Abstract:With the layers of mucus and nonspecific immune defenses, the fish gill is exposed to the external environment and forms the initial barrier to invasio n by pathogens. As one of the mucosaassociated lymphoid tissues in teleost fishes, gill contains populations of leucocytes. In order to probe the immunerelated genes in red seabream (Pagrus major), the cDNA library of gill s was constructed from the red seabream inoculated with several species of pathogenic bacteria. The total RNA was extracted by TRIZOL and mRNA was isolated by biotinoligo (dT). Then, cDNA was synthesized from 5 μg mRNA. Through running gel electrophoresis of doublestrand cDNA, the fragments of 500-4000 bp were harvested, concentrated, ligated with λZAP vector and packaged in vitro. The primary cDNA library contained 1.75 ×10⁵ recombinants. The titer of amplified library was tested up to 1×10⁹ pfu·mL^-1, and the insert size of random picked phagemid was between 500-2000 bp. PCR with specific primers of hepcidin was performed to cloning of the homologous genes from cDNA library and a fragment of hepcidin cDNA sequence was amplified. The identity was above 85% in comparison with the hepcidin cDNA isolated from the liver of also red seabream. Therefore, the construction of cDNA library provides the basis for screening immunerelated genes from red seabream.

Abstract:13 different MHC class ⅡB alleles which encoded 13 different amino acid seq uences were identified among 411 clones from 42 resistant Japanese flounder individuals and 42 susceptible Japanese flounder individuals using the primer pairs of fmhcN1 and fmhcC1. Among the PCR products of 280 (268) nucleotides, 32 (11.4%) nucleotide positions were variable. Among the putative 61 amino acid sequence, 13 amino acid positions were polymorphic and 6 (46.15%) positions occurred in PBR region. Comparison of the ratio of nonsynonymous substitutions and synonymous substitutions (dN/dS) in the PBR (23.091) and in the NonPBR (0.523) indicated that the selection for amino acid replacement occurred mainly at the PBR region. Phylogenetic analysis using MEGA2.0 software showed that the putative 13 MHC class II B amino acid sequences of Japanese flounder assembled together as a cluster firstly. The phylogenetic relationship with the corresponding sequence of striped sea bass was relatively near, and the phylogenetic relationship with that of mammalian, such as with mouse and human, was relatively distant. With analysis of the occurrence percentages about the 13 different alleles in resistant and sus ceptible stock, most alleles such as Paolallele*aPaolallele*f, Paolallel e*j, Paolallele*k, Paolallele*i, Paolallele*m were commonly found in oth stocks. Using Chisquare Test, Paolallele*d was significantly more prevalent in individuals from susceptible stock than in individuals from resistant stock, and their percentages were 23.80% and 7.14%, respectively. In addition, the Paolallele*g and Paolallele*h only occurred in 13 individuals from 42 resistant individuals, and their percentages were 21.4% and 9.52%, respectively. While Paolallele*l was only found from 42 susceptible individuals, its percentage is 19.05%.

Abstract:Nibea albiflora is a commercially important marine fish species, which is widely distributed in Chinese, Japanese and Korean waters. Nibea albiflora has been intensively studied in China, including its basic biological characteristics and artificial reproduction. However, very few studies on N. albiflora were focused on genetic diversity, which has become an obstacle to further researches on the protection, management, rational exploitation and sustainable utilization of N. albiflora resources. The yield of N. albiflora has declined quickly due to increased efficiency of capture techniques and the leap of catch intensity. The information of genetic diversities of different populations will give us theoretical guidance in breeding and genetic improvement. In this study, a new molecular techonology, amplified fragment length polymorphism (AFLP) was used to analyze the genetic diversity and variation of two populations of N. albiflora from Qingdao and Xiamen sea areas. Procedures of AFLP analysis were essentially based on Vose et al. and Wang et al. A total of 47 individuals were studied using five primer combinations (E-AAC/M-CAG, E-AGG/M-CTT, E-AGG/M-CAG, E-AGC/M-CAT, E-AGC/M-CAG). AFLP bands were scored for presence (1) or absence (0) and transformed into 0/1 binary character matrix. AMOVA and mismatch distribution analysis were performed in Arlequin, Nei genetic diversity and Shannon genetic diversity index were calculated in POPGEN. A total of 461 loci ranging in size from 60 bp to 550 bp were detected from 47 individuals, of which 265 were polymorphic. The number of bands per primer combination varied from 66 to 126 and the percentage of polymorphic bands per primer combination ranged from 52.58% to 62.70%. The proportion of polymorphic loci, the Neigenetic diversity and Shannon genetic diversity index of Qingdao and Xiamen populations were 51.70% and 51.99%, 0.1022 and 0.0996, 0.1643 and 0.1622, respectively. The results showed that the genetic diversity of these two populations was at the same level. G_st value, Shannon genetic diversity index and AMOVA analysis indicated that the genetic variation mainly came from individuals within populations and there was no significant genetic differentiation between populations. The UPGMA tree based on AFLP data supported this result and no distinct clade was detectd. Mismatch distribution analysis and dominant gene frequency revealed that these two populations have the same genetic population structure. The results of AFLP analysis indicated that there is no significant genetic differentiation between two populations due to frequent gene flow. At present, the natural populations of marine fish, including N. albiflora, have been destroyed due to over exploitation and increasing marine pollution which caused the number of parent fish to be extremely diminished. These results provide information on the geneticst ructure of N. albiflora, which can be utilized to make informed mana gement decisions. However, AFLP markers are inherited as dominant markers, further studies utilizing codominant markers are needed for a better understanding of the genetic diversity of this fish.

Abstract:Pampus argenteus and Pampus cinereu are two of the most important commercial species of inshore waters in China. The reproductive characteristics of the two species were discussed by means of biological investigation and histological slides analysis with optic microscope. The samples of these two species had been collected continually all year by fishing vessels at sea. The results indicated that the two species had same sex ratio(1∶1) during the spawning season. There was little variation in their gonad and oocyte development. The egg diameter of oocyte at the 5phase was 700-870 μm with one oil globule at 260-400 μm diameter, and the thickness of primary egg membrane was 12-19 μm. They spawn several batchs during one reproductive season. Average fork lengths of female Pampusargenteus and P. cinereu in their reproductive season were 22.5 cm and 29.0 cm, respectively, and the fork length of male Pampus argenteus and P. cinereu in their dominant population was 14.0-17.0 cm and 23.0-25.5 cm, respectively. The reproductive season of Pampus argenteus was from early April to early June and the ovary gonadosomatic index (GSI) went up to its peak from middle of April to middle of May with the maximum 25%. However the reproductive season of P. cinereu was from early June to middle of August and its maximal ovary maximal GSI was 12.6% from late June to late July. In addition, the water temperature in reproductive period for Pampus argenteus and P. cinereu was 18-24℃ and 24-28℃, respectively. The effect of environmental factors on amount of spawning population was also discussed in this paper.

Abstract:In order to assess the relative trophic importance of the spotted velvetfish in Changjiang Estuary and adjacent sea in summer, carbon(δ¹³C) and nitrogen(δ¹⁵N) stable isotope ratios were determined in spotted velvetfish and its potential food sources. Based on an organism's stable isotope ratios are an integration of the isotopic signatures of food sources that have been assimilated through time. The δ¹³C and δ¹⁵N values showed that the spotted velvetfish's food sources were composed of plankton >900 μm, Leptochela gracilis, common squid, Lantern cuttlefish, Beka squid, Common Japanese squid, larvae and juvenile and Japanese sand shrimp. plankton >900 μm was the most important food source, accounting for 69%-75% of the total food by weight. Food proportion of the spotted velvetfish varied among different classes of sizes. Cluster analysis dived the body lengths of the spotted velvetfish into two groups according to stable isotope ratios, >60 mm and <60 mm, respectively. The food proportion of the plankton >900 μm was 68%-87% to 75%-88% for the > 60 mm to <60 mm, while the proportions of larvae and juvenile，mollusk，Crusta ceans to the larger group have a degree of increasing. The proportions of all food sources showed that plankton >900 μｍ was the most important prey, while others were eaten occasionally by the spotted velvetfish. If the production of plankton >900 μｍ decrease in some cases, the spotted velvetfish would increase the other food proportion, sharing the large size food with other org anisms,which would decrease the production of high trophic level organisms or s horten the length of food chain.

Abstract:A trip of productionpossibility frontier on exploring purple flying squid （Sthenoteuthis oualaniensis） has been made by Chinese squid jigging fleet in the northwestern Indian Ocean during September and November in 2003. It aimed to estimate the resources biomass of squid and its distribution. During the investigation period, the main fishing ground was found in the waters near 16°N, 61°E , where the averaged catch per day was more than 5 tons. But the fishing ground disappeared suddenly 10 days later. The situation of fishing ground changing and disappearing have been discussed based on the marine environmental data from the trip and satellite data from website. The upwelling might occur in the area along the coast of Oman. The upwelling might bring the oxygenpoor seawater from deeper layer, which might result that fish gather in pockets of relative oxygenrich water, at the same time fish tend to gather around areas where there is ample food, areas where there is high sea surface chlorophyll-α(Chl-a)) and lower sea surface temperature (SST). The fishing ground had been found at the area with high SST and Chl-a gradient. From the surface winds direction and speed data, it is showed that the appearance of lower cyclone will change the SST and Chla distribution, and plays a very important role in the forming of squid fishing ground.

Abstract:In order to solve the problem of which food conversion ratio was higher and how nutritional disease such as fat liver occurred, when Monopterus albus fed formulated diet, this paper studied the effects of supplement of choline to the diets on the growth, food conversion ratio, lipid content in muscle and liver, hepatosmatic index and activities of 4 digestive enzymes of Monopterus albus. By adding choline of different content to the diets at 9 levels: 0.0%, 0.1%, 0.3%, 0.5%, 0.8%, 1.0%, 1.2%, 1.5% and 2.0%, cultivating M. albus for 90 d. Then the study measured and compared 9 levels of choline to the diets on the growth, food conversion ratio, lipid content in muscle and liver, hepatosmatic index and activities of 4 digestive enzymes of M. albus. The results show that when M. albus feed on diets with different content of choline, the growth rate, food conversion ratio, lipid content in muscle and liver, hepatosmatic index, activities of 4 digestive enzymes are different. Between 0.0% to 2.0% of supplement of choline to the diets, with the increase of choline content in the diets, its growth rate increases, the activities of protease, trypsin, amylase and lipase in foregut, hindgut and liver increase, and food conversion ratio,lipid content in muscle and liver and hepatoamatic index decrease. When the choline content in the diets increases from 0.0% to 0.8%-1.0%, these changes are significant. This suggests that the suitable supplement of choline in the diets is 0.8%-1.0%.

Abstract:This is the first report of sex reversal induced by injection of androgenic gland (AG) extract in brachyuran crabs. In the present study, the extract of AG from the mud crab Scylla paramamosain and the Chinese mitten crab Eriocheir sinensis was injected into newly developed female (i.e. just after sex differentiation) crabs E. sinensis. Masculinization was observed in the injection with E. sinensis as well as S. paramamosain. AG cross activity might exist between the two species. Sex reversal could be achieved at a low dosage of 0.14 (E. sinensis) or 0.06 (S. paramamosain) AG equivalent.

Abstract:The immunologic enhancement of Flavobacterium columnare exopolysacchar ide (EPS) was examined by semi-quantitative RT-PCR in grass carp (Ctenopharyngodon idellus) immunized with intraperitoneal injection of EPS. The total RNA was extracted from liver, spleen, trunk and head kidney of EPS and saline injected fish and reversetranscribed into cDNA, respectively. Five immune genes, i.e. CRP, IL-1, MHC-I, IgM and IFN were then amplified through polymerase chain reaction and semiquantified by comparing brightness of electrophoresis band of the gene PCR product with that of β-actin, respectively. The gene expression of CRP in liver of immunized fish increased significantly, and gene expression of MHC-I in spleen and trunk kidney of immunized fish decreased significantly in comparison with control fish. No significant change was observed in the gene expression of IL-1 between immunized and control fish, and the gene expression of IFN was weakly detected in tissues of both immunized and control fish with no significant difference observed. However, the gene expression of IgM in four tissues of immunized fish increased significantly when compared with control fish. It is thus indicated that the innate and specific humoral immunity of grass carp are effectively activated by immunization with EPS of F. columnare.

Abstract:Environmental hormones and endocrinedisrupting chemicals, are outside factors, which can interfere in the normal function of natural hormone, suc h as production, release, transport, metabolism, link, effect or exclusion. These polluters disordering estrogen and reproduction could link with estrogen receptor and even produce estrogen effects. As a typical kind of environmental hormones, polychlorinated biphenyls(PCBs) are a class of chlorinated hydrocarbon compounds with high chemical stabilities and toxicity, and often extant in environment and bioaccumulated in food chains, leading to a ubiquitous contamination in nature and man. In laboratory studies, technical PCB mixtures have been shown to cause reproductive disorders in fish. But the interferential mechanism of PCB was not found, the aim of the present study was to provide the research of the mechanism with the effects of PCB₁₂₅₄ on testosterone and estradiol of serum and culture ovary in allogynogenetic silver crucian carp. The effects of PCB₁₂₅₄ on sex steroids in allogynogenetic silver crucian carp were investigated by using in vitro injection of PCB₁₂₅₄ 0.5, 5, 50 μg·kg^-1body weight. In injection groups, the levels of testosterone in the serum decreased with time and concentration of PCB₁₂₅₄. Meanwhile, the levels of estradiol in the serum changed in the opposite trend. In the groups of incubated ovarian fragments in vitro with PCB₁₂₅₄, the levels of estradiol secretion from ovary increased with time and concentration of PCB₁₂₅₄ treatments.

Abstract:The cDNA of crayfish (Procambarus clarkii) infected with white spot syn drome virus(WSSV) were used as tester and that of the control as driver to construct a forward subtraction.A reverse subtraction was performed by used tester as driver and driver as tester at the same time.The subtracted products were cloned into the PinPoint^TM Xa-1 T-Vector.Then the recombinated plasmid was transformated into DH5α.A forward subtracted cDNA library including 1620 clone and a reverse subtracted cDNA library including 400 clone were constructed respectively. Using sequences flanking the cloning site as primers,1514 clones were amplified from the 2020 clones constructed by suppression subtractive hybridization.1221 clones of these were got from forward subtraction library and 293 clones from reverse subtraction library. After concentration and degeneration the PCR fragments were robotically printed onto nylon membrane to make a cDNA chip. In the end, 255 positive clones from forward subtraction library and 23 positive clones from reverse subtraction library were detected by cDNA chip.

Abstract:The effects of danofloxacin by oral administration for 20 days at dosage of 20,50 and 100 mg·kg^-1 on the toxicity of Acipenser schrencki were studied. The activity of superoxide dismutase(SOD) ,catalase(CAT),Na⁺，K⁺ATPase，glutathioneStransferase(GSHPX)，glutathione peroxidase(GST) and Maleic Dialdehyde(MDA) content were investigated by kit, so as to appraise security of danofloxacin used in Acipenser schrencki. Meanwhile blood smears were prepared to investi gated the micronucleus and nuclear anomalies rate. The results showed:the SOD activity in erythrocyte remains higher level than that of control group；CAT activity in erythrocyte is higher than control group obviously；The activity of Na⁺, K⁺ATPase in erythrocyte is not significant compared with control group；The GSHPX activity in erythrocyte of 100 mg·kg^-1 is higher than other test groups（P<0.05）；the MDA concentration in erythrocyte remains unchanged；the activity of GST inerythrocyte is not affected by danofloxacin；The erythrocyte micronucleu s remained stable, but nuclear anomalies in test group rose markedly compared with control group. The result showed that the nuclear anomalies are the highest at 10th day, then began to decrease but still is higher obviously than control group. The result showed that danlfloxacin affected SOD，CAT，GSHPX activity, does not changed the Na⁺， K⁺ATPase，GST activity and MDA content. The nuclear anomalies rate of test fish is higher than control group.

Abstract:The hydrolysis of sodium tripolyphosphate (STPP) in the fresh Aristichthys nobilis minced meat and myofibrils (Mf) were studied by using ion chromatog raphy (IC). The results showed that STPP was hydrolyzed to pyrophosphate（PP）and orthophosphate(Pi), whereas pyrophosphate was further hydrolyzed to orthophosphate in Aristichthys nobilis minced meat. In the case of myofibrils, STPP was also hydrolyzed to pyrophosphate and orthophosphate. Myofibrils from Aristichthys nobilis muscle had the activity of tripolyphosphatase (TPPase). The optimum temperature and pH for the TPPase activity of myofibrils from Aristichthys nobilis muscle were 25 ℃，pH 5.5.The TPPase activity of myofibrils was activated at the condition of below 5 mmol·L^-1 Mg²⁺, but was inhibited when the content of Mg²⁺ over 5 mmol·L-1. At the condition of 0.3 mol·L^-1 KCl, the strongest activity was determined at 5 mmol·L^-1 Mg²⁺. The TPPase activity of myofibrils from Aristichthys nobilis was also affected by KCl. The optimum consistence of KCl for the TPPase was 0.3 mmol·L^-1 at the condition of 5 mmol·L^-1 Mg²⁺. EDTA-Na₂ could inhibit activity of TPPase.

Abstract:The method of adding chemical compounds in the induction of settlement and metamorphosis of eyespot larvae of Coelomactra antiquata was adopted in this paper. The aim is to research the induction of epinephrine, γ-aminobutyric acid(GABA),LDOBA and Ca²⁺ on the settlement and metamorphosis of C. antiquata larvae,and to assess optimum induction concentrations of four chemicals. The results indicated that exposure to 10^-4 mol·L^-1 epinephrine induced significant levels of both settlement and metamorphosis in larvae of C. antiquata, and settlement, metamorphosis and survival rates were 88.3%, 92.8% and 98.14% respectively. Larval settlement and metamorphosis rates of C. antiquata were 86.9% and 87.6% respectively, in the presence of GABA at the concentration of 10^-5 mol·L^-1. L-DOPA could induce the metamorphosis of eyespot larvae of C. antiquata. Maximum metamorphosis rate of 73.3% and settlement rate of 84.9% were achieved after exposure to 10^-6 mol·L^-1 L-DOPA. The results also showed that the settlement and metamorphosis rates were 86.1% and 42.8% respectively, and the survival rate of the larvae was 87.4% when induced by 20 mmol·L^-1Ca²⁺. When the larvae were induced by exposure to the optimal concentrations of above four kinds of chemicals respectively, the settlement and metamorphosis rates of the larvae were observed significantly higher than those in controls.It was discussed that the induction technique of using Ca²⁺ could apply to the artificial breed of C. antiquata in large scale production.