Abstract:Thirteen microsatellite markers were used to analyze the genetic structure and genetic diversity of the breeding populations “Guanjingyang Youkuai 01” from F₁ to F₄. The results showed that the genetic diversity of the breeding population was decreasing, the average polymorphism information content (PIC) of the 13 microsatellite markers decreased from 0.638 to 0.524, the average allele number from 5.462 to 4.308, the average heterozygosity from 0.779 to 0.532, and the average Shannon’s gene diversity index from 1.356 to 1.092, respectively. The genetic identity between F₁ and its descendant generations (F₂, F₃, F₄) decreased (from 0.719 4 to 0.581 3) while the genetic distance increased. The genetic identity between the adjacent generations increased and the F_ST values decreased (0.061 9 in F₁-F₂; 0.051 1 in F₂-F₃; 0.047 5 in F₃-F₄). The allele frequency of the loci LYC0002 and LYC0054 changed regularly in the four breeding generations and they might correlate with the selected traits, which should be proved by further research. Our study suggested that the selective breeding work was efficient, some adverse genes were phased out, the hereditary basis of the population was getting pure and the genetic structure would be stable with the continuation of the breeding work.

Abstract:Infectious pancreatic necrosis (IPN) virus, the etiologic agent of infectious pancreatic necrosis in salmonid fish, causes significant losses to the aquaculture industry. The gene for the viral inner capsid protein (VP3) was amplified by RTPCR method from IPNV, and cloned into pET30b vector. The expression of recombinant plasmid pET30bVP3 in E.coli BL21(DE3) was induced and detected by SDSPAGE analysis. The predicted molecular weight for unmodified rtrunc VP3 was approximately 30 ku and this was found to be the case for E.coli protein. The amount of expression made up 30 percent of the bacteria protein total expression by thin layer scanning analysis. The results showed that the VP3 gene of IPNV can express successfully in E.coli BL21. The fusion protein was purified with ProBond^TM resin from the suspension centrifuged and the antisera against VP3 protein was produced. The pET30bVP3 fusion protein can be recognized by the positive serum of IPNV by Western-blotting analysis. The prepared antisera reacted specifically with IPNV antigen by indirect ELISA. The antisera against VP3 protein had OD values at least twice that obtained for the negative control serum at a dilution of 1∶25 600. The results showed that the expressed VP3 protein was immunogenical and antigenical which is the same as the natural IPNV VP3 protein. In this experiment the IPNV VP3 protein was expressed successfully by using prokaryotic expression system. The expressed fusion protein was active and the antisera against VP3 protein were produced.

Abstract:Japanese flounder (Paralichthys olivaceus) is a commercially important marine finfish in China. The growth rate of Japanese flounder is significantly faster in females than in males. Effective methods for gynogenetic inducement in this species are important for allfemale population construction. Meiogynogenesis was induced in Japanese flounder （P.olivaceus） by using UV irradiated sperm of red sea bream （Pagrus major）. The sperm of red sea bream exhibited a significant Hertwig effect after irradiating by UV. The embryo had the lowest hatching rate (2.0%±0.1％) when UV dosage was 3.4 mJ/cm², and recovered with the increase of UV dosage. It reached the highest hatching rate (57.7%±3.1％) when UV dosage was 73 mJ/cm², and ploidy analysis by flow cytometer showed that the embryos were all haploids. After a series of tests for initiation and duration at water temperature of （0±0.5） ℃, the best parameters for meiogynogenesis inducing in Japanese flounder were determined that cold shock begun at 3 min after insemination and continued for 45 min.The fertilization rate and hatching rate at this scenario were 80.7%±1.1% and 57.3%±2.3％,respectively, which were significantly higher than all the other treatment groups(P＜0.05). Fries of gynogenesis were confirmed to be diploids via flow cytometer. Further analysis of RAPD analysis showed that genetic material of red sea bream sperm was destroyed after UV irradiation and was not transmitted to progenies. This study provided an effective method to induce meiogynogenesis for all female population construction in Japanese flounder.

Abstract:Singapore grouper iridovirus (SGIV), as a causative agent of serious systemic disease, resulted in significant economic losses in grouper aquaculture. In this study, recombinant eukaryotic vector pEGFP-ICP46 which was inserted with SGIV-ICP46 (infected cell polypeptides 46) gene was transfected into fathead minnow (FHM) cells, and ICP46-GFP fusion protein was successfully expressed in the cytoplasm of FHM cells. Candidate siRNA targeting SGIV-ICP46 gene (siRNA-ICP46) was designed and chemically synthesized. To investigate the inhibition effect of siRNAICP46, pEGFPICP46 and siRNA was co-transfected into FHM cells, and the green fluorescence was observed by fluorescence microscope at the indicated time points (24, 36, 48, 60 and 72 h post transfection). During 24-48 h after transfection, the green fluorescence in FHM cells co-transfected with siRNAICP46/pEGFPICP46 were similar to that of positive control (co-transfected with siRNAGFP/pEGFPICP46) , and both of them are about 70% weaker than that of negative control (co-transfected with siRNANegative / pEGFP-ICP46), which show the siRNAICP46 can effectively inhibit the extrinsic SGIV ICP46 gene in FHM cells during 24-48 h after transfection.

Abstract:The mitochondrial DNA control region (D-loop) of Siniperca chuatsi was amplified by PCR using a pair of specific primers. The sequences with the complete nucleotide control region from S.chuatsi mitochondrial was cloned and directly sequenced. The length of this region (Dloop) contained 833 bp nucleotides and the T, C, A and G contents were 30.4%, 20.4%, 33.7%and 15.5% respectively. The genetic diversity of S.chuatsi from the Nanhai, Guangdong; Huizhou, Guangdong; Nanchang, Jiangxi and Poyang Lake (Jiangxi) was analyzed. The results indicated, compared to Jiangxi S.chuatsi species, S.chuatsi species in Guangdong have a problem that the quality of species has low polymorphism. The polymorphism analysis has evaluated S.chuatsi’s heredity multiple levels, provides the scientific basis for S.chuatsi’s idioplasm protection, provides the genetics background material for development and utilization of S.chuatsi’s resources, in order to formulate corresponding resources conservation measures. Meanwhile it is significantly meaningful that the intensification of China’s main production area Guangdong Province’s S.chuatsi industry should be promoted to highly effective, highquality direction with sustainable healthy development. 

Abstract:The pathogen Virio harveyi was isolated from the focus of illed Pseudosciaena crocea was infected back to the healthy P．crocea artificially． The challenged fish indicated the typical ulcered syndrome． The blood chemistry parameters were tested．The challenged fish indicated significantly（P<0.01）lower percentages of monocytes,polymorphonuclear neutrophil granulocytes(PMN) and thrombocytes but higher percentages of leucocytes; it differed from the control fish. Significant decreases in total protein, globulin, triacyl glycerol and blood glucoe concentrations were found in the challenged fish compared to the control fish（P<0.01）,with the total protein values at （12.8 ±3.95） g/L and （19.4±4.33） g/L, respectively. Significant increases of alanine aminotransferase（ALT）and aspartate aminotransferase（AST）concentrations were found in the challenged fish compared to the healthy fish（P<0.01）. The concentrations of ALT and AST were （37±5.11） IU/L and （164±47.19） IU/L in the challenged fish, while in the control fish were （28±5.54） IU/L and （117±21.84） IU/L. The results indicated liver and kidney damage， and disturbances in hepatic and osmoregulatory function of P.crocea.Observations by histopathology showed that head kidney and liver of the tested P.crocea were severely damaged and various denaturalization and necrosis. Denaturalization and necrosis emerged in the epithelial cell of renal tubule under electronic microscope. Fatty were accumulated in the cytoplasm of hepatic cells. The serum chemistry parameters determinations was a good inicator of bacterial infection of the challenged fish.

Abstract:Abstract: A total of 7 half-sib families and 21 full-sib families (A1、A2、A3……G1、G2、G3) of Sunray surf clam Mactra chinensis Philipi were established by using the method of unbalanced nest design in 2007, 9. Phenotypic traits of different families, such as egg-diameter, fertilization rates, hatching rates, and larval and juvenile growth, survival and metamorphosis were analyzed. The results showed that egg-diameter, fertilization rates, hatching rates, and D larval size were non-significant difference among families (P＞0.05). Considering growth, growth was varied at different development stages for each family and was influenced on maternal effect, paternal effect and male within female effect. G1 had the largest shell length with the growth rate 10.04±1.67 μm•d-1; whereas A2 had the smallest shell length with 6.07±1.32 μm•d-1during pelagic period of larvae. B3 grew fastest in filial generation of family with 29.93±3.67 μm•d-1; whereas D1 grew slowest in filial generation of family with 16.72±2.73 μm•d-1 during plantigrade period. G3 had the largest shell length with the growth rate83.14±5.85μm•d-1; whereas A2, A3 of juvenile indoor period had the smallest shell length with growth rate 57.78±5.44μm•d-1, 55.86±4.48μm•d-1 respectively. Considering survival, A2, C1, E3, F2, G1 and G2 had higher survival rate (＞85%) in filial generation of family, whereas B2、D1 had lower survival rate(＜35%) in filial generation of family during pelagic period of larvae. G1 had the highest survival rate (70.40%) in filial generation of family, whereas A1(23.40%), C2 (20.90%), F1(19.30%) had lower survival rate in filial generation of family during juvenile indoor period. Considering metamorphosis, growth rate was varied for each family, which the growth rate of G1and G3 were 7.94±1.53 μm•d-1, 7.96±1.52 μm•d-1 respectively, but the growth of D1 was only 5.23±0.75μm•d-1. B1, E1 and G2 had higher metamorphosis (＞80%) in filial generation of family, whereas E2(41.24%)、F1(43.25%) had lowest metamorphosis in filial generation of family. However, as the metamorphic time prolonged, the metamorphic size decreased. G1、G2、G3 had the largest metamorphic size (240μm) with the shortest metamorphic time (13d), whereas D1 had the lowest metamorphic size (226.65±5.65μm) with the longest metamorphic time (17d). By summarizing the early phenotypic traits of families, G3, F2 and G1 respectively inherited the excellent traits such as fast growth, strong stress resistance, and high yield, and were further used as the breeding material for rearing of new sunray surf clam strains.

Abstract:Spinibarbus sinensis Bleeker in the Yangtze River and its tributaries is of great economical value. However， it suffers from serious diseases， particularly a bacterial disease named “muscle ulceration” in intensive aquaculture. In order to effectively control the disease of S.sinensis， it is necessary to clarify the changes of immune parameters of periphery blood in S.sinensis immunized with a kind of dead bacteria. One hundred and eighty healthy fish (S.sinensis) with an average body weight of (100±25) g were divided randomly into immunized and control groups. Each individual was intraperitoneally injected with 0.2 mL 1.0×10⁸ mL formalinkilled Aeromonas sobria in immunized group，and with 0.2 mL 0.65% sterile saline solution in control group. Then，approximate 2 mL of blood was taken from each fish caudal vein on Day 0，1，2，4，7，14，21，28 and 35 post injection to study the changes of immune parameters，such as haematocyte number，differential leucocyte count(DLC)，serum antibody titer，phagocytic activity of phagocytes and serum protein content. The results were as follows：the number of erythrocytes and leucocytes in peripheral blood of immunized S.sinensis increased during the first 4 days post injection and peaked on 4th day，then，gradually decreased to the control level(Fig.1a & 1b). The differential leucocyte count(DLC) of the immunized group increased and was significantly(or very significantly) higher than that of control group for monocytes and neutrophils from day 4 to day 7(Fig.2a & 2b)，and for lymphocytes from the 14th day to 28th day(Fig.2c)，but lower than that of control group for thrombocytes on day 4 and 21(Fig.2d)，and was the same level as that of control group for the four kinds of cells at the other sampling times. The phagocytic percentage(PP) and phagocitic index(PI) of phagocytes in immunized S.sinensis were significantly higher than those of the nonimmunized fish between day 2 and day 7，and of the maximum value on day 4，but were not significantly different from those of the nonimmunized S.sinensis at the other sampling times(Fig.3a & 3b). The serum antibody titers gradually increased in immunized S.sinensis during the first 21 days post injection and reached the greatest value on day 21，then，gradually dropped off，but always kept the low level of 1∶4-1∶5 in the nonimmunized fish(Tab.1). The serum globulin content gradually increased in the immunized fish from day 4 to day 21， peaked and significantly higher than that of the nonimmunized fish on day 21(Tab.2). The serum globulin changes resulted in the same change model of serum total protein，which was not of significant difference between immunized and control groups(Tab. 2). However，the serum albumin quantity was not significantly different between the immunized and control groups，and fluctuated a little in the two groups(Tab.2). Relative percent survival was 65.21% for the immunized S.sinensis after challenge by means of injection with 0.2 mL live A.sobria at a concentration of 5.0×107 mL for every individual(Tab.3). This indicated that the formalinkilled Aeromonas sobria(F_AS) stimulated S.sinensis to produce a significantly protective immunity against the pathogen bacteria infection. All the results stated above suggested that the vaccine(F_AS) made S.sinensis enhance the nonspecific cell immunity，such as monocytes，neutrophils and erythrocytes，during the early period(from day 1 to day 7)，and the specific cellular and humoral immunity，such as lymphocytes and antibodies，during the later period against the bacterial pathogen infection post single dose immunization，and the relative percent survival reached 65.21% for the immunized S.sinensis after challenge with live A.sobria.

Abstract:The formation, release and germination of tetraspores, together with the morphogenesis of gametophyte of Kapphycus alvarezii were investigated in Lian Bay, Hainan Province, China from November, 2007 to April 2009. The results showed the brown Kappaphycus alvarezii, originally gained from Phillippines in 1985, were tetrasporophytes and the tetraspores were mainly formed and released from April to October. Both tetraspore germination and embryogenesis could be induced artificially in the laboratory and great variations of embryos in color from brown red, yellowgreen, olivine green to olivinebrown chimera were observed. Rhizoids were formed and then disappeared during the embryogenesis period. The germination rate of tetraspores was about 87.1%±7.2% and the mean specific growth rate of embryos was about 6.3%±1.1 %/d for indoor dish culture. After 160 days indoor cultivation, the germinated tetraspores developed to young gametophytes. Then, 20 young gametophytes were cultured on the soften raft in the sea. The average specific growth rate measured was over 10%/d in the first month culture in the sea, and the highest specific growth rate for young gametophytes was over 21.2 %/d. However, the specific growth rate gradually reduced accompanied with the frond increase in size in the following cultivation. Four types of gametophytes with different morphology, density of branches and growth rate were obtained as follows: TypeⅠ shows thick buds, cylindershaped trunk, taper terminal, and sparse branches. Type Ⅱ shows thin buds, cylindershaped trunk, blunt terminal, sparse branches. Type Ⅲ shows thin buds, cylindershaped trunk, taper terminal, and dense branches. Type Ⅳ shows thin buds, cylindershaped trunk, oblate terminal stem, taper terminal, and dense branches. During our experimental period, a large scale of iceice disease broke out, and the vitality and resistance to the iceice disease in some strains of the gametophytes improved significantly compared to their parent plants.

Abstract:Vibrio harveyi is one of the most serious marine pathogen that can infect a number of aquaculture species. It has caused severe losses to aquaculture industries worldwide. Attempts to control the infection are hampered by lack of effective vaccines and rapid diagnostic kits, the formulation of which could be facilitated by the identification of immunogenic proteins. In this study, an immunoproteomebased approach was developed to identify candidate antigens of Vibrio harveyi for vaccine development. A 2DE map has been constructed for Vibrio harveyi, in the pI range of 4.0 to 7.0. Strain HY99 was grown in TSA medium for 18 hours at 28 ℃. Total soluble proteins were extracted using lysis buffer and purified with a 2D cleanup kit. Protein concentrations were determined by 2D Quant Kit, and the proteins were separated by 2DE under immobilized pH gradients (IPG). The 2DE map was obtained from 3 gels run with 7 cm immobilized pH gradient strips and 12.5% SDS-PAGE gels. The electrophoregrams were obtained by coomassie brilliant blue staining. 2DE gels were scanned with Image Master 2D Platinum analyzed by 300 dpi 2DE image analysis revealed (429±18) protein spots. Then Vibrio harveyi HY99 antisera was analyzed for reactivity by Westernblotting against Vibrio harveyi total soluble proteins separated by 2DE. The 3 maps analyzed revealed 45 pair protein spots by ImageMaster 2D Platinum. 15 spots are nonspecificimmunoreactive proteins of Vibrio harveyi, and 30 spots are specificimmunoreactive proteins of Vibrio harveyi. These 30 spots were chosen for mass spectrometry identification ,and 29 spots were successfully matched with the proteins of NCBInr database（http://www.matrixscience.com）. Two isoforms of formate acetyltransferase were proposed. The 30 spots from the 2DE map corresponded to 28 proteins. None of these identified proteins have previously been reported as immunogenic in Vibrio harveyi. 6 proteins are known from other bacterial immunoproteomic analyses. They may be considered to be crossreactive antigens from other bacterial infections. OmpN were identified a number of times during the immunoproteome analysis of other bacteria, such as Shigella flexneri, Pasteurella multocida, Escherichia coli. OmpW is one of the major outer membrane proteins of Vibrio alginolyticus, and it is an immunoprotein in the report. OmpU is an important virulence factor involved in the adherence of Vibrio vulnificus to the host cells. alanine dehydrogenase, Elongation factor Ts (EFTs), cysteine synthase were recognized by anti-sera of Staphylococcus epidermidis. To the best of our knowledge, there are no reports about the immunogenicity of the other remaining 18 identified proteins. Their role in immunoreaction is not fully understood. It is suggested that this study may be valuable for the immunoproteomics research on Vibrio harveyi. These immunoreactive proteins could be novel candidates for vaccine development. Future studies will evaluate the protection of the 28 proteins by a nasal immunization and challenge.

Abstract:According to the samples of Dosidicus gigas collected by Chinese squid jigging vessels during the investigation from 2007 to 2008 off the coast of Chile, the ratio of statolith maximum width (MW) to total length (TSL) as a indicator of statolith growth, the statolith growth impacted by sex maturity and individual size is analyzed by the analysis of variance. The results indicated that the ratio MW to TSL (MW/TSL) for female individual is closely related with gonad maturity, for the female squids with sexual maturity stage I to stage III, Stage II to stage III, here was a significant difference between their MW/TSL (P <0.05). However, for male squid there has no obviously relationship between MW/TSL and gonad maturity accordingly. The MW/TSLs for female squids with different mantle length (ML) group, group with ML 300-400mm to ML 500-600mm and Ml 600-700mm, group ML 400-500mm to ML 500- 600mm and ML 600-700 mm, exist significant difference (P <0.05). When the female individual grow from ML 400-500mm to ML 500-600mm, the MW/TSL increase rapidly and the difference was significant (P <0.01). For male squid, the MW/TSLs between ML 350-400mm and ML 400-500mm, ML 400-450mm and ML 450-500mm also exist significant differences (P <0.05), when the squid attain ML 400-450mm, the MW/TSL is the smallest. The studies have shown that sexual maturation and size of Dosidicus gigas have a obviously effect on its statolith growth, but the difference exist between male and female.

Abstract:In order to study the relationship between silver carp surimi gelling temperature and thermal stability of myosin, surimi gel properties treated with different gelling temperature, formation of myosin solution aggregates, α-helix structure change and thermal denaturation temperature of myosin solution were determined respectively. The results were as follows: The suitable gelling temperature of silver carp surimi was 40 ℃. The aggregation rate of myosin increased greatly at 39 ℃、51 ℃、54 ℃, and the maximum aggregation rate was 39 ℃. Myosin α-helix unwinded largely to random crimp structure at 40 ℃ and 54 ℃, and the maximum unwinding rate was 40℃. There are two myosin denaturation temperature of 43.32 ℃ and 51.59 ℃. Silver carp surimi gelling temperature was relative to the first unwinding temperature and denaturing peak temperature of myosin α-helix. The gelling temperature is essentially the first denaturing peak temperature of myosin.

Abstract:In the Yellow Sea, larger, higher trophic level, commercially important demersal species were gradually replaced by small, lower trophic level, pelagic, lessvaluable species. Such transition inevitably leads to increase prey on low trophic level species, especially zooplankton. Therefore, it is an important means to discuss control mechanism of biological production in the marine ecosystem and to assess predation pressure on zooplankton. At present, there is still lack of systematic research on combination production of fish community with zooplankton at domestic and overseas. This paper aims to study seasonal variations of the functional groups of fish community and their consumption of zooplankton in the Yellow Sea. The fish samples, which accounted for 90% of total biomass, were collected during five bottom trawl surveys in the Yellow Sea in September, December 2006 and March, May and August 2007 which covered the range of 120.50°-124.53°E，31.77°-36.55°N. Except silver pomfret Pampus argenteus, the other 18 kinds of fish, 344 2 stomach content samples were analyzed. The prey items were weighed to the nearest 0.001 g after removing the surface water, and identified to the lowest possible taxonomic level. According to stomach content analysis results, cluster analysis and 60% of BrayCurtis similarity level were used as criterion to divide functional groups of fish community. The results suggest that fish community in the Yellow Sea is divided into seven functional groups, i.e., planktivores, shrimp predators, shrimp/crab predators, shrimp/fish predators, benthivores, piscivores and generalist predators. Except early spring, the dominant functional group of fish community in other seasons is planktivores. The composition of functional groups in spring and summer is simple, but the composition in autumn and winter becomes complicated. By estimating food consumption of zooplankton in each month, fish community in the Yellow Sea feeds mainly on 11 kinds of zooplankton, including Euchaeta marina, Corycaeus sp., Calanus sinicus, Euphausia pacifica, Themisto gracilipes, Oxycephalus sp., Sagitta crassa, Acetes chinensis, Maeruran larva, Brachyura sp., and Squilla alima. The consumption of zooplankton prey is 218 735 tons in spring, and is the largest consumption is all seasons, and then gradually reduces in summer and autumn. Zooplankton consumption come to the least amount in winter, and the consumption of zooplankton prey picks up in early spring. The kinds and consumption of zooplankton prey of fish community in different seasons have remarkable difference. Only Euphausia pacifica appeared in the food of fish in each month and the other kinds of zooplankton prey are seasonal. It is obvious that krill is most important zooplankton and occupies a very important position in the Yellow Sea ecosystem. So, the resource situation of krill in the Yellow Sea directly affects competition degree among planktivores. This study provides basic information for going deep into study on effect of zooplankton on dynamics of fish community at high trophic level. It emphasizes combination study between demand of fish production for zooplankton with biomass of zooplankton, and discussing the interaction mechanism between them.

Abstract:In order to understand the nutritive quality and nutrient components in the muscle of Glossogobius giuris and to evaluate the potential of the species as a diet source for Chinese sturgeon that is an endangered species only found in the coastal China Seas and the Yangtze River, the main nutrient components in the muscle of G.giuris were analyzed in this paper. Samples ［body weight, (9.07±1.87) g; body length, (8.17±0.63) cm］ were collected in the coastal area of Zhejiang Province. Analysis methods of nutrient components were based on the standards of GB 5009-85 and BT/T 14965-1994. Evaluation standards of the nutritive quality were based on the FAO/WHO’s standard mode. The general nutrient compositions were determined with routine methods. Amino acid compositions were determined with amino acid analyzer. Fatty acid compositions were inspected with GC. The results showed that the content of moisture, crude protein, crude fat and crude ash in fresh muscle of G.giuris were 79.92%±0.96%,16.76%±0.30%, 0.91%±0.07% and 2.25%±0.14% respectively. Eighteen common amino acids were found in the muscle of G.giuris including 8 kinds of essential amino acids (Thr, Val, Met, Phe, Ile, Leu, Lys and Trp), 2 kinds of halfessential amino acid (His and Arg ), 8 kinds of nonessential amino acids (Asp, Glu, Ser, Gly, Ala ,Tyr, Cys and Pro). In dry sample, the total content of amino acids (TAA) was 78.70%±0.57%; the contents of EAA, HEAA and NEAA were 32.26%±0.55%, 7.55%±0.09% and 38.88%±0.34% respectively. Four kinds of delicious amino acids (DAA) accounted for 29.67%±0.30% in dry sample; the ratio of total delicious amino acids to total amino acids (W_DAA/W_TAA) was 37.70%. The essential amino acids index (EAAI) was 63.88, the ratio of total essential amino acids to total amino acids (W_EAA/W_TAA) was 40.99%, and the ratio of total essential amino acids to total nonessential amino acids (W_EAA/W_NEAA) was 82.96%. It was clear that the content of the different amino acids was stable and the constitutional rate of the essential amino acids met the Food and Agriculture Organization of the United Nations/Word Health Organization (FAO/WHO) standards. According to nutrition evaluation in amino acids score (AAS) and chemical score (CS), the first limited amino acid was Trp, the second limited amino acid was (Met+Cys). 5 saturated fatty acids (SFA), 5 monounsaturated fatty acids (MUFA) and 6 poly unsaturated fatty acids (PUFA) were found in the muscle of G.giuris. The contents of EPA and DHA in fatty acids were 3.91%±0.11% and 8.10%±0.60% respectively, which were much higher than those of most fish species. In conclusion, the study results indicated that the muscle of G.giuris contains plenty of PUFA, EPA and DHA, thus, it is a high quality food fish for human. It is also a potential diet for the juvenile Chinese sturgeon, Acipenser sinensis, when they stay in the Yangtze Estuary area.

Abstract:Feeding experiments were conducted with protein diets(PD)and proteinfree diets(FPD) for Hucho taimen（6.8-7.3 g）, the diets(PD) was prepared by casein and gelatin as protein source with high biological values. The whole body amino acid composition was determined at the beginning and the end of the study. The requirements of Hucho taimen for essential amino acids(EAA) were determined based on the daily increment and daily maintenance requirement of each EAA in fish body. Fish were raised in tanks. There were 2 diets, each diet was randomly assigned to triplicate groups of 25 fish. During the experiment, the water temperature fluctuated from 23 ℃ to 25 ℃, and the dissolved oxygen was 6.4 mg/L to 7.5 mg/L. The experiment was conducted for 28 days. Compared with the FPD fish, the survival rate, the feed coefficient rate, the weight gain and crude lipid were increased significantly(P<0.05).The daily maintenance requirement of EAA(except trytophan) accounted for 20% to 30% of the sum of daily increment and daily maintenance requirement, and accounting for 54.52% of tryptophan. So the daily maintenance requirement of EAA can not be ignored. The minimum requirements of EAA needed to satisfy the requirements of Hucho taimen［g/(100 g body weight)/d］ were: Threonine 0.040, Valine 0.041, Methionine 0.027, Isoleuline 0.034, Leucine 0.067, Phenylalanine 0.035, Lysine 0.068, Histidine 0.110, Arginine 0.050. When dietary protein level was 42％, the minimum contents of EAA in the dietary(g/kg diet) were as follows: Threonine 4.17, Valine 4.24, Methionine 2.84, Isoleuline 3.56, Leucine 6.99, Phenylalanine 3.64, Lysine 7.18, Histidine 11.53, Arginine 5.19 and Trytophan 0.76.

Abstract:A specific and sensitive chemiluminescent immunoassay (CLIA) was developed for quantification of a major vitellogenin (Vg) subtype (B-type Vg; VgB) in the serum of red lip mullet (Liza haematocheila). The mullet VgB CLIA was performed using twosite method, with subtypespecific antiserum developed for purified red lip mullet VgB (a-VgB). Assay conditions were optimized with regard to antibody concentration, as well as incubation time, resulting in the typical assay range from 3.91 to 500 ng/mL. Dilution of vitellogenic female mullet serum and the serum from estrogentreated juvenile mullet appeared to be parallel to purified mullet Vg, while dilution of male mullet serum hardly revealed any positive immunoreactivity in the developed CLIA. Serum levels of VgB were quantified in 10 individuals of red lip mullet reared in an aquaculture center in Tianjin City, China. In females (n=9), production of VgB was evident but varied in their serum, the levels were ranged from 3.0 to 2 700.1 μg/mL, showing a typical trend of increase during the ovarian growth. In contract, serum level of VgB in a matured male mullet was extremely low (2.7 μg/mL), indicating no sign of estrogenic activities in this environment. In addition, no trace of gonadal abnormality was evident when histological observation was performed for all individuals. The present study provided a new tool for the quantification of the major estrogeninducible biomarker (i.e., VgB) in red lip mullet, which appeared to be better in the sensitivity in comparison with our previous assay and thus enabled us to evaluate a variety level of estrogenic activities in aquatic environment. Basic information such as typical, albeit preliminary, reproductive changes in circulating VgB levels were also provided and will endow to set a “normal” baseline, which is necessary to be established prior to interpreting “abnormal” inductions of this biomarker.

Abstract:The promoting effect of nutrient addition on aquacultural wastewater purification of microorganism products, SYMCORE BZT^TM was investigated， and furthermore, the relationship between removal rates of chemical oxygen demand (COD), ammonia nitrogen (NH₄⁺-N) and nitrite nitrogen (NO₂^--N) and the ratio of C∶N∶P was studied. The experiments were composed of two parts. Part I consisted of one control group and two trial groups, promoting effect of nutrient addition on removal rate of microorganism was proved. Part II was made up of one control group and 15 trial groups to analyze the relationship between the mount of added nutrients and the removal rates of pollutants. Moreover, the relationship between the ratio of carbon, nitrogen and phosphorus and the removal rates of COD, NH₄⁺-N and NO₂^--N was calculated, and the mathematics models were set up. The results showed that the purification ability of microorganism products was significantly improved by adding suitable nutrients. The removal rates of COD, NH₄⁺-N and NO₂^--N of the treatment group increased 21.92%,34.43%,57.41% by adding glucose and 30.02%, 53.45%, 15.08% by adding phosphate compared with the control, respectively. At the same time, the amount of phosphate was found insufficient in control group. The removal rates for COD, NH₄⁺-N and NO₂^--N were the highest, 97.07%, 96.88% and 93.41%, when the ratio of C∶N∶P was 62.5∶4.85∶1，37.5∶4.51∶1 and 25∶2.97∶1, respectively. The relationship of glucose and phosphate concentration with COD, NH₄⁺-N and NO₂^--N removal rates could be described by quadratic equations, V=aX²+bX+c. The individual COD, NH₄⁺-N and NO₂^--N removal rates of the microorganism products are positively correlated with the concentration of glucose and phosphate in range of 0-40 mg/L and 0-1 600 mg/L, respectively. The process of microbial remediation could be featured as three sequential stages：Firstly, dissolved organic matter (DOM) was degraded; secondly ammonia was oxidized; and at last, nitrite was oxidized. From day 1 to day 7, DOM was directly utilized as the carbon and nitrogen sources of the heterotrophic bacteria．The removal rates of COD at this stage were significantly higher than those of the control. From day 7 to day 13, bacteria mainly utilized ammonium as nitrogen sources and produced nitrite. So the concentration of NH₄⁺-N reduced sharply, while NO₂^--N concentration increased suddenly. From day 13 to 19, NO₂^--N concentration decreased rapidly, because the nitrite was oxidized to nitrate. This paper supplied theoretical and technical foundation for the biopurification of aquaculture wastewater and the extending application of microbial products.

Abstract:Aiming at exploring the sequence differences of IGFⅠ gene of Nile tilapia (Oreochromis niloticus NewGifts，ON), blackchin tilapia (Sarotherodon melanotheron，SM) and their First Generation of Hybrid (HB), investigating the relationships between the great differences of saltresistant property among the three genotypes, we cloned partial cDNA fragments of insulinlike growth factor Ⅰ(IGF-Ⅰ) from gill extracts through rapid amplification of 3′ cDNA ends（3′RACE）methods. The results demonstrated to be the IGF-Ⅰb gene cDNAs with length of 1 076 bp,1 075 bp and 1 079 bp respectively and comprising of a stop codon, a 546 bp open reading frame (ORF) encoding a 182 amino acid peptides(aa), and 3′untranslated regions（3′UTR）. The deduced amino acid sequence of IGF-Ⅰb precursor is composed of signal peptide, mature peptide（including conserved B, C, A and D domains consisting of 29aa,10aa,21aa and 8aa respectively）and E domain of 70aa. Results of the predicted secondary structures were typed as “mixed patterns”. The results of multiple alignments showed highly homologous with 75.8% to 100% amino acid identity. The B and A domains are highly conserved in all of the allied species. There are 2aadeletion between No.82 and 83 sites in C domain, 3aa and 1aadeletion next to NO.131 and No.159 site, respectively. Ala / Pro substitution occurred at No.133 of E domain of blackchin tilapia compared with ON and HB, which is identical to Oreochromis mossambicus and Oreochromis urolepis hornorum. We presumed that the differences of saltresistant property of ON, SM and HB may be due to the alterative splicing of the IGF-Ⅰ E domain.

Abstract:In order to provide firsthand scientific data and to utilize domestic cultured puffer fish safely, four tissues(skin, muscle,liver and gonad) of four cultured pufferfish species (Takifugu rubripes, Takifugu fasciatus, Takifugu flavidus, Takifugu bimaculatus) collected from five coastal provinces(Liaoning, Hebei, Shandong, Jiangsu and Fujian) in China were monitored quantitatively by mouse method after grouping and concentration. The skin, muscle and liver groups of T.rubripes from Shandong Province and several ovaries were measured by enzymelinked immunosorbent assay(ELISA) at same time. The results indicated that the average TTX content in each group of skin, muscle and liver of the four species specimens were ‘nontoxic’ in both of the mouse assay (less than 0.5 μg/g) and ELISA (less than 0.8 μg/g), and all the testes have no detectable TTX . However, a moderately toxic ovary sample was still detected. The annual variation of the average TTX content in groups of the muscle, liver and skin of cultured pufferfish can be observed. The results of the mouse method and ELISA method have no significant difference. It is confirmed that the skin, muscle and testis of the collected cultured pufferfish can be used with safety. The livers of the four species of puffer fish could be safely consumed as a delicious and nutritive food. The market size of cultured pufferfish was small, but still toxic. It is necessary to prevent the edible parts contamination.

Abstract:A rapid, sensitive and simple method for white spot syndrome virus (WSSV) detection was established. Loopmediated isothermal amplification (LAMP) assay is a novel method of gene amplification with high specificity, sensitivity and rapidity, which can be applied for disease diagnosis in shrimp aquaculture. The method is performed under isothermal conditions with a set of four specially designed primers that recognize six distinct sequences of the target. In the present study, a set of four specific primers were designed based on the conservative sequence of envelope protein gene VP28 using Primer Explorer v4 software, and a rapid detection of WSSV was established by using LAMP assay. The parameters of reaction time and temperature were optimized, and its specificity and sensitivity were assessed. The reactions were carried out at 58,60,62,63,64,65,66,67,68 ℃ for different time (15 -75 min). A plasmid pMDWSSV carrying target sequence of LAMP detection was constructed. Tenfold diluted pMDWSSV (107-100 copies/μL) was used as template for LAMP assay to investigate the detection limit. To determine the specificity, LAMP assays were carried out with DNA templates from other pathogens (Infectious Hypodermal and Hematopoietic Necrosis Virus IHHNV, Taura Syndrome Virus TSV, A.hydrophila, V.alginolyticus, V. parahaemolytious, E.coli).The results showed the optimum LAMP assay for the rapid detection of WSSV was performed at 64 ℃ for 60 min. The LAMP assay had an unequivocal detection limit of 100 copies/μL, and it was 100 times lower than that of the nested PCR. The nucleic acids of other pathogens were not amplified by this LAMP system with the specific primers, which showed a good specificity. The resulting amplicons were detected using visual observation after the addition of SYBR Green I and gel electrophoresis. We investigated the efficacy of uracilNglycosylase (UNG) in preventing contamination in the LAMP assay procedure and explored its effect on the amplification efficiency. Products of LAMP with dUTP adding could be lysed by UNG. UNG could prevent LAMP products contamination effectively, but its effect on amplification efficiency was visible. The LAMP assay could be finished within an hour, requiring only a regular laboratory water bath or heat block for reaction and the result could easily be detected using visual observation. Clinical suspected WSSVinfected diseased shrimp samples were detected by both LAMP and PCR assay, which indicated LAMP could be more sensitive for WSSV detection.