Effects of calanoid copepod Schmackeria poplesia as a live food on the growth, survival and fatty acid composition of larvae and juveniles of Japanese flounder, Paralichthys olivaceus

Zooplankton constitutes a major part of the diet for fish larvae in the marine food web, and it is generally believed that copepods can meet the nutritional requirements of fish larvae. In this study, calanoid copepod Schmackeria poplesia, rotifer Brachionus plicatilis and anostraca crustacean Artemia sp. were analyzed for fatty acid contents, and were used as live food for culturing larval Japanese flounder, Paralichthys olivaceus. The total content of three types of HUFAs (DHA, EPA and ARA) in S. poplesia was significantly higher than that in the other two live foods (P<0.01). Three live organisms were used for raising larvae and juveniles of Paralichthys olivaceus respectively for 15 and 10 d. Then the growth, survival and fatty acid composition of the larvae and juveniles were investigated. The results showed that the larvae and juveniles fed with copepods (S. poplesia) had significantly higher growth rate than those fed with the other two organisms (P<0.01). The survival of the flounder larvae fed with copepods was significantly higher than that of the others (P<0.01), and the survival of the juvenile fish fed with copepods was higher than that fed with Artemia (P<0.05). The contents of three types of HUFAs (DHA, EPA and ARA) and the ratio of DHA/EPA in larval and juvenile flounder P. olivaceus were analyzed. The results showed that the contents of DHA, EPA and ARA in the larvae and juveniles fed with S. poplesia were higher than those fed with a mixed diet or Artemia only, and the ratio of EPA/ARA in larvae and juveniles of P. olivaceus fed with S. poplesia was lower than that in the case of feeding with a mixed diet or Artemia only. The present data showed that copepod is the best choice for feeding the larvae and juveniles of fish considering its effects on the survival, growth and nutrition composition of the fish.     

Otolith Microstructure Analysis Reveals Different Growth Histories of Japanese Sardine (Sardinops melanostictus) in the Oyashio Waters

After decades of low year classes,the stock of Japanese sardine(Sardinops melanostictus)has begun to recover since the mid-2000s.The hatch dates and otolith growth rates of age-0 juvenile sardine,which were collected in the subarctic Oyashio waters in autumn 2018,were determined from an otolith microstructure analysis.The sardines were hatched from late January to late April,while mostly in February and March.The otolith growth rate increased continuously up to 60 d after hatching and thereafter de-creased.The revealed growth rate in a crucial growth period is faster than that reported for juvenile sardines collected in the 1990s,which is coincided with the recent recovery trend of the sardine stock.Two groups with different hatch dates,growth histories,and migration routes were identified using unsupervised random forest clustering analysis.They were considered inshore and offshore migration individuals in accordance with recent researches.In the offshore group,a high proportion of sardine juveniles hatched late and grew faster in the Kuroshio-Oyashio transitional waters,a finding consistent with the hypothesis of growth-rate-dependent re-cruitment.This finding on the population composition and growth rate of juvenile sardine in the Oyashio waters can be a basis for an improved prediction of their survival and provides us with valuable information on the recruitment processes of this stock during the period of stock recovery.     

Using otolith weight to predict the age of <Emphasis Type="Italic">Pennahia macrocephalus</Emphasis> in the mouth of the Beibu Gulf

The relation between otolith weight (OW) and the age of marine fish is studied. A total of 222 individuals of bighead white croaker, Pennahia macrocephalus were sampled seasonally in the mouth of the Beibu Gulf, the South China Sea, in 2007. Since there are no significant differences in sagittal OW between otolith in pairs (P≥0.05), the undamaged left sagittal otolith is used for age determination. The highest correlations among standard length, OW and fish ages are confirmed by linear, exponential and multinomial regression. Results show that sagittal OW overlaps only occasionally among age groups, and to individuals with similar standard length, the older and slower-growing fish has a heavier otolith because of the continued otolith material deposition. There are differences in sagittal OW among different age groups and significant positive linear relationship with age (P<0.05). The age readings can be verified by plotting the sagittal OW versus the standard length for age groups, and the individuals with similar standard length but in different ages can be separated by sagittal OW frequency analysis. Mostly, the predicted ages using the regression between sagittal OW and ages are closed to the observed ages by counting annulus on scale. It indicates that the sagittal OW analysis is a useful technique for validating the accuracy of age determination by annuli counts, especially for individuals of similar size. Furthermore, the technique is applied for Pennahia macrocephalus with discussion in this paper. Supported by the National Natural Science Foundation of China (No. 30771653) and Bureau of Fisheries, Ministry of Agriculture     

Embryonic and larval development in barfin flounder Verasper moseri (Jordan and Gilbert)

Broodstock of Verasper moseri (Jordan and Gilbert) aged 3–4 years old were selected, and reinforced cultivation was conducted to promote maturation under controlled water temperature and photoperiod conditions. Fertilized eggs were obtained by artificial fertilization, and the development of embryos, larvae and juveniles was observed continuously. The results showed that the fertilized eggs of V. moseri were spherical, with transparent yolk and homogeneous bioplasm, and had no oil globule inside. The average diameter of the eggs was 1.77±0.02 mm. The eggs of V. moseri were buoyant in water with salinity above 35. The cleavage type was typical discoidal. Young pigment cells appeared when olfactory plates began to form. Hatching occurred at 187 h after fertilization at a water temperature of 8.5°C. The newly hatched larvae, floating on the water surface, were transparent with an average total length of 4.69±0.15 mm. During the cultivation period, when the water temperature was raised from 9 to 14.5°C, 4-day old larvae showed more melanophores on the body surface, making the larvae gray in color. The pectoral fins began to develop, which enabled the larvae to swim horizontally and in a lively manner. On days 7–8, the digestive duct formed. The yolk sac was small and black. The yolk sac was absorbed on day 11. Larvae took food actively, and body length and body height clearly increased. The rudiments of dorsal and anal fin pterygiophores were discernible and caudal fin ray elements formed on day 19. On day 24, the larval notochord flexed upwards, and the rays of unpaired fins began to differentiate. Pigment cells converged on the dorsal and anal fin rays, and the mastoid teeth on the mandible appeared. On day 29, the left eyes of juveniles began to move upwards. Depigmentation began in some juveniles and they became sandy brown in color on day 37. Most juveniles began to settle on the bottom of the tank. The left eyes of juveniles migrated completely to the right side on day 50, when the average body length attained 2.5±0.18 cm, and juveniles accomplished metamorphosis to young. The embryonic and larval characters of several flounder species are compared.     

Use of calcein and alizarin red S for immersion marking of black rockfish Sebastes schlegelii juveniles

Black rockfi sh Sebastes schlegelii juveniles(30–40 mm total length) were immersed in a range of calcein(CAL) solutions at concentrations ranging from 50 to 250 mg/L and alizarin red S(ARS) solutions at concentrations ranging from 100 to 500 mg/L in fi ltered seawater(salinity 30) for 24 h. Fluorescent marks were detected in otoliths(sagittae, asteriscus), scales, fi n rays(dorsal, pectoral, ventral, anal, and caudal fi n rays), and fi n spines(dorsal, ventral, and anal fi n spines) after a 60-d growth experiment. With the exception of 50–100 mg/L CAL, acceptable marks were produced in the otoliths and fi n spines by all concentrations of CAL and ARS. In particular, marks were clearly visible under normal light in the sagittae, asteriscus, and fi n spines of fi sh immersed in 200–500 mg/L, 300–500 mg/L, and 200–500 mg/L ARS, respectively. Scales and fi n rays had acceptable marks at much higher concentrations(≥50 mg/L CAL, ≥300 mg/L ARS for scales and ≥50 mg/L CAL, ≥200 mg/L ARS for fi n rays). The mark quality was highest(i.e., acceptable marks were observed in all sampled structures after immersion marking) in fi sh immersed in 150–250 mg/L CAL or 300–500 mg/L ARS. In addition, there was no signifi cant difference in survival and growth of marked fi sh compared with controls 60 d post-marking( P 0.05).     

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.     

Use of Otolith Shape for the Identification of Trumpeter Sillago （Sillago maculata） and Silver Sillago （Sillago sihama）

Otolith shape is species-specific in fish. In this study, 161 samples of Trumpeter sillago Sillago maculata and 164 of Silver sillago Sillago sihama were collected from Beibu Gulf in July 2009. The main objective of this study was to use morphological variables and Fourier harmonics to describe otolith characteristics and use discriminant analyses to separate the two species. Otoliths were measured by traditional one and two dimensional measures(otolith length, width, diameters, area, perimeter, rectangularity and circularity), as well as by Fourier analysis to capture the finer regions of the otolith. Analysis of covariance(ANCOVA)showed that there was significant correlation between morphological variables(diameter, perimeter, otolith length, otolith width, otolith area, density, harmonics 3, harmonics 4, harmonics 5, harmonics 13, harmonics 15, and harmonics 16)and body length. To minimize size effects on the morphological variables between species, only fish with a body length between 90 and 140 mm were included in the data analysis and the variables which had significant relation with body length were transformed using the residual. The result showed that the accuracy of discriminant analysis was 97. 8% and 100% for Trumpeter sillago and Silver sillago, respectively. This high accuracy indicated that otolith shape was described accurately by morphological variables and Fourier harmonics, and discriminant analysis was an effective way to identify and separate the two species.     

The effects of temperature and substrate on ontogenetic behavior of bastard halibut, Paralichthys olivaceus （T.et S.）

The effects of temperature and substrate on ontogenetic patterns and settlement behavior of bastard halibut, Paralichthys olivaceus （T. et S.） were studied in a series of laboratory experiments. Analysis on stage-based data revealed that except for settlement, and compared to the substrate, the ambient temperature dominantly controlled the hatch success and subsequent development of the species. The oosperm optimum survival rate of 60% during hatching occurred at 20℃, and survival rate of 29%, at 18℃ for larval rearing. The survival rate of larvae reared under variable temperature of 3.5-4.5℃ was higher than that of those reared under stable temperature. The maximum survival rate of 29%, corresponded to variation at temperature of 4℃. With increase in age newly settled juveniles preferred to lie on the fine substrate and gradually moved on the coarse substrate. GLM （general linear model） analysis showed that the combination of temperature and substrate had no significant impact on the survival of settling larvae, but definitely affected the duration of settlement.     

尖吻鲈苗种工厂化培育模式的初步研究

采用室内水泥池及活饵培育配套技术进行尖吻鲈苗种阶段工厂化育苗。投放刚孵化出膜的仔鱼１６０万尾,经室内池８ｄ的培育,成活率为９０％;经１３ｄ的培育其规格超过１ｃｍ,成活率为５６．９％;经２４ｄ的培育平均规格达２ｃｍ左右,平均成活率为５０％,共育出８０万尾。苗种生长快,体质健壮,成活率高。关键技术是：基础设施配套;采用不同活饵,做到适口、及时转化、足量;定期过筛分级培育;水质控制良好;注意防病。２ｃｍ以上鱼种阶段采用多种方式培育,经１５～２０ｄ的饲养,规格达３～４ｃｍ,平均成活率为６５％～７５％。     

Development of retinal structure and visual acuity in Japanese flounder (Paralichthys olivaceus)

The retinal structure and visual acuity in Japanese flounderParalichthys olivaceus at different stages of development were examined by light microscopy. The resolving power of the retina, the visual axis and the best visual field were estimated based on the distribution of cone cells in the retina. The visual system of the larvae appears poorly developed at hatching. The larvae with total length (TL) of less than 10 mm, have single cones only and the eyes were well pigmented. At 10–11 mm TL, most single cones fused to form double cones, with the single and double cones forming a mosaic pattern. From larvae to early juvenile the retina stretches, the cones increase in diameter and rods increase in number. Based on the highest density of the cones in the ventro-temporal region, the visual axis was orientated upforward. The resolving power of the retina in 40–530 mm TL Japanese flounder was found to range from 25.1 to 11.5 min. The results indicated continual improvements in the visual system of the growing fish towards higher resolving power, visual acuity and sensitivity. Project 39970578 supported by the NSFC, and also supported by the Ministry of Education Foundation for University Key Teachers.     

Effects of rearing temperature and density on growth, survival and development of sea cucumber larvae, Apostichopus japonicus (Selenka)

In laboratory conditions, effects of rearing temperature and stocking density were examined on hatching of fertilized egg and growth of auricularia larvae of Apostichopus japonicus respectively. Data series like larval length and density, metamorphic time, and survival rate of the larvae were recorded. Statistics showed that for A. japonicus, survival rate (from fertilized egg to late auricularia) decreased significantly with the increasing rearing temperature (P<0.05). At different temperatures SGR was statistically significant as well (P<0.05) from day 1, and maximal SGR was found on day 9 at 24°C (159.26±3.28). This study clearly indicated that at low temperature (<24°C), metamorphic rate was remarkably higher than at higher temperature (>26°C). Hatching rate was significantly different between 0.2–5 ind./ml groups and 20–50 ind./ml groups. Rearing larvae at the higher density had the smaller maximal-length, whereas needed longer time to complete metamorphosis. This study suggested that 21°C and 0.4 ind./ml can be used as the most suitable rearing temperature and stocking density for large -scale artificial breeding of A. japonicus’s larvae.     

Length frequency analysis ofPleuragramma antarcticum,Electrona antarctica,Protomyctophum bolini

British Antarctic Survey (BAS) data collected near the Antarctic Peninsula area were used in this 1988/1989 study on the length frequency of three Antarctic fish species (Pleuragramma antarcticum, Electrona antarctica and Protomyctophum bolini) showing that the standard length (SL) ofP. Antarcticum ranged from 26.0 mm to 161.0 mm, that the 26.0 mm to 54.0 mm SL juveniles could be considered to be the age group 1 fish, and that the SL ofE. antarctica ranged from 39.4 mm to 89.7 mm for the males and 48.0 mm to 118.0 mm for the females, which may contain at least 6 age groups. This is more than previously thought.P. Bolini had 29.2 mm to 60.2 mm SL and complicated age composition.     

Using otolith weight to predict the age of Pennahia macrocephalus in the mouth of the Beibu Gulf

The relation between otolith weight (OW) and the age of marine fish is studied. A total of 222 individuals of bighead white croaker, Pennahia macrocephalus were sampled seasonally in the mouth of the Beibu Gulf, the South China Sea, in 2007. Since there are no significant differences in sagittal OW between otolith in pairs (P≧>0.05), the undamaged left sagittal otolith is used for age determination. The highest correlations among standard length, OW and fish ages are confirmned by linear, exponential and multinomial regression. Results show that sagittal OW overlaps only occasionally among age groups, and to individuals with similar standard length, the older and slower-growing fish has a heavier otolith because of the continued otolith material deposition. There are differences in sagittal OW among different age groups and significant positive linear relationship with age (P<0.05). The age readings can be verified by plotting the sagittal OW versus the standard length for age groups, and the individuals with similar standard length but in different ages can be separated by sagittal OW frequency analysis. Mostly, the predicted ages using the regression between sagittai OW and ages are closed to the observed ages by counting annulus on scale. It indicates that the sagittal OW analysis is a useful technique for validating the accuracy of age determination by annuli counts, especially for individuals of similar size. Furthermore,the technique is applied for Pennahia macrocephalus with discussion in this paper.     

A light and transmission electron microscopic study ofHexamite capsularis sp. nov. (Diplomonadina: Hexamitidae) in fish (Xenocypris dividi)

A diplomonad flagellateHexamita capsularis sp. nov. from the posterior intestine of the fish (Xenocypris dividi) from Liangzi Lake, Hubei Province, is described. The flagellate was oval, with smooth surface and homogenous cytoplasm, 6.21±0.28 μm in length, 3.43±0.48 μm in width and with an elliptical capsule. Two recurrent flagellae arranged between two nuclei ran longitudinally to the posterior end. Fibrillar bands connected the accessory organelles of two cytostomal tubes, the body depressed at the posterior end of the body. This work is a part of the M. S. thesis of Xiao Wu-han and was supported by the National Natural Science Foundation of China (Grant No. 3870489).     

A successful microbound diet for the larval culture of Chinese shrimp Fenneropenaeus chinensis

A 13-day feeding trial was carried out to evaluate the effectiveness of a microbound diet for rearing the larvae of Chinese shrimp Fenneropenaeus chinensis in comparison with the live foods that consist of Isochrysis galbana, Chlorella vulgaris, Tetraselmis chuii, rotifer (Brachionus plicatilis) and Artemia sp. Larvae of 0 to 13d post-hatch (dph) were reared in a temperature-controlled semi-open culture system and stocked at a density of 100 larvae L⁻¹ in tanks, each containing 50 L sterilized seawater with salinity 30–32. Larvae were manually fed either the live foods or the microbound diet 6 times per day. At 13dph, the growth of the larvae fed on the microbound diet was approximately 84% of that fed on the live foods. The survival rate of the larvae fed on the microbound diet was 44.29% at 13 dph, which was not significantly different from that of larvae fed on live foods (63.55% ). The body length and development index (DI) of the larvae fed on the microbound diet were always lower than those of larvae fed on live foods. However, the differences reached significant levels only at 11 and 13dph (P<0.05). The mean dry weight loss of the microbound diet was 9.2% after 90min immersion in seawater, indicating that this diet has a good water stability. The microbound diet contains 5223% crude protein and 10.27% lipid and is easy to prepare. These characteristics of the diet suggest good potentials for its successful use in the larviculture of other penaeid and fish species.     

Effects of base analogues 5-bromouracil and 6-aminopurine on development of zebrafish Danio Rerio

Zebrafish (Danio rerio) genetic screens allow isolation of a wide array of problems in vertebrate biology. The effects of base analogues 5-bromouracil and 6-aminopurine on the development of zebrafish embryos are reported for the first time in this study. The early development of the zebrafish embryos was little affected by 5-bromouracil and 6-aminopurine, while the late development (organogenesis) was significantly impaired. Embryos exposed to 5-bromouracil mainly showed curled tail, wavy body, golden pigmentation and the mouth with protruding lower jaw. 6-aminopurine-treated embryos had defective anterior structures, curled tails and wavy body. RAPD analysis showed that the majority of 5-bromouracil- and 6-aminopurine-treated larvae and fish shared banding patterns in common with the control, suggesting that most mutagenesis induced by these agents are point mutations. However, some fish derived from 5-bromouracil-treated embryos had golden (gol) pigmentation; and RAPD analysis revealed that their band patterns differed from those of the control. Possibly, 5-bromouracil can occasionally cause relatively extensive changes in the fish genome. The results of this study may provide valuable help for detailed studies of mutagenesis. This work was supported by Ministry of Science and Technology (MOST) of China (G 1999012005).     

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.     

Effects of salinity on embryonic development,survival, and growth of Crassostrea hongkongensis

This study examined the effects of salinity on embryonic development, survival, and growth of the Hong Kong oyster Crassostrea hongkongensis. The embryos, larvae, and juveniles of C. hongkongensis were held in artificial seawater at three different salinities (low, 15; medium, 23; and high, 30) to determine the optimum hatchery and nursery conditions for mass production of the seeds. Results showed that the percentage production of straight-hinged larvae from fertilized eggs was significantly lower at the high salinity than at the low- and medium-salinities (P < 0.05). The survival rates of larvae and juveniles differed significantly among the three salinity trials, with the highest survival rate observed at the low salinity (P < 0.05). The shell height of larvae was significantly larger at the low salinity than at the high and medium salinities from days 9 to 15 (P < 0.05), whereas that of juveniles was significantly larger at the low salinity than at the high and medium salinities on day 70 (P < 0.05). These results indicate that the larvae and juveniles of C. hongkongensis are tolerant to a wide range of salinities (15 to 30), but show better growth and survival at relatively low salinities. Thus, it is recommended to use relatively low salinities in hatchery and nursery systems for improved yields of C. hongkongensis.     

方斑东风螺不同批次卵质量的比较

2005年5月~8月在雷州市覃斗镇海威水产种苗基地的室内水池进行方斑东风螺亲螺培育,对亲螺不同批次产出的卵囊、卵、及其幼虫进行比较。结果表明:随着亲螺产卵批次的增加,亲螺产卵囊数、卵囊含卵数、产卵量逐次减少;卵囊长、宽逐次变小,厚逐次增加,形状由葵花子片状的比例大变为棒状的比例大;幼虫浮游期逐次延长;变态期幼虫壳长逐次变小;各批次之间的变态率逐次降低;卵径、孵化率、出囊幼虫大小差异不显著。