栉孔扇贝发病期间存活率较高的原因进行了讨论。     

海湾扇贝病原菌-黑美人弧菌的分离与致病性

1997年8～9月,大连金州区大李家乡养殖场筏养海湾扇贝(Argopectenirradians)突发大面积死亡。其主要病症是外套膜收缩,重者全部脱落,鳃丝呈土黄色或糜烂,肠管空并伴有不同程度的肾肿胀。从病贝血淋巴液和鳃组织中分离的2株优势菌(h9701-1,g9701-2)分别做纯培养,采用肌注法感染健康贝,得到与原发病相同症状,证实是病原菌。经多项形态、生理生化测定,2株分离菌皆是黑美人弧菌(Vibrionigripulchritudo)。     

胶州湾不同水层中养殖海湾扇贝生长与死亡的比较

本试验于1985年在胶州湾进行。结果表明:表面下2米水层养殖海湾扇贝最好,个体生长较大,死亡较少,1米水层养者次之,3米者再次之,海底者最差。养殖笼上的附着物随水深加大而数量减小,1米水层者为2米者的2倍,3米者的8倍,底层者的20倍。1米水层大量滋生的附着物对该水层养殖的海湾扇贝造成较大的不良影响。     

海湾扇贝消化系统粘液细胞的类型与分布

采用组织化学方法，AB—PAS染色，对人工养殖的1龄海湾扇贝(Argopecten irradians，壳长35—40mm)进行显微观察，发现海湾扇贝唇瓣、口唇、胃、肠、直肠、晶杆囊和肝脏中均有粘液细胞分布，其中，内唇瓣的内表皮和外唇瓣的外表皮粘液细胞数量较多，其中Ⅱ型(AB—PAS染色呈蓝色)最多，Ⅳ型(蓝紫色)次之，Ⅲ型(紫红色)较少，I型(红色)极少；内唇瓣的外表皮和外唇瓣的内表皮粘液细胞仅在每一横褶的皱褶处集中分布，多为Ⅱ型，另有少量Ⅳ型。口唇粘液细胞较多，排列比较紧密，Ⅱ型最多，Ⅳ型次之。胃粘液细胞数量很少，均为Ⅱ型。肠粘液细胞(Ⅱ型)较多，在肠上皮细胞较薄的区域分布密集。直肠粘液细胞(Ⅱ型)较多。晶杆囊粘液细胞(Ⅱ型)仅分布于局部。粘液细胞(Ⅱ型)在肝脏仅有少量分布于输出管壁和外上皮中。     

海湾扇贝类衣原体寄生及人工感染试验

于1999年10月至2001年3月海湾扇贝(Argopectenirridians)大量死亡期间,对不同生长阶段的海湾扇贝进行了组织学及超微结构观察,在贝消化腺上皮细胞内发现嗜碱性的类衣原体包涵体,该包涵体为圆形及不规则形状,大小约9μm×7μm,HE染色呈兰色。包涵体发育至晚期破裂,宿主细胞崩解,类衣原体逸出。超微结构观察显示,包涵体内部含有大小不同的3种个体形态,即网状体、原体和中间体。网状体大而疏松,呈圆形或不规则形,大小为(890.5±164.6)nm×(623.6±129.3)nm(n=20);原体个体小而致密,呈较为整齐的短棒状或纺锤形,大小为(317.5±40.1)nm×(180.3±40.0)nm(n=18);中间体数量较少,大小介于上述2种个体之间,可能处于二者的过渡阶段。将类衣原体分离后对海湾扇贝的幼体、稚贝和成贝进行了人工感染,结果表明,类衣原体可以感染海湾扇贝不同发育阶段(幼贝、稚贝、成贝)的个体,受感染个体生长缓慢,内脏干瘪,易从附着基上脱落。但不同阶段的贝体感染率与死亡率差异较大,幼体组感染率和死亡率分别达80%和90%以上;稚贝组分别达50%和70%左右;成贝组的感染率达50%以上,但死亡率与空白组及对照组无显著差异。说明类衣原体在一定条件下可造成对幼体和稚体扇贝的危害,但对成贝影响不大,不能导致成贝宿主的大量死亡。