七带石斑鱼人工繁育技术研究进展

七带石斑鱼(Epinephelus septemfasciatus)具有经济价值高、生长速度快和低温耐受性强等特点,是东北亚温带海域理想的网箱养殖品种之一,已成为中、日、韩三国海水鱼类繁育研究的热点。由于存在雄鱼性成熟年龄高、人工驯养条件下难以实现自然产卵和初孵仔鱼弱小等难题,其稳定和高效的种苗规模化繁育技术尚未完全建立起来。本文综述了七带石斑鱼人工繁育技术研究现状,重点总结日本近20年在亲鱼繁殖、仔稚鱼培育和苗种中间培育等方面取得的研究进展,并就其发展前景进行分析,旨为中国七带石斑鱼苗种规模化繁育技术研发提供参考。     

七带石斑鱼仔鱼摄食习性的观察

在人工育苗条件下,采用轮虫与牡蛎受精卵两种开口饵料,3种不同的投喂方式：开口单喂轮虫(A)、开口单喂牡蛎受精卵(B)、开口将牡蛎受精卵和轮虫混合投喂(C),分析对比不同饵料对仔鱼开口效果。通过观察5日龄仔鱼日摄食与4~20日龄仔鱼消化道饱满度的变化,探讨仔鱼摄食习性及光照与摄食的关系。结果发现,七带石斑鱼孵化开口后,投喂3d牡蛎受精卵者比直接投喂轮虫对仔鱼生长有明显的效果;10日龄前,B组在全长、肛前距、口裂宽方面都大于A、C两组,B组全长的特定生长率为4.969%,分别是A、C两组的2.5与2.4倍,生长速度最快;5日龄仔鱼在09∶00~10∶00与13∶00~14∶00时间段,有两个明显的摄食高峰期,光照强度分别为720~1 018 lx与865~923 lx;4日龄仔鱼其饱食率较低,只有40%达到了3级摄食量,4~10日龄的仔鱼饱食率波动幅度较大,10日龄后开始逐渐上升,13日龄后基本可以达到100%。     

Skeletal development and deformities in cultured larval and juvenile seven-band grouper, Epinephelus septemfasciatus (Thunberg)

The seven‐band grouper, Epinephelus septemfasciatus (Thunberg), is currently recognized as a potential new species for aquaculture in Japan. This study describes normal and abnormal skeletal development of the jaw and vertebrae in cultured larvae and early juveniles of E. septemfasciatus. The ontogenetic stages at which skeletal deformities of jaw and vertebra developed were also described for this species. Osteological observations were made using a clearing and staining method for larvae and soft X‐ray photographs for juveniles. A high incidence of skeletal deformities was observed in the jaws and vertebral column during the larval and juvenile stages. Most of the jaw deformities were explained by an abnormal maxilla curvature. Jaw deformities were visually evident from flexion stage after ossification of the deformed elements. Deformities in the vertebral column (mostly lordosis) were observed from the post‐larval stage and became more evident as growth proceeded. The lordosis generally occurred on the positions of the 8–11th vertebra. These types of deformities are compared with those of other species, and possible causative factors of the skeletal deformities are discussed.
     

Effect of tank proportions on survival of seven-band grouper Epinephelus septemfasciatus (Thunberg) and devil stinger Inimicus japonicus (Cuvier) larvae

We examined the effects of rearing‐tank proportions on early survival, surface death and growth of the seven‐band grouper Epinephelus septemfasciatus (Thunberg) and the devil stinger Inimicus japonicus (Cuvier). Fertilized eggs were introduced into three differently shaped 100 L rearing tanks. The three tanks had different water surface areas, and included a shallow tank (S; 71 × 26 cm in diameter and depth, respectively), an intermediate tank (I; 57 × 39 cm) and a deep tank (D; 44 × 70 cm). Both species showed their highest survival rate and the lowest numbers of surface death in the D tank (P<0.05). There were no significant differences between fish reared in the three tank shapes in notochord length, total length, growth rate and dry weight. Rearing‐tank shape affected larval movement in the water column, with the duration of larval movement under the water surface being the shortest in the D tank. These results suggest that using a rearing tank of a suitable shape could significantly reduce the surface death of marine fish larvae.     

Improvement of the survival in the seven-band grouper Epinephelus septemfasciatus larvae by optimizing aeration and water inlet in the mass-scale rearing tank

The water flow in larval rearing tanks has been indicated to cause mass mortality of the seven-band grouper Epinephelus septemfasciatus larvae. Therefore, a new aerating method was tested in an actual scale intensive rearing tank (8.0 m in diameter, 1.87 m of water depth, 100 m³ of volume), in which an aerator was positioned at the center of the rearing tank surrounding cylindrical drain (1.2 m in diameter) to generate the flow field, and seven larval rearing trials were performed. The survival rate with the former aeration methods were compared, in which several aerators were located in the rearing tank. The survival rate at 10 days after hatching with the new aeration method (61.5±5.1%, n=7) was approximately three times higher than the former methods (21.2±13.7%, n=6). The flow environment of rearing tanks was also examined by quantifying the flow field, and the relationship between the flow field in the rearing tank, behavior of larvae and survival discussed. It was confirmed that the vertical circulating flow was observed in rearing tanks, and determined effectively the survival and the behavior of grouper larvae in patchiness.     

Development of the musculature and muscular abnormalities in larval seven-band grouper Epinephelus septemfasciatus

There are a number of reports documenting skeletal development in groupers, but surprisingly little is known about muscle development. We have documented muscle development in the seven-band grouper Epinephelus septemfasciatus (Thunberg), focusing primarily on the cranial muscles from 0 days post-hatching (dph) to 28 dph, using a modified whole mount immunohistochemical staining method. The eye muscles developed first followed by the cheek muscles, which completed development prior to first feeding. The muscles associated with the opercle and the levator internus 1 developed gradually and were not linked to first feeding. Later, between 12 and 16 dph, the muscle composition in the dorsal branchial arches changed to the adult form. We observed a number of cultured larvae that exhibited abnormal musculature in the body and jaw. Our results provide the first documentation of muscle development in larval seven-band grouper. This information can be used to detect abnormalities during development and provide a marker to assess the effect of changes in larval rearing techniques.     

七带石斑鱼的形态和生物学性状

2010年在广东省饶平县柘林湾进行了七带石斑鱼（Epinephelus septemfasciatus）海水网箱人工养殖研究,观察和研究了人工养成的七带石斑鱼的形态特征和生物学性状。通过观察、测量、解剖,对114尾七带石斑鱼的外部形态、可量比性状和内部结构进行了研究。结果表明,七带石斑鱼体呈椭圆形,侧扁,体侧具7条褐色横带,第1条横带仅伸达主鳃盖骨上方边缘,第6条横带分成2条,体侧横带具有不规则的白色斑纹,各鳍边缘白色,背鳍鳍棘部褐色和白色横带相间,鳍条部具有由白色横带延伸而成的3个白斑。胃呈卜型,幽门盲囊发达;肠道长度为体长的0.7～1.4倍,与七带石斑鱼为肉食性鱼类的特征相一致。     

Molecular cloning of gonadotropin cDNA in sevenband grouper, Epinephelus septemfasciatus

The cDNA for glycoprotein hormone (GPH) α-subunit, β-subunits of the FSH (GtH I) and LH (GtH II) were isolated, cloned, and sequenced from sevenband grouper pituitary using RACE PCR. These results show that sevenband grouper has two different types of gonadotropins FSH (GtH I) and LH (GtH II), as other teleosts.     

七带石斑鱼染色体核型研究

采用PHA体内直接注射法制备了七带石斑鱼(Epinephelus septemfasciatus)头肾组织染色体标本并分析其核型。结果表明,七带石斑鱼二倍体染色体数为48,24对染色体均为端部着丝粒染色体,其染色体臂数(NF)为48,核型公式为:2n=48t,NF=48。未发现与性别相关的异型染色体,第24对染色体长度明显小于其他染色体。通过与其他22种石斑鱼染色体核型进行比较,发现七带石斑鱼具有石斑鱼属鱼类的原始核型特征,属于石斑鱼属鱼类的原始类群。     

Protective immunity of sevenband grouper, Epinephelus septemfasciatus Thunberg, against experimental viral nervous necrosis

This paper describes the protective immune responses of sevenband grouper, Epinephelus septemfasciatus Thunberg, immunized with live piscine nodavirus, the causative agent of viral nervous necrosis (VNN), or the Escherichia coli – expressed recombinant coat protein. Nodavirus-neutralizing antibodies were detected at titres ranging from 1:158 to 1:1257 in serum of sevenband grouper which survived intramuscular injection with the virus, by a cell culture assay system. The virus-neutralizing ability of immune serum was also confirmed by injecting virus previously treated with serum into fish. This indicates establishment of acquired immunity in survivors and thus explains why survivors from natural infection are resistant to recurrence of the disease. Young sevenband grouper were immunized twice by intramuscular injections with the recombinant coat protein. Immunized fish produced neutralizing antibodies at high titres for at least 110 days and showed significantly lower mortalities in virus challenge tests. These results suggest the potential for vaccination against VNN in sevenband grouper, which is susceptible to piscine nodavirus at all life-stages.     

Histopathological studies on viral nervous necrosis of sevenband grouper, Epinephelus septemfasciatus Thunberg, at the grow-out stage

Abstract Viral nervous necrosis caused by sevenband grouper nervous necrosis virus (SGNNV) has occurred in grow-out stages (0-3 years old) of sevenband grouper, Epinephelus septemfasciatus, since the 1980s. In the present study, based on histopathological features of the central nervous system (CNS) in naturally diseased fish, pernasal infection experiments using grow-out fish were performed and pernasal infection was established as a putative invasion route of SGNNV. The definite SGNNV-targeted cells were determined by histopathological studies including indirect fluorescent antibody test and electron microscopy. Nerve cells in the olfactory lobe were most extensively necrotized with vacuolation followed by infiltration of microglia and macrophages. Purkinje cells and Golgi cells were extensively infected in the cerebellum. Megalocells and small nerve cell nuclei were also infected in the preoptic area, thalamus, medulla oblongata and spinal cord. Only a few small nerve cells were infected in the olfactory bulb and optic tectum. The retina of some diseased fish displayed vacuolated bipolar cells of the inner nuclear layer and in the ganglion cell layer. These SGNNV-infected nerve cells displayed viroplasmic inclusions containing virions, vacuoles and myelin-like structures. Based on observed histopathological changes, the lesion of the CNS was characterized by encephalitis but not encephalopathy.     

温度对云南盘鮈仔稚鱼生长及存活率的影响

为探究不同温度对云南盘鮈(Discogobio yunnanensis)仔稚鱼生长及存活率的影响,实验设置14、17、20、23和26℃五个温度梯度,以刚出膜且发育正常的云南盘仔鱼为实验对象,定期观察仔稚鱼的生长发育情况并统计死亡数量,测定全长、体长、体重等生长参数。结果显示:温度显著影响云南盘鮈仔稚鱼的生长发育及存活。随着温度的升高,发育进程加快;实验结束时,23℃组云南盘鮈口吸盘最早发育完整,20℃组云南盘鮈存活率最高。在整个实验过程中,20、23和26℃组间全长和体重无显著性差异,说明这几个温度对云南盘的全长、体重生长产生的效应大致相同。综合多种因素考虑,建议在实际生产中,以20~26℃作为云南盘的养殖温度。     

盐度对七带石斑鱼胚胎发育和卵黄囊仔鱼生长的影响

观察比较了8个盐度梯度[10、15、20、25、30(自然海水)、35、40和45]下七带石斑鱼Epinephelus septem fasciatus受精卵的孵化周期、孵化率和畸形率及卵黄囊仔鱼的畸形率,并在上述条件下对七带石斑鱼进行了耐饥饿试验,测定其生存活力指数(SAI).结果显示,受精卵孵化的最适盐度范围为30～35,盐度高于35,孵化率随着盐度的升高而降低,仔鱼畸形率随之升高;盐度低于25,孵化率随盐度的降低而降低,仔鱼畸形率随之升高.仔鱼的最适生存盐度为30～35,此盐度下的仔鱼存活系数为24.7±0.36和23.27土3.01.     

Effects of delayed first feeding on the survival and growth of tiger grouper,Epinephelus fuscoguttatus (Forsskål, 1775), larvae

The effects of delayed first feeding on the survival and growth of tiger grouper, Epinephelus fuscoguttatus (Forsskål 1775), larvae were examined under controlled conditions. The total length, yolk sac volume, oil globule volume, yolk sac absorption time and nutritional transition period (NTP) of the larvae fed at different first times (0, 6, 12, 18 and 24 h after the mouth opening stage; h AMO) were compared. Larval first feed intake was observed at 54 h after hatching (h AH) at 27.5 ± 0.5 °C. The yolk sac was consumed more rapidly with an increase in delayed first feeding and was significantly different among treatments (P<0.05). Larvae first fed at 0 h AMO had the longest yolk sac absorption (72 h AH) and NTP (20 h) times and had the highest survival and growth rates at the end of the experimental period (360 h AH), being significantly higher (P<0.05) than the other treatments. First mortality was observed at 69 h AH, approximately 2 h after point of no return (PNR) occurred. This study suggests that first feeding of tiger grouper larvae should commence at 0 h AMO for enhancement of larval survival and growth.     

Effects of salinity, aeration and light intensity on oil globule absorption, feeding incidence, growth and survival of early-stage grouper Epinephelus coioides larvae

ABSTRACT: A series of experiments were conducted to examine the effects of salinity, aeration and light intensity on oil globule absorption, feeding incidence, and growth and survival of early-stage Epinephelus coioides larvae. Newly hatched larvae were transferred to 40-L aquaria at a density of 1500 individuals/aquarium. Larvae were exposed to different levels of aeration (0 mL/min per L, 0.62 mL/min per L, 1.25 mL/min per L, 2.50 mL/min per L, or 3.75 mL/min per L); salinity (8 ppt, 16 ppt, 24 ppt, 32 ppt, or 40 ppt); and light intensity (0 lx, 120 lx, 230 lx, 500 lx, or 700 lx) for 4–6 days. Twenty larvae were sampled daily at 11:00 hours to measure for total length (TL), oil globule volume, and feeding incidence. Survival rates were determined by counting the total number of larvae remaining in each aquarium at the end of the experiment. Significantly higher survival rates ( P   <  0.05) were observed at aeration levels of 0.62 mL/min per L and 1.25 mL/min per L, at salinity levels of 16 ppt and 24 ppt, and at light intensities of 500 lx and 700 lx. The influence of aeration level, salinity and light intensity on oil globule absorption, feeding incidence, and growth and survival of early-stage grouper larvae are discussed.     

Effect of temperature on survival,growth and development of Mytilus galloprovincialis larvae

Mussel aquaculture is widely prevalent worldwide, but generally relies on natural seed collection, which does not always meet the needs of the producers. Thus, development of mussel hatcheries is of economic interest in some parts of the world, such as Europe; it provides opportunities not only on annual reliability of seed but also on genetic improvements. To broaden knowledge on mussel larval physiology, we carried out temperature treatments (17, 20 and 24 °C) on Mytilus galloprovincialis larvae under laboratory conditions. The trials ended when 30% of the larval population was in the post‐larval stage. The temperature coefficient Q₁₀ indicated a strong relationship between temperature and increase in growth from 17 to 20 °C, but not between 20 and 24 °C. Exposure of M. galloprovincialis larvae to 17 °C resulted in poor growth, low survival and a delayed development and was considered to be inadequate for M. galloprovincialis larval culture. Rearing the larvae at 20 or 24 °C produced better growth, higher survival rates and faster metamorphosis as compared with 17 °C. The temperature region within 20 and 24 °C was suggested as adequate for the mussel M. galloprovincialis larval culture, and implications of these results on the development of commercial hatcheries were discussed.     

Effect of temperature,hypoxia and disinfection with ozonated seawater during somitogenesis on muscular development of the trunk in larval seven‐band grouper,Epinephelus septemfasciatus (Thunberg)

The cause of abnormal musculature on both sides of the same myotome in artificially cultured seven‐band grouper Epinephelus septemfasciatus (Thunberg) larvae remains unclear. We evaluated the effect of three factors: changes in temperature (between 22 and 24°C), disinfection with ozonated seawater (0.3 mg TROs L^?1, 1 min), and hypoxia (22 or 14% saturation) during the early (20 h post‐fertilization, hpf) and mid‐ (24 hpf) somite stages on trunk muscular development in larval seven‐band grouper. The thermal regimes and disinfection with ozonated seawater had no effect on muscle development (P > 0.05, G‐test). In contrast, exposure to hypoxia during both the early and mid‐somite stages increased the incidence of abnormal musculature in the trunk (P < 0.01, G‐test). These larvae had abnormal musculature on both sides of the trunk. In addition, the number of abnormal myotomes differed among these larvae. Exposure in the early and mid‐somite stages resulted in the highest frequency of abnormalities in the 11th and 13th myotome respectively. Our results suggest that hypoxia during somitogenesis can lead to abnormal muscle development in the trunk of artificially cultured seven‐band grouper larva.     

温度变化对七带石斑鱼早期发育及开口摄食的影响

观察比较了9个温度梯度(13、15、17、19、21、23、25、27、29℃)对七带石斑鱼(Epinephelus septemfasciatus)受精卵孵化时间、孵化率和畸形率的影响。用在(21±0.5)℃条件下孵出的健康仔鱼进行耐饥饿和摄食实验,实验温度处理设两种方式:一、处理温度始终保持不变;二、将温度在处理48 h后统一调节至21℃。实验期间,每天统计仔鱼死亡数,测定不投饵条件下的存活指数(SAI),并观察仔鱼投饵后的开口摄食情况及其形态发育状况。结果表明:1)受精卵孵化的最适温度范围为17~23℃,当温度低于13℃或高于27℃时,受精卵不能孵化。21℃时,受精卵孵化率最高,为(93.67±1.52)%;而畸形率最低,为(1.06±1.06)%。当温度低于或高于21℃时其孵化率降低,而畸形率升高;2)在13~28℃范围内,初孵仔鱼的SAI值随着温度的上升先升高后逐渐降低,21℃时的SAI值最高,为(20.26±0.44)%。各组温度保持不变时,13℃和27℃的SAI值分别为(2.18±0.01)%、(8.47±0.28)%;调整一致后分别为(6.90±0.44)%、(13.30±0.31)%。温度调节组与不调节组相比,其SAI值明显提高。温度调节后,每个梯度组的畸形率也出现了不同程度的降低;3)不同温度处理的仔鱼开口摄食存在差异。21℃时,仔鱼的初始摄食率和饱食率分别可达80.0%和40.0%。随着温度偏离21℃,摄食率和饱食率均逐渐降低。温度调节恢复组的仔鱼摄食情况改善明显,其中17℃不变和恢复组对比变化最为明显:当保持17℃不变时仔鱼的摄食率和饱食率分别为20.0%、13.3%,而处理48 h后恢复到21℃后,两者分别提高到66.7%和33.3%,分别提高了2.3和1.5倍。     

Changes in Na+,K+-ATPase activity and gill chloride cell morphology in the grouper Epinephelus coioides larvae and juveniles in response to salinity and temperature

The activity of the enzyme Na⁺,K⁺-ATPase and morphological changes of gill chloride cells in grouper, Epinephelus coioides larvae and juveniles were determined 6–48 h after abrupt transfer from ambient rearing conditions (30–32 ppt, 26.5–30 °C) to different salinity (8, 18, 32, 40 ppt) and temperature (25, 30 °C) combinations. Na⁺,K⁺-ATPase activity in day 20 larvae did not change at salinities 8–32 ppt. Activity decreased significantly (P <0.01) after exposure to 40 ppt at 25–30 °C, which was accompanied by an increase (P <0.05) in density and fractional area of chloride cells. Enzyme activity in 40 ppt did not reach a stable level and larvae failed to recover from an osmotic imbalance that produced a low survival at 25 °C and death of all larvae at 30 °C. Enzyme activity and chloride cell morphology in day 40 groupers did not change in 8–40 ppt at 25 °C and 8–32 ppt at 30 °C. A significant decrease and a subsequent increase in Na⁺,K⁺-ATPase activity in 40 ppt at 30 °C was associated with the increase in chloride cell density resulting in an increased fractional area but a decreased cell size. Enzyme activity and chloride cells of day 60 grouper were unaffected by abrupt transfer to test salinities and temperatures. These results demonstrate that grouper larvae and juveniles are efficient osmoregulators over a wide range of salinities. Salinity adaptation showed an ontogenetic shift as the larvae grew and reached the juvenile stage. This development of tolerance limits may reflect their response to actual conditions existing in the natural environment.