Development study on some digestive enzymes of Takifugu rubripes larvae and juvenile

The activities of some digestive enzymes are studied for Takifugu rubripes larvae and juvenile from the first feeding to 27d after hatching at selected stages of development. The homogenate of whole larvae body is used for enzymatic determination. Activity of acid protease decreases notably during the beginning days after the commencement of completely exogenous feeding and the days before the beginning of the juvenile stage. Alkaline protease specific activity also decreases at metamorphosis. The activities are associated with the morphology of the developing digestive tract. Amylase activity increases before the first feeding, followed by a decreasing and then a rather constant level. Lipase activity remains low during the larvae and juvenile periods. Alkaline phosphatase activity increases gradually. This reflects the development of brush border membranes of enterocytes.     

军曹鱼仔鱼期的摄食与生长

以人工孵化的军曹鱼(Rachycentron canadum)卵黄囊期仔鱼为材料,对仔鱼早期阶段的摄食及生长情况进行研究。结果表明:在水温28~32℃条件下,军曹鱼仔鱼3日龄开始摄食,6日龄时卵黄完全耗尽,6~7日龄饥饿仔鱼进入不可逆点(PNR)期;混合营养期3~4 d。饥饿仔鱼的初次摄食率开始较低,此后上升,到卵黄囊快要耗尽时达高峰(50%),以后又下降。内营养期,仔鱼日平均生长率为0.86 mm/d;PNR前的摄食期,饥饿仔鱼的日平均增长率为0.068 mm/d,而摄食仔鱼的日平均生长率为0.64 mm/d;PNR后至死亡前,饥饿仔鱼体长呈负增长,而摄食仔鱼的日平均生长率达1.04 mm/d。摄食仔鱼在孵化后8 d内的全长和日龄呈指数关系;前10 d的口径与全长呈正比关系。     

Ontogenetic changes in digestive enzyme activities and the amino acid profile of starry flounder Platichthys stellatus

Ontogenetic changes in digestive enzyme activities and the amino acid(AA) profile of starry flounder,Platichthys stellatus,were investigated and limiting amino acids were estimated compared with the essential AA profile between larvae and live food to clarify starry flounder larval nutritional requirements.Larvae were collected at the egg stage and 0,2,4,7,12,17,24 days after hatching(DAH)for analysis.Larvae grew from 1.91 mm at hatching to 12.13 mm at 24 DAH.Trypsin and chymotrypsin activities changed slightly by 4 DAH and then increased significantly 4 DAH.Pepsin activity increased sharply beginning 17 DAH.Lipase activity increased significantly 4 DAH and increased progressively with larval growth.Amylase activity was also detected in newly hatched larvae and increased 7 DAH followed by a gradual decrease.High free amino acid(FAA) content was detected in starry flounder eggs(110.72mg/g dry weight).Total FAA content dropped to 43.29 mg/g in 4-DAH larvae and then decreased gradually to 13.74 mg/g in 24-DAH larvae.Most FAAs(except lysine and methionine) decreased 50%in 4-DAH larvae compared with those in eggs and then decreased to the lowest values in 24-DAH larvae.Changes in the protein amino acid(PAA) profile were much milder than those observed for FAAs.Most PAAs increased gradually during larval development,except lysine and phenylalanine.The percentages of free threonine,valine,isoleucine,and leucine decreased until the end of the trial,whereas the protein forms of these four AAs followed the opposite trend.A comparison of the essential AA composition of live food(rotifers,Artemia nauplii,and Artemia metanauplii) and larvae suggested that methionine was potentially the first limiting AA.These results may help develop starry flounder larviculture methods by solving the AA imbalance in live food.Moreover,the increased digestive enzyme activities indicate the possibility of introducing artificial compound feed.     

饥饿和再投喂对尼罗罗非鱼蛋白酶活性的影响

测定了饥饿和再投喂的尼罗罗非鱼胃、肠、肝胰脏蛋白酶活性。结果表明:(1)尼罗罗非鱼蛋白酶的活性以胃为最高,肠为最低,且胃蛋白酶的活性极显著高于肠、肝胰脏(P<0.01),肠和肝胰脏之间差异不显著(P>0.05);(2)饥饿后,蛋白酶活性呈降低的趋势,饥饿处理25d后,胃、肠、肝胰脏的蛋白酶活力分别降低到饥饿处理前的42.7%、41.0%、61.5%,均与对照组差异极显著(P<0.01);(3)再投喂后,饥俄15d以内的鱼蛋白酶活性上升迅速,经过15d恢复投喂,基本上就恢复到了正常水平,且消化酶活性甚至高于一直投喂对照组,而饥饿20、25d的鱼蛋白酶活性虽缓慢上升,但仍无法恢复到正常水平,与对照组差异显著(P<0.05)。     

Ontogenetic changes in digestive enzyme activities and the amino acid profile of starry flounder <Emphasis Type="Italic">Platichthys stellatus</Emphasis>

Ontogenetic changes in digestive enzyme activities and the amino acid (AA) profile of starry flounder, Platichthys stellatus, were investigated and limiting amino acids were estimated compared with the essential AA profile between larvae and live food to clarify starry flounder larval nutritional requirements. Larvae were collected at the egg stage and 0, 2, 4, 7, 12, 17, 24 days after hatching (DAH) for analysis. Larvae grew from 1.91 mm at hatching to 12.13 mm at 24 DAH. Trypsin and chymotrypsin activities changed slightly by 4 DAH and then increased significantly 4 DAH. Pepsin activity increased sharply beginning 17 DAH. Lipase activity increased significantly 4 DAH and increased progressively with larval growth. Amylase activity was also detected in newly hatched larvae and increased 7 DAH followed by a gradual decrease. High free amino acid (FAA) content was detected in starry flounder eggs (110.72 mg/g dry weight). Total FAA content dropped to 43.29 mg/g in 4-DAH larvae and then decreased gradually to 13.74 mg/g in 24-DAH larvae. Most FAAs (except lysine and methionine) decreased >50% in 4-DAH larvae compared with those in eggs and then decreased to the lowest values in 24-DAH larvae. Changes in the protein amino acid (PAA) profile were much milder than those observed for FAAs. Most PAAs increased gradually during larval development, except lysine and phenylalanine. The percentages of free threonine, valine, isoleucine, and leucine decreased until the end of the trial, whereas the protein forms of these four AAs followed the opposite trend. A comparison of the essential AA composition of live food (rotifers, Artemia nauplii, and Artemia metanauplii) and larvae suggested that methionine was potentially the first limiting AA. These results may help develop starry flounder larviculture methods by solving the AA imbalance in live food. Moreover, the increased digestive enzyme activities indicate the possibility of introducing artificial compound feed.     

盐度对虎斑乌贼(Sepia pharaonis)受精卵孵化及幼体活力的影响

水温22±1℃条件下,研究不同盐度(15.0、18.0、21.0、24.0、27.0、30.0、33.0)对虎斑乌贼(Sepia pharaonis)受精卵孵化及幼体活力的影响。结果表明,受精卵孵化适宜盐度范围为27.0~33.0,最适盐度范围为30.0~33.0,随着盐度增加,培育周期和孵化周期呈逐渐缩短趋势,而孵化率则呈先升后降趋势。盐度15.0环境中,初孵幼体经24h累计死亡率为100%;盐度18.0、21.0中,24h的累计死亡率分别为4.15%和5.83%,不投饵存活8d和9d;盐度24.0~33.0中,24h幼体累计死亡率均为0,不投饵存活9~10d。随着盐度的增加,幼体的不投饵存活系数(SAI值)先升后降,在盐度30.0时达到最大值,盐度(y)与SAI值(x)间的函数关系可表示为:y=-0.1764x2+9.807 9x-104.33(r2=0.927 8)。     

Effects of Pseudoalteromonas sp. BC228 on digestive enzyme activity and immune response of juvenile sea cucumber (Apostichopus japonicus)

A marine bacterium, Pseudoalteromonas sp. BC228 was supplemented to feed in a feeding experiment aiming to determine its ability of enhancing the digestive enzyme activity and immune response of juvenile Apostichopus japonicus. Sea cucumber individuals were fed with the diets containing 0(control), 105, 107 and 109 CFU g-1 diet of BC228 for 45 days. Results showed that intestinal trypsin and lipase activities were significantly enhanced by 107 and 109 CFU g-1 diet of BC228 in comparison with control(P 0.01). The phagocytic activity in the coelomocytes of sea cucumber fed the diet supplemented with 107 CFU g-1 diet of BC228 was significantly higher than that of those fed control diet(P 0.05). In addition, 105 and 107 CFU g-1 diet of BC228 significantly enhanced lysozyme and phenoloxidase activities in the coelomic fluid of sea cucumber, respectively, in comparison with other diets(P 0.01). Sea cucumbers, 10 each diet, were challenged with Vibrio splendidus NB13 after 45 days of feeding. It was found that the cumulative incidence and mortality of sea cucumber fed with BC228 containing diets were lower than those of animals fed control diet. Our findings evidenced that BC228 supplemented in diets improved the digestive enzyme activity of juvenile sea cucumber, stimulated its immune response and enhanced its resistance to the infection of V. splendidus.     

An integrative study of larval organogenesis of American shad <Emphasis Type="Italic">Alosa sapidissima</Emphasis> in histological aspects

We describe organogenesis at a histological level in American shad (Alosa sapidissima) larvae from 0 until 45 days after hatching (DAH). Larval development was divided into four stages based on the feeding mode, external morphological features, and structural changes in the organs: stage 1 (0–2 DAH), stage 2 (3–5 DAH), stage 3 (6–26 DAH) and stage 4 (27–45 DAH). At early stage 2 (3 DAH), American shad larvae developed the initial digestive and absorptive tissues, including the mouth and anal opening, buccopharyngeal cavity, oesophagus, incipient stomach, anterior and posterior intestine, differentiated hepatocytes, and exocrine pancreas. The digestive and absorptive capacity developed further in stages 2 to 3, at which time the pharyngeal teeth, taste buds, gut mucosa folds, differentiated stomach, and gastric glands could be observed. Four defined compartments were discernible in the heart at 4 DAH. From 3 to 13 DAH, the excretory systems started to develop, accompanied by urinary bladder opening, the appearance and development of primordial pronephros, and the proliferation and convolution of renal tubules. Primordial gills were detected at 2 DAH, the pseudobranch was visible at 6 DAH, and the filaments and lamellae proliferated rapidly during stage 3. The primordial swim bladder was first observed at 2 DAH and started to inflate at 9 DAH; from then on, it expanded constantly. The spleen was first observed at 8 DAH and the thymus was evident at 12 DAH. From stage 4 onwards, most organs essentially manifested an increase in size, number, and complexity of tissue structure.     

Ontogenetic development in the morphology and behavior of loach （Misgurnus anguillicaudatus） during early life stages

Loach （Misgurnus anguillicaudatus） is a commercially important fish in China and an ideal aquaculture species. However, culturists experience high larval and juvenile mortality during mass production. To provide insight into ways to improve larviculture techniques, we describe the morphological characteristics and behavior of loach during the larval and early juvenile stages. Yolksae larvae ranged from 2.8 to 4.0 mm body length （BL） between days 0 to 4; preflexion larvae ranged from 3.6 to 5.5 mm BL between days 4 to 6; flexion larvae ranged from 4.8 to 8.1 mm BL between days 5 and 14; and post-flexion larvae ranged from 7. l to 15.7 mm BL between days 11 to 27; the minimum length and age of juveniles was 14.1 mm BL and 23 d, respectively. Loach are demersal from hatch through to the early juvenile stages. A suite of morphological characteristics （e.g., external gill filament and ventral mouth opening） and behavioral traits have developed to adapt to demersal living. We observed positive allometric growth in eye diameter, head length, head height, and pectoral fin length during the early larval stages, reflecting the priorities in the development of the organs essential for survival. Our results provide a basis for developing techniques to improve the survival of larval and juvenile loach during mass production.     

饲喂频率对奥尼罗非鱼仔稚鱼生长、体成分和消化酶活力的影响

研究在(28.0±1)℃条件下,5种不同饲喂频率对奥尼罗非鱼(Oreochromis niloticus×O.aureus)仔稚鱼的生长、体成分及几种消化酶活力的影响。结果显示:随着饲喂频率的增加,奥尼罗非鱼仔稚鱼的摄食率和特定生长率逐步提高,1次/d组的摄食率和特定生长率显著低于其他组(P<0.05);2、3、4、5次/d组的相对增重率之间差异不显著(P>0.05),但显著高于1次/d组;各组的饲料转化率随饲喂频率的增加而降低;胃蛋白酶、脂肪酶和淀粉酶活力随饲喂频率的增加而显著降低(P<0.05);各组胰蛋白酶变化差异不明显(P>0.05);随饲喂频率的增加,奥尼罗非鱼仔稚鱼的灰分和蛋白质含量无显著变化(P>0.05);脂肪含量显著上升,水分含量则明显下降(P<0.05)。分析表明,奥尼罗非鱼仔稚鱼最适饲喂频率为3次/d。     

Histological Study on the Digestive System Development of Takifugu rubripes Larvae and Juvenile

The digestive tract of Takifugu rubripes during early life stages was studied with light microscopy. At the beginning of hatching, the digestive tract is represented by a simple and undifferentiated straight tube and does not communicate with the exterior, as the mouth and anus are not open yet. At 2 d after hatching, a constriction between intestine and rectum that will become the intestino-rectal valve is visible. During the endogenous feeding phase, the yolk sac is resorbed and the digestive tract becomes functional and differentiated. The liver and pancreas also become apparent at this time. At onset of exogenous feeding （3 d after hatching）, yolk sac resesves are not completely depleted, suggesting a period of mixed nutrition. The digestive tract differentiates fully into b.uccopharynx, esophagus, stomach, intestine and rectum. At 9 d after hatching, the yolk sac reserves are completely depleted. The most noticeable events occurring from 5 to 17 d after hatching are the transformation of the epithelia type, the differentiation of the pneumatic sac, the epithelial cell, gut convolution, mucosal fold increase, and the growth of liver and pancreas. From 18 to 27 d after hatching, the numbers of intestine folds and mucus cells increase progressively. From then on, morphological changes of digestive features are almost completed. At 27 d after hatching, the morphology and function of digestive system are similar to that of the adult.     

Effects of Hanseniaspora opuntiae C21 on the growth and digestive enzyme activity of juvenile sea cucumber Apostichopus japonicas

The effects of a diet containing Hanseniaspora opuntiae C21 on growth and digestive enzyme activity were estimated in juvenile Apostichopus japonicus. Groups of sea cucumbers were fed diets containing H. opuntiae C21 at 0(control), 10 4, 10 5, and 10 6 CFU(colony-forming units)/g feed. Results showed that after 45 d the specific growth rate(SGR) of sea cucumbers fed a C21-supplemented diet at 10 4 CFU/g feed was significantly higher than that of the control( P 0.05). Intestinal trypsin and lipase activities were significantly enhanced by C21 administration at 10 4 and 10 5 CFU/g feed compared with the control( P 0.05). After feeding for 23–42 d, C21 was demonstrated by denaturing gradient gel electrophoresis to be present in the intestine of sea cucumbers. In addition, after feeding the C21-supplemented diets for 15 d, the sea cucumbers were switched to an unsupplemented diet and C21 was confirmed to be capable of colonizing the intestine for at least 31 d after cessation of feeding. In conclusion, C21 was shown to successfully colonize the intestine of juvenile A. japonicus via dietary supplementation, and improve growth and digestive enzyme activity.     

罗非鱼开口饵料的初步研究

研究了配合饵料、轮虫、蛋黄、轮虫+人工配合饵料(3～12日龄投喂轮虫,13日龄起投喂配合饵料)等4种开口饵料对奥尼罗非鱼仔鱼生长、发育和存活影响。结果表明,1)投喂不同饵料对奥尼罗非鱼仔鱼生长具有显著影响(P<0.05),轮虫+配合饵料组生长较快,绝对增重率和特定增重率分别为0.011g/d和17.18%d-1;蛋黄组生长最慢,绝对增重率和特定增重率与其他各组差异显著(P<0.05)。2)轮虫+配合饵料与轮虫组的仔鱼成活率较高,分别为90.3%和89.7%,2组差异不显著(P>0.05),且仔鱼发育较好,大部分个体进入稚鱼阶段;蛋黄组成活率最低,只有40.7%,个体发育较为缓慢,与其他各组差异显著(P<0.05)。可见,投喂鲜活的天然饵料(轮虫)+配合饵料是罗非鱼苗种培育的最佳开口饵料。     

Effect of dietary genistein on growth performance,digestive enzyme activity,and body composition of Nile tilapia <Emphasis Type="Italic">Oreochromis niloticus</Emphasis>

An 8-week feeding experiment was performed to evaluate the effect of dietary genistein on growth performance, body composition, and digestive enzymes activity of juvenile Nile tilapia (Oreochromis niloticus). Four isonitrogenous and isoenergetic diets were formulated containing four graded supplements of genistein: 0, 30, 300, and 3 000 μg/g. Each diet was randomly assigned in triplicate to tanks stocked with 15 juvenile tilapia (10.47±1.24 g). The results show that 30 and 300 μg/g dietary genistein had no significant effect on growth performance of Nile tilapia, but the higher level of genistein (3 000 μg/g) significantly depressed the final body weight and specific growth rate. There was no significant difference in survival rate, feed intake, feed efficiency ratio or whole body composition among all dietary treatments. An assay of digestive enzymes showed that the diet containing 3 000 μg/ggenistein decreased stomach and hepatopancreas protease activity, and amylase activity in the liver and intestine, while a dietary level of 300 μg/g genistein depressed stomach protease and intestine amylase activities. However, no significant difference in stomach amylase activity was found among dietary treatments. Overall, the results of the present study indicate that a high level of dietary genistein (3 000 μg/g, or above) would significantly reduce the growth of Nile tilapia, partly because of its inhibitory effect on the activity of major digestive enzymes. Accordingly, the detrimental effects of genistein, as found in soybean products, should not be ignored when applied as an alternative ingredient source in aquaculture.     

温度对奥尼罗非鱼仔稚鱼生长、饲料利用和消化酶活力的影响

研究了温度22、25、28、31和34℃下奥尼罗非鱼(Oreochromis niloticus×O.aureus)仔稚鱼生长、饲料利用和消化酶活力。结果表明:1)不同温度对奥尼罗非鱼仔稚鱼生长有显著影响(P<0.05),温度为31℃时,仔稚鱼生长速度和绝对增重率最高,分别为0.08cm/d和0.035g/d;温度为22℃时最低,仅0.06cm/d和0.020g/d。2)饲料系数在28℃时最低,仅1.90;22和34℃时较高,达2.14和2.17,各组差异显著(P<0.05)。3)在仔稚鱼发育和存活率上,当温度在22～34℃时,温度越高,仔稚鱼发育越好;各组仔稚鱼的存活率介于93.7%～94.7%,差异不显著(P>0.05)。4)在消化酶活力上,胃蛋白酶、胰蛋白酶和淀粉酶活力随温度升高而升高,均在34℃时活力最高;脂肪酶活力在31℃时达到最高,22℃时活力最低。     

花胚胎发育及仔鱼阶段同工酶的初步分析

<正>鱼类胚胎发育是一个复杂而有序的系统工程,是基因时空综合表达结果。已有的研究证明:机体在生长发育过程中会发生一系列复杂的同工酶时空表达变化,它是基因特异性表达的一种主要作用形式。同工酶研究是研究鱼类个体发育的有力手段之一,     

BIOCHEMICAL CHANGES DURING DEVELOPMENT OF EGGS AND YOLK-SAC LARVAE OF HERRING AND PLAICE

Changes in quantities of water, sodium, potassium, lipid and protein were compared in developing eggs and yolk-sac larvae of herring dupes harengus L., a species with demersal eggs, and plaice Pleuroncctcs platessa L., a species with pelagic eggs. The principal difference between the two species was the occurrence in herring, but not in plaice of considerable increases in water content and in one or both of the above ions at three distinct stages: at fertilization,during the middle stage of egg development, and in the days immediately after hatching. Newly hatched larvae of the two species differed greatly in water content because of the difference in mass and water content of the yolk, not in the water content of the rest of the body. In herring, there were increases in sodium and potassium which coincided broadly with the increase in water content. These differences are related to the gradual acquisition of buoyancy by herring during this period, comparable with the buoyancy conferred on plaice eggs a     

池养条件下军曹鱼早期发育阶段的生长和摄食

研究了军曹鱼(Rachycentron canadum)早期阶段的生长和摄食特性。结果表明:1~29日龄间,全长与日龄之间呈指数函数相关:y=3.6922e0.0995x(R=0.9913);体重与日龄间呈指数函数相关:y=0.0002e0.276x(R=0.9830);体重与全长之间为幂函数相关:y=5×10-6x2.786(R=0.9957);口径与日龄的关系为直线函数关系:y=0.1034x-0.0044(R=0.9939)。鱼苗早期阶段的生长速度存在显著性差异。军曹鱼早期鱼苗在3~6日龄时主要摄食原生动物、单轮幼虫和轮虫;7~12日龄时主要摄食桡足类无节幼体;13日龄以后主要摄食桡足类成体。仔、稚鱼阶段,军曹鱼的摄食活动呈现明显的昼夜节律,其饱满度指数均在12时前后出现最高峰,日摄食率分别为57.36%和14.78%。     

The potential of ocean acidification on suppressing larval development in the Pacific oyster Crassostrea gigas and blood cockle Area inflata Reeve

We evaluated the effect of pH on larval development in larval Pacific oyster(Crassostrea gigas) and blood cockle(Arca inflata Reeve).The larvae were reared at pH 8.2(control),7.9,7.6,or 7.3beginning 30 min or 24 h post fertilization.Exposure to lower pH during early embryonic development inhibited larval shell formation in both species.Compared with the control,larvae took longer to reach the D-veliger stage when reared under pH 7.6 and 7.3.Exposure to lower pH immediately after fertilization resulted in significantly delayed shell formation in the Pacific oyster larvae at pH 7.3 and blood cockle larvae at pH 7.6 and 7.3.However,when exposure was delayed until 24 h post fertilization,shell formation was only inhibited in blood cockle larvae reared at pH 7.3.Thus,the early embryonic stages were more sensitive to acidified conditions.Our results suggest that ocean acidification will have an adverse effect on embryonic development in bivalves.Although the effects appear subtle,they may accumulate and lead to subsequent issues during later larval development.