Triploidy induction with heat shocks toPenaeus chinensis and their effects on gonad development

Heat shocks effectively produced triploids inPenaeus chinensis. Fertilized eggs heat shocked (28–32°C) for 8 to 16 minutes, starting from 8 to 20 minutes after spawning, resulted in triploidy induction rates of 39%–75%. Several triploid populations were cultured to 10 cm. In a triploid population, two kinds of ovaries were observed. Histological examination showed apparent differences between these two kinds of ovaries, whereas among male shrimp, there were no such differences. Contribution No. 3465 from the Institute of Oceanology. Chinese Academy of Sciences. The study was supported by the International Foundation for Science, Asian Fisheries Society and Chinese Climbing Project PDB 6–2, and Chinese National 863 Project (No. 863-819-01-05).     

Chromosome Behavior of Heat Shock Induced Triploid in Fenneropenaeus Chinensis

INTRODUCTIONTriploidanimalsareusefulbecausetheyaresterile ,characterizedbypositivegrowthinthere productiveseason ,usefulfortesting ,forintroductionofnon nativespecies ,andforprotectionofde velopedstrains.Theinductionoftriploidyhadbeenreportedinmanyaquaculturespecies .Triploidshavebeensuccessfullyinducedinshrimpusingtemperatureshockorchemicalshock (CB ,6 DMAP)in 4species :Sicyoniaingentis (Xiangetal.,1 991 ) ,Fenneropenaeuschinensis (Xiangetal.,1 992 ;Daietal.,1 993;Baoetal.,1 993;Li…     

Tetraploid induction by inhibiting mitosis I in scallopChlamys farreri

Tetraploid induction was carried out by inhibiting mitosis I in fertilized eggs ofChlamys farreri. Mitosis I was blocked with cold shock (5–7°C), Cytochalasin B (0.75 mg/L) and 6-dimethylaminopurine (6-DMAP) (60–75 mg/L) when 60% fertilized eggs released polar body II at 20°C. At 4-cells embryo stage, the ploidy was determined by counting chromosome number. In control groups, most embryos were diploids (72.22%) and aneuploids (24.78%). In Cytochalasin B, cold shock and 6-DMAP treated groups, tetraploids were respectively 10.51%, 4.08%, and 13.34%; aneuploids were 43.10%, 35.93% and 29.16%, and triploids were 7.84%, 8.52% and 18.33%. At D-larva stage, ploidy was determined by flow cytometry (FCM). The ploidy analysis of day 2 larvae showed diploids in control group and also in three treated groups. Juvenile scallops (0.2–0.3cm) which were harvested in two control groups and two CB treated groups were all diploids through checking ploidy individually by FCM. Contribution No. 3648 from the Institute of Oceanology, Chinese Academy of Sciences. Contribution No. 238 from the Experimental Marine Biology Laboratory, Chinese Academy of Sciences. Project funded by grant (No. 819-01-07) from the Chinese Science and Technology Ministry and also by “Hundred Talents Plan” grant from the Chinese Academy of Sciences.     

STUDY ON EMBRYONIC DEVELOPMENT AND EARLY GROWTH OF TRIPLOID AND GYNOGENETIC DIPLOID LEFI—EYED FLOUNDER, PARALICHTHYS OLIVACEUS （T. et S. ）

The early effects of chromosomal manipulation of eggs and sperm on the yields of trip-loid and gynogenefic diploid larvae of Paralichthys olivaceus were investigated. Triploidy was achieved by cold shocking fen.ilized eggs at 0 - 2℃ for 45 minutes duration 5 minutes after fen.ilization, and the in-duced triploidy rates were 31.2% - 50% and the relative hatching rates were 53.3% - 99%. Gynnge-aetic diploids were obtained when eggs were inseminated with irradiated sperm and cold shocked at 0 -2℃ for 45 minutes duration 5 minutes after fertihzation. The induced gynogenetic diploid rates and the relative hatching rates were 94 % - 96 % and 48.5 % - 68.5 % respectively. The embryonic development of the triploid experimental group and of the gynogenetic diploid experimental group was delayed at first compared with the control group. But from the gastrula stage, it was not delayed anymore. There were no significant differences in the growth of the triploid experimental group larvae and the control group larvae, and in the growth of the gynogenetic diploid experimental group larvae and the control group larvae ac-cording to Student‘s t-test (α = 0.05). The relationship between the early growth of the triploid experi-mental group larvae and that of gynogenetic diploid experimental group larvae was also studied.     

Use of rapd for detecting and identifyingPorphyra (Bangiales, Rhodophyta)

Randomly amplified polymorphic DNA (RAPD) technique was applied to assess the genetic variations and phylogenetic relationships in 4 species ofPorphyra. The samples were collected from the coast of Canada, Vietnam, Zhoushan, Fujian and Qingdao in China. Amplifications with 20 primers were carried out under predetermined optimal reaction conditions (samples were first heated at 94°C for 5 min. and followed by 45 cycles of 1 min at 94°C, 1 min at 36°C, and 2 min at 72°C, then held at 72°C for 10 min). The amplified products were scored as present (1) or absent (0) for each DNA sample and an index of genetic similarity (F) was calculated by using Nei & Li’s matching coefficient method (1979). The value of (1−F) was used to quantify the genetic distances between species and construct a phylogenetic tree. The relationship indicated by the UPGMA and NJ cluster analysis on the values of the genetic distance is in good overall agreement with classical taxonomy. The obvious differences between natural and cultivated population ofP. haitanensi suggest that variation or hybridization with other species occurred during the culture. Contribution No. 100 from the Experimental Marine Biology Laboratory, IOCAS. Contribution No. 3289 from the Institute of Oceanology, Chinese Academy of Sciences. The research supported by the Foundation of the Chinese Academy of Sciences.     

Properties of proteolytic toxin of Vibrio anguilolarum from diseased flounder

Extracellular products (ECP) produced byVibrio anguillarum strain M3 originally isolated from diseased flounder (Paralichthys olivaceus) were prepared. ECP of M3 showed gelatinase, casinase, amylase and haemolytic activity on agarose plates. High protease activity against azocasin was detected. Bacterium M2 showed highest growth and protease activity at 25°C. The protease present in ECP showed maximal activity at pH 8 and 55°C; was completely inactivated by application of 80°C heat for 30 min; was completely inhibited by EDTA and HgCl₂, and was partially inhibited by PMSF, SDS, MnCl₂ and iodoacetic acid; but not inhibited by CaCl₂ and MgCl₂. The ECP was toxic to flounder fish at LD₅₀ values of 3.1 μg protein/g body weight. The addition of HgCl₂ and application of heat at 50°C decreased the lethal toxicity of ECP. When heated at 100°C, ECP lethality to flounder was completely inhibited. After intramuscular injection of ECP into flounder, it showed evident histopathological changes including necrosis of muscle, extensive deposition of haemosiderin in the spleen, dilated blood vessels congested with numerious lymphocytes in the liver. These results showed that ECP protease was a lethal factor produced by the bacteriumV. anguillarum M3. Contribution No. 4213 from the Institute of Oceanology, Chinese Academy of Sciences. Funded by projects under the Major State Basic Research Development Program, Grant G1999012003.     

Competition Among Dinoflagellate Alexandrium Tam—arense, Raphidophyte Heterosigma Carterae and Diatom Skeletonema Costatum under Combinations ofTwo Temperatures and Five Salinities

Competition among HAB (Harmful Algal Bloom) species Dinoflagellate Alexandrium tamarense, Raphidophyte Heterosigma carterae, and Diatom Skeletonema costatum was studied in the laboratory. Experiments with these three major HAB species under combinations of different salinities (10, 18, 25, 30, 35) and temperatures (19℃, 25℃) were carried out. The results showed that S. costatum successfully competed with the other two species at salinities of 18, 25,30, and 35 at temperatures of 19℃ and 25℃. However, H. carterae showed its advantage at low salinity of 10 and became the single dominant species at salinity 10 and 25℃. A. tamarense could not compete successfully with the other two species especially at low salinities. However, it could remain at low density in the presence of higher densities of other algae.     

Triploidy induction in Pacific oysterCrassostrea gigas by caffeine with thermal shock

Zygotes of Pacific oyster (Crassostrea gigas) were treated for triploid induction with caffeine (10 mmol/L, 15mmol/L and 20mmol/L) in combination with thermal shocks (at 40 minute post-fertilization) lasting for 5 and 10 minutes. The highest yield of triploids, 41.5%, was obtained from the treatment with 20 mmol/L caffeine at 34°C lasting 10 minutes. The triploid levels were less than 30.0% in other treatments. Triploid induction was more effective in treatment with 15–20 mmol/L caffeine at 34–38 °C than with lower concentrations of caffeine at temperatures below 32 °C. Our results suggest that triploid induction with caffeine in combination with thermal shocks is less efficient than some other methods reported previously.     

Heat-shock protein 70 expression in shrimp Fenneropenaeus chinensis during thermal and immune-challenged stress

Using western immunoblotting we obtained heat-shock protein 70 (HSP70) induction data and distribution in different tissues from shrimpFenneropenaeus chinensis during thermal and immune-challenged stresses. This is probably the first report of the effects of various stressors on the expression of HSP70 in shrimp. HSP70 was prominently induced in hepatopancreas and gills, but not in muscle, eyestalk and hemolymph, when the shrimp were exposed to heat shock andVibrio anguillavium-challenged stresses. Cold shock and WSSV treatment had no significant effects on the levels of HSP70 expression in all tissues examined. HSP70 induction was greatest after 2 h exposure to heat shock stress, which was elevated after acute heat shock exposure of 10°C above ambient temperature. Supported by the National Key Fundamental Research Program “Study of the Major Diseases and Resistance Mechanism of Mariculture Organisms”, NSFC, No. 30140017, and the international cooperative program “Immunaqua” by the European Commission, No. ICA4-CT-2001-10023.     

STUDY ON MIXIS POTENTIAL OF ROTIFER RESTING EGGS（ BRACHIONUS PLICATILIS ） WITH DIFFERENT COLLECTION TIMES AND DIFFERENT PRESERVATION PERIODS

The present study investigated the possible changes in the mixis potential of rotifer resting eggs produced by a single stock of Brachionus plicatilis and collected and preserved annually from 1985 - 1998. Several clones derived from each batch of resting eggs were cultured under the same conditions for 21 days. The percentage of clones appearing resting eggs and the average yield of resting eggs produced from each clone were recorded and statistically analyzed to find the differences between the mixis potential of those resting egg batches. Results showed that different batches of resting eggs had different mictic levels among their descendent clones; but no regular relationship was found between the mixis potential of resting eggs and their collection times/preservation periods. Several internal and external factors that might affect the mixis potential of resting eggs were discussed.     

Effect of ration, temperature and body weight on growth of black porgy,Sparus macrocephalus (B.)

Growth of black porgy,Sparus macrocephalus, was studied experimentally at four ration levels from starvation to satiation and four temperatures ranging from 14.8 to 26.8°C. At maximum rations, the specific growth rate (SGR) increased with temperature and was a negative linear function of body weight. The relationship between SGR and ration levels was a decelerating curve. Stepwise regression was used to develop the predictive models for the specific growth rates. Maintenance rations and optimum rations both increased with increased temperature. The relationship between maintenance, optimum rations and body weight was influenced by temperature. Conversion efficiencies increased with ration from zero at maintenance ration to a peak at the optimum ration, then declined with further increased ration. Contribution No. 3127 from the Institute of Oceanology, Chinese Academy of Sciences. Project 39370542 supported by NSFC and study also supported by National Key Laboratory in the Institute of Hydrobiology, Chinese Academy of Sciences.     

Production of bioplastics and hydrogen gas by photosynthetic microorganisms

Experiments on salimity tolerance of embryos and larvae of the southern bay scallopArgopecten irradians concentricus Say under controlled temperature of 28°C showed that although the eggs were fertilized and cleaved at salinity 18.1–37.4, they could further develop into normal D-shaped larvae only at salinity 23.4–36.1, while the trochophore larvae could develop into normal D—shaped larvae at salinity 22.1–39.9. For 72 hours, the larvae could tolerate salinity of 18.4–43.9, the salinity favoring growth was 19.8–42.4; the suitable range was 21–36, and the optimum at about 28. They metamorphosed at salinity 18.9–43.5, optimally at 24–32. Contribution No. 3301 from the Institute of Oceanology, Chinese Academy of Sciences. The research was supported by the National Climbing Project B(PDB6-3-2).     

Effects of Body Weight and Water Temperature on Maximum Food Consumption of Juvenile Sebastodes fuscescens （Houttuyn）

Maximum rate of food consumption ( Cmax ) was determined for juvenile Sebastodes fuscescens (Houttuyn) at water temperature of 10, 15, 20 and 25℃. The relationships of Cmax to the body weight (W) at each temperature were described by a power equation: lnCmax = a b lnW. Covariance analysis revealed significant interaction of the temperature and body weight. The relationship of adjusted Cmax to water temperature (T) was described by a quadratic equation: Cmax = -0.369 0.456T - 0.0117T^2. The optimal feeding temperature calculated from this equation was 19.5 ℃. The coefficients of the multiple regression estimation relating Cmax to body weight (W) and water temperature (T) were given in the Table 2.     

Relation of body weight and food consumption to metabolic rate of juvenile Japanese sea bass,Lateolabrax japonicus

ＩＮＴＲＯＤＵＣＴＩＯＮＴｈｅＪａｐａｎｅｓｅｓｅａｂａｓｓ (ＬａｔｅｏｌａｂｒａｘｊａｐｏｎｉｃｕｓＣ＆Ｖ)ｄｉｓｔｒｉｂｕｔｅｓｗｉｄｅｌｙｉｎｔｈｅｃｏａｓｔａｌｗａｔｅｒｓａｌｏｎｇｔｈｅＢｏｈａｉＳｅａ ,ＹｅｌｌｏｗＳｅａ ,ＥａｓｔＣｈｉｎａＳｅａ,ａｎｄＳｏｕｔｈＣｈｉｎａＳｅａ .Ａｓａｃａｒｎｉｖｏｒｏｕｓｆｉｓｈ ,ｉｔｏｃｃｕｐｉｅｓｔｈｅｈｉｇｈｔｒｏｐｈｉｃｌｅｖｅｌｉｎｔｈｅｍａｒｉｎｅｅｃｏｓｙｓｔｅｍａｎｄｈａｓａｄａｐｔｅｄｔｏｖａｒｉｏｕｓｅｎｖｉｒｏｎｍｅｎｔａｌｒ…     

A NEW SPECIES OF THE GENUS NEROCILA （ISOPODA： CYMOTHOIDAE） FROM THE EAST CHINA SEA

A new species of cymothoid Isopeda, Nerocila donghaiensis, collected from the body of a sciaenid fish, Johnius sp., is described in the present paper. The new species can be distinguished from the other species of genus Nerocila by its endoped of uropods with deep notch, sharply serrated on the lateral margin and strongly serrated on the medial margin; antennule with 1 st article swollen; dactyli of pereopods Ⅰ, Ⅱ, Ⅳ, Ⅴ swollen.     

Effects of reduced salinity on oxygen consumption and ammonia-N excretion ofChlamys farreri

Effects of reduced salinity on the oxygen consumption rate (OCR) and the ammonia-N excretion rate (AER) of scallopChlamys farreri (3.2–5.9 cm in shell height, 0.147–1.635 g in soft tissue dry weight) were studied in laboratory from March 21, 1997 to April 16, 1997. Under the controlled conditions of reduced salinity from 31.5 to 15.0 and ambient temperature 17°C and 23°C, the concentrations of dissolved oxygen and ammonia-N were determined by the Winkle method and the hypobromite method, respectively. Results showed that with controlled reduced salinity, the mean values of the OCR were 2.17 mg/(g.h) at 17°C, and 2.86 mg/(g.h) at 23°C and that the mean values of the AER were 178.0 μg/(g.h) at 17°C and 147.0 μg/(g.h) at 23°C. The OCR and the AER decreased with reducing salinity from 31.5 to 15.0 both at 17°C and 23°C. The effects of reduced salinity on the OCR and the AER of scallopC. farreri could be represented by the allometric equation and the exponential equation, respectively. Contribution No. 3295 from the Institute of Oceanology, Chinese Academy of Sciences. Project supported by the National Ministry of Science and Technology of China and by the Chinese Academy of Sciences, Grant No. 96-922-02-04 and KZ951-A1-102-02.     

IMPACT OF STARVATION ON SURVIVAL，MEAT CONDITION AND METABOLISM OF CHLAMYS FARRERI

The effects of 60-day starvation on survival rate. condition index (CI) , changes ofnutrient composition of different tissues, respiration and excretion of scallop ChlartnCs farreri were studiedin laboratory from Oct. 17 to Dec. 15 ,1997. Two groups (control and starvation with 200 individualseach) were cultured in two 2 m3 tanks, with 31 to 32 salinity water at 1 7 “C . Starvation effects were mea-sured alter 1 0, 20, 40 and 60 days. There was no mass mortality of scallops cf the two tanks and survivalrates of the control and starvation groups were 93 .5 % and 92.0 %, respectively. Starvation had strongeffect on the meat condition of the scallops, especially after 10 days; when relative lipid percentagedropped sharply while relative protein percentage increased. The impact of starvation on the oxygen consumption mtc (OCR) and the ammonia-N cxcretion mtc (AER) was obvious.     

Reconstruction of vertical thermal structure from several subsurface temperatures in the China Seas and adjacent waters

Empirical Orthogonal Function (EOF) analysis is used in this study to generate main eigenvector fields of historical temperature for the China Seas (here referring to Chinese marine territories) and adjacent waters from 1930 to 2002 (510 143 profiles). A good temperature profile is reconstructed based on several subsurface in situ temperature observations and the thermocline was estimated using the model. The results show that: 1) For the study area, the former four principal components can explain 95% of the overall variance, and the vertical distribution of temperature is most stable using the in situ temperature observations near the surface. 2) The model verifications based on the observed CTD data from the East China Sea (ECS), South China Sea (SCS) and the areas around Taiwan Island show that the reconstructed profiles have high correlation with the observed ones with the confidence level >95%, especially to describe the characteristics of the thermocline well. The average errors between the reconstructed and observed profiles in these three areas are 0.69°C, 0.52°C and 1.18°C respectively. It also shows the model RMS error is less than or close to the climatological error. The statistical model can be used to well estimate the temperature profile vertical structure. 3) Comparing the thermocline characteristics between the reconstructed and observed profiles, the results in the ECS show that the average absolute errors are 1.5m, 1.4 m and 0.17°C/m, and the average relative errors are 24.7%, 8.9% and 22.6% for the upper, lower thermocline boundaries and the gradient, respectively. Although the relative errors are obvious, the absolute error is small. In the SCS, the average absolute errors are 4.1 m, 27.7 m and 0.007°C/m, and the average relative errors are 16.1%, 16.8% and 9.5% for the upper, lower thermocline boundaries and the gradient, respectively. The average relative errors are all <20%. Although the average absolute error of the lower thermocline boundary is considerable, but contrast to the spatial scale of average depth of the lower thermocline boundary (165 m), the average relative error is small (16.8%). Therefore the model can be used to well estimate the thermocline. Supported by the Knowledge Innovation Program of Chinese Academy of Sciences (No.KZCX-3W-222; KZCX2-YW-Q11-02) and National Basic Research Program of China (No.2007CB411802; 2006CB403601)     

Interannual variability of the southern Yellow Sea Cold Water Mass

Temperature data collected in the sections of 34°N, 35°N and 36°N in August from 1975 through 2003 were analyzed using Empirical Orthogonal Function (EOF) to investigate interannual variability of the southern Yellow Sea Cold Water Mass (YSCWM). The first mode (EOF1) reveals variations of basin-wide thermocline depth, which is mainly caused by surface heating. The second mode (EOF2) presents fluctuations of vertical circulation, resulting mainly from interannual variability of cold front intensity. In addition, it is found that the upward extent of upwelling in the cold front is basically determined by wind stress curl and the zonal position of the warm water center in the southern Yellow Sea is correlated with spatial difference of net heat flux.     

Artificial induction of mito-gynogenetic diploids in large yellow croaker (Pseudosciaena crocea) by hydrostatic pressure

The present study investigated conditions for inducing mito-gynogenetic (endomitosis) diploids by hydrostatic pressure in the large yellow croaker Pseudosciaena crocea. In haploid control groups, the development of eggs was activated with ultraviolet radiated semen. All fry presented typical haploid syndrome in the haploid control groups, and were verified as haploids using cytometry. After hydrostatic pressure treatment, morphologically normal fry reappeared at different frequencies according to the intensity and time of pressure shock. Fry with normal appearance in the pressure treated groups were verified as gynogenetic double haploids (GDHs), containing only one allele from the female parent at all four diagnostic microsatellite loci. For a fixed duration of 3 min, the optimal intensity of blocking the first mitosis was determined to be 40 Mpa, which was similar to that of blocking the second meiosis. There was a “window” of starting time, from 36.1 min to 38.1 min post-insemination at 25.0±1.0°C, within which the production of GDHs was not significantly different. Maximum production of morphologically normal fries, 9.36%±2.97% of developed eggs, was found when the eggs were shocked with hydrostatic pressure at 40 Mpa for 3 min, starting from 38.1 min post insemination at 25.0±1.0°C.