Abstract:The main biological characteristics, the main spawning period and fecundity characteristics of jack mackerel spawning stock were studied based on the sample data obtained in Diaoyu Islands waters in March, 2005. The results showed that the Diaoyu Islands waters was the main jack mackerel spawning grounds in the southern East China Sea in spring, and the main spawning period was in the middle and late March. According to the fork length and weight frequency distribution, the main spawning stock had three dominant folk length groups, 200~220mm, 230~240mm, and few individuals of 260~270mm. The composition characteristics of body weight were consistent with the fork length composition. Since the samples were taken in the late spawning period, the conditional growth factor was relatively small. The absolute individual fecundity was 12 264±37 629 grain and the mean individual absolute fecundity was 27 844±10 179 grain. The fecundity per unit length was in the range of 50~157 grain/mm, at an average of 121±42 grain/mm. The fecundity per unit net weight was in the range of 82~238 grain/g,at an average of 202±68 grain/g.

Abstract:Based on the fish sampling data during fish migration season (March to June, 2006) in the Buyuan River, which is a first order tributary at the downstream of the Lancang River, the fish species diversity was analyzed. With reference to the water quality and hydrological data in the same period, relationship between the environment and the number and weight of fishes were analyzed using artificial neural network (ANN). Results showed that 34 species belonging to 3 orders, 8 families and 22 genera, were collected, among which 8 species were migratory species. Diversity index (H') was at the highest in March, while domination index (Sp) was higher in April and June for the appearance of Mystacoleucus chilopterus and Opsariichthys bidens. There was no significant difference of evenness index (E) among all the sampling months. The lowest water level (LWL) was the most influential factor on the number of fish species, followed by the average runoff velocity (ARV) and average water temperature (AWT). The highest water temperature (HWT) had significant influence on the weight of fish, followed by the total phosphorus and the dissolved oxygen. The simulant values obtained by using ANN were similar with the actual values, which indicated that the ANN method was applicable.

Abstract:According to the biological data of small yellow croaker Larimichthys polyactis (Bleeker，1987) collected by the trawls in the central and southern Yellow Sea in the late of 2008, the growth characteristics of this fish were studied. The results showed that the body length and body weight increased 15.22±5.65 mm and 5.46±0.65 g every month from July to December. The relationship between the body length and body weight was W=3.76×10-5L2.830 3. The relationship between the otolith and body length and body weight was discussed. The Power function was the fittest one among the three models. Growth models VBGF(von Bertalanffy function), LGF(Logistic function), GGF(Gompertz function), Logarithmic function, Inverse function, Quadratic function and Cubic function were chosen as the candidates for fitting the growth model of small yellow croaker underyearling to establish the optimum growth equation of body length. Based on the R2 and RSS criterion, the VBGF, LGF and GGF had the similar fitness, while the Logarithmic, Inverse, Quadratic and Cubic had the similar fitness, and the effect was better than VBGF, LGF and GGF. The Cubic function was the best one to describe the growth of small yellow croaker underyearling.

Abstract:An outbreak of serious mortality among the cultured black sea bream Sparus macrocephalus, characterized by a swollen intestine containing yellow fluid, occurred in early spring of 2008 in Xiangshan Bay, Zhejiang Province. Four strains of bacteria, BB01, BB02, BB03 and BB04 were isolated from the intestinal yellow fluid of the moribund black sea bream with Zobell2216E agar and/or thiosulfate citrate bile salt sucrose agar. The former three were identified as causative agents via artificial challenge. The 96h LD50 value of BB01 was 461×104 CFU/g bream body weight. The former three strains were characterized as Pseudomonas putida through morphological, physical and biochemical profiles tests and 16S rRNA sequencing. The three strains were Gram negative, short rod shaped, oxidase and glucose oxidation positive, gas negative, maltose and citrates utilization positive, arginine dihydrolase positive, nitrate reduction positive, DNA enzyme and acetamidase negative. The 16S rRNA sequence of BB01 shared the identity of over 99% with those of P.putida strains. Drug sensitivity tests showed that the strain BB01 was susceptible to most of the tested antibiotics while resistant to ofloxacin.

Abstract:The effects of Ca2+ and Mg2+ concentrations (1∶3) in artificial seawater on growth, superoxidase dismutase（SOD）and catalase（CAT） enzymatic activity of juvenile Paralichthys olivaceus were investigated at 20℃ and salinity of 30. Five concentrations of Ca2+ and Mg2+ were set as follows, 700 (group A), 1 100 (group B), 1 600 (group C), 2 200 (group D), and 2 800 (group E) mg/L, respectively. It was found that stress response occurred including bend of fish body and reduction of ventilation rate and food intake in juvenile fish under high Ca2+ and Mg2+ concentrations and the fish recovered after a week. After 60 d experiment, the survival rates of all groups were over 90%, and there were no significant differences in food conversion efficiencies (FCE) among different groups (P>005). However, the effects of Ca2+ and Mg2+ concentrations in artificial seawater on specific growth rates (SGR) and food intake (FI) at 0~20 d were significant (P<005). After 20 d, there was no significant difference in FI (P>005). During 40~60 d, the SGRs in low Ca2+ and Mg2+ concentrations groups were lower than that in high concentrations groups. The SGR and FI of all groups decreased when Ca2+ and Mg2+ concentrations increased. At 60 d, the SGR and FI of group A were 113 and 104 times as that of group E, respectively. Significant influences were also observed in immuno enzymatic activity of liver among these groups (P<005). The activity of SOD and hepatosomatic index of group D were significantly higher than those of other groups . However, the activity of SOD of group E was lower than other groups. The activity of CAT of group E was also lower than other groups but no significant difference (P>005). The results indicated that the concentrations of Ca2+ and Mg2+hindered the growth of juvenile P. olivaceus by changing their food intake. The fish in low Ca2+ and Mg2+ concentration groups grew fast and those in high Ca2+ and Mg2+ concentration groups grew slowly. There were no significant differences in the SGR of all the groups except for group E after 60 d rearing experiment. High Ca2+ and Mg2+ concentrations could restrain immuno enzymatic activity of juvenile P. olivaceus. It is suggested that high concentration of Ca2+ and Mg2+ should be avoided in the culturing of juvenile P. olivaceus.

Abstract:Fenneropenaeus chinensis 5-Bromouracil(5-BrUra)Disease resistance Growth

Abstract:he spatial and temporal variations of planktobacteria quantity in Apostichopus japonicus culture ponds during autumn and winter(from September,2008 to Februray,2009)were investigated by plate culture and fluorescence microscope techniques.The total number of planktobacteria estimated by acridine orange direct count(AODC)and Direct Viable Count(DVC)ranged in 1.0×106~2.1×107 cell/ml and 4.0×105~5.7×106 cell/ml,respectively.The amount of heterotrophic bacteria detected by heterotrophic plate count(HPC)ranged from 0.6×102 to 1.66×105CFU/ml.The amount of Vibrio was below 6.6×103CFU/ml using TCBS selective medium cultivation.The number of planktobacteria varied in different season,which reached the maximum in October and the minimum in January.The number of bacteria in the ponds was significantly higher in autumn than in winter(P<0.01).The total number of bacteria in different water layers was determined.The results showed that the total number of bacteria in the bottom water was significantly higher than that in the surface water(P<0.05).There was significant correlation(P<0.01)between data deduced by using different bacterial count methods.

Abstract:

Abstract:Genetic linkage map of Portunus trituberculatus was constructed using “double pseudo testcross” strategy with AFLP and SSR molecular labeling techniques. In this study, maternal and paternal broodstock were sampled from Laizhou Bay and Zhoushan Island wild populations of P. trituberculatus, respectively. Through the directional mating (1♂×3♀), the wild parents were used to generate F1 family. The 108 progenies of directional mating F2 family coming from sisterhood intercross between F1 family and its parents were used as experiment materials. Sixty AFLP primer combinations and 3 pairs of SSR primers were screened to produce 214 maternal markers and 195 paternal markers. There were 155 markers which included 153 AFLP markers and 2 SSR markers segregating as 1∶1 model from 214 maternal markers. 139 markers which contained 138 AFLP markers and 1 SSR marker segregated in 1∶1 model in 195 parental markers. The female linkage map consisted of 100 markers distributed in 9 linkage groups (more than three markers), 6 triplets and 15 doublets, which covered a total length of 1 544 cM. Average marker spacing of female linkage map was 22.0 cM and the observed genome length accounted for 52.9% of the estimated genome length. The paternal linkage map consisted of 71 markers distributed in 6 linkage groups, 6 triplets and 11 doublets, spanning the distance about 1 174.2 cM with average marker spacing of 24.0 cM and its genome coverage was 49.5%. The Pearson analysis results showed that all the genetic markers distributed in linkage map uniformly. This study provided a starting point for further construction of high density linkage maps, as well as for the mapping of functional genes and the location of quantitative trait loci (QTL) in P. trituberculatus.

Abstract:The fish scale of tilapia taken as the raw material, the orthogonal test design was adopted to study the pretreatment of the fish scale of tilapia by using alkali method, acid method, enzyme method, acid and saline method, and saline and alkaline method. Then the five optimized processes were compared and the gelatin extraction process was studied. The results showed that the enzyme method was the optimum extraction process of gelatin from tilapia fish scale. The process was as the following: pretreatment with 0. 2% acidic protease solution with pH adjusted to 3.5, enzymatic hydrolysis at 35 ℃ for 1h, then the fish scale was rinsed to pH 5~6 after being soaked for 4 h in 2% hydrochloric acid. The gelatin extraction process includes 3 heating steps,heating at 65 ℃ for 3 h, heating at 70 ℃ for 35 h and heating at 75 ℃ for 35 h, then the final product was filtrated and the filtrate was pooled.

Abstract:Two sets of specific oligonucleotide primers for Haplosporidium sp. and Marteilia refringens, along with two TaqMan probes specific for each protozoan parasite were designed with Primer Express 20 software. The reaction parameters such as the concentration of two pair of primers, two TaqMan probes and the reaction buffer were optimized to develop duplex real time PCR assay for the rapid detection of Haplosporidium sp. and Marteilia refringens. The sensitivity of duplex real time PCR assay was 40 template copies for Haplosporidium sp. and Marteilia refringens. The duplex real time PCR assay was found to be specific and to be able to detect and differentiate Haplosporidium sp. and Marteilia refringens, and no positive results were observed when nucleic acid from Perkinsus sp.,Pseudomonas fluorescens, Vibrio parahaemolyticu, V. Alginolyticu, V.Fluvialis and V. Mimicus were used as duplex real time PCR templates. This duplex real time PCR assay is a rapid, sensitive, and specific test for detection of Haplosporidium sp and Marteilia refringens and will be useful for the control of these protozoan parasites in shellfish.

Abstract:Two trials for Sargassum thunbergii seedling production and three times seedling nursery were conducted between 2007 and 2009 in Xiaoshidao of Weihai and Xuejiadao of Qingdao in Shandong Province. About 10 concrete ponds were employed as seedling production facilities, each with 2×5 m2. Parent strains were collected from different sites in Weihai, and it was proved that the local strain in Xiaoshidao was better for seedling production than the other S. thunbergii collected. Six types adherences substratum were used for seedling adherence, and it was indicated that bamboo curtain was the most efficient one, which yielded over 30 ind./cm2 of seedling densities. The total yielded seedling was 7,507,000 individuals in 2007, with average of 3.5 mm in length; while in 2008, it was 30,154,200 individuals with 3.2 mm in length. Early developmental observation showed that S. thunbergii seedlings owned higher survival rate and went through the normal development, but seedlings may be detached from the substrate when reached to 4 mm in length.The seedling density decreased to 0.2~1.0 ind./cm2 when the seedling length reached to 10 mm.

Abstract:According to the monitoring data of water quality and runoff volume of the Guan River and Sheyang River from 2004 to 2008,water quality assessment was done using Nemerow pollution index method and the volume of major pollutants entered the sea was assessed. Based on the research results, the influence of the pollutants on the coastal marine environment was studied, and the relationship between the change of pollutant volume and the outbreak Enteromorpha prolifera in the central and southern Yellow Sea was also analyzed. The results showed that the water quality of Guan River was good from 2004 to 2008, but the pollution index showed an upgrade tendency. The water in Sheyang River was slightly polluted in 2005 and 2007, while the water quality was good in 2004, 2006 and 2008. The runoff volume of the Guan River and Sheyang River varied obviously from 2004 to 2008, which were 15.72×108 m3 to 41.93×108 m3 and 28.74×108 m3 to 59.96×108 m3, respectively. The mean annual total volume of main pollutants(CODMn, BOD5, NH4-N and phenol) entered the sea from the Guan River and Sheyang River were 18,794.87 t and 43919.24 t, respectively. The volume of CODMn and BOD5 accounted for over 89% from 2004 to 2008 and both were the leading factors. The pollutants entered the sea from the Guan River and the Sheyang River had effected the coastal marine environment obviously, and had been related to the breaking out of Enteromorpha prolifera in the central and southern Yellow Sea in recent years.

Abstract:A multiplex polymerase chain reaction (PCR) was developed for synchronous detection of three major pathogens of aquatic animals, including Vibrio anguillarum, Aeromonas hydrophila and Edwardsiella tarda. Three important and special virulent genes were selected for the multiplex PCR, such as positive transcriptional regulator toxR gene of V. anguillarum, aerolysin gene aerA, which encodes important virulent factor of A. hydrophila, and evpA gene, which encodes the apparatus protein of secretion system T6SS of E. tarda. The reaction condition was optimized and the specificity and sensitivity was tested for this method. The detection for real samples also proved its reliability.

Abstract:Vibrio harveyi is a kind of important pathogenic bacteria for seawater animals and can bring about severe loss to their culture production. V.harveyi strain GYC1108-1, which was isolated from diseased Pseudosciaena crocea in Zhejiang Province in 2003, was identified after morphological observation, biochemical characteristic analysis, 16sRNA and HSP60 gene sequence detection. Earlier research in our laboratory demonstrated that the extracellular protease（ECPase）was the major virulent factor of GYC1108-1 and was identified as cysteine protease with a molecular mass of 55 kDa as estimated by SDS-PAGE．The protease can decompose the casein of degrease milk and was named as 1108 ECPase．The method of solid LB culture medium containing 1% skim milk was established to detect relative activities of 1108 ECPase and YZ ECPase (YZ is a recombinant bacteria that can express the target ECPase). The optimum cultivation time of GYC1108 1 and YZ excreting ECPase was 36 h and the optimal concentration of ammonium sulfate for ECPase purification was 70% for the above mentioned method．In this study, 1108 ECPase and YZ ECPase were purified by 70% ammonium sulfate and Sephadex G-25 gel filtration for antigens standby. Three different substances A, B and C were chosen to be adjuvants. A was a common adjuvant which was a mixture of white oil and Tween 80; B was propolis adjuvant and C was Freund's adjuvant. The purified 1108-ECPase and YZ-ECPase antigens were mixed with A, B and C before being applied separately to immunize 3 ICR mouse. The adjuvant free and blank treatments were designed as control. There were nine treatments in the immunization test. The immune sera of the mouse were collected after three immunizations. Except the blank control, multi clone antibodies showed high ELISA titers in the sera of eight treatments, which were between 1∶1.6×104～1∶2.56×105. The titers of immune sera in different treatments were ranked as C>A>B>adjuvant free, and it is suggested that the adjuvants chosen followes this effectiveness sequence. The result of Western blot revealed one clear band in ECPase, which had a molecular mass of 55 kDa as tested by mouse anti serum. These results indicated that the prepared 1108-ECPase and YZ-ECPase had good immunogenicity, and this work has provided some basic knowledge for further study on genetic engineering vaccines.

Abstract:Immunoassays are demonstrated to be simple, rapid, and cost effective. For the past decades, more and more techniques are being developed which combine the advantages of approaches in highly powerful analytical tools. The convenience and speed of the test have been achieved by a novel concept of colloidal gold based immunochromatographic assay, which depends on the transport of a color labeled reactant to its binding partner immobilized on the surfaces of the membrane. This new technique has been proved to be a well established and appropriate technique for a variety of point of care and field use applications, which was widely used as a convenient test for human being and terrestrial animal diseases in nowadays. In this paper, progress on the study of this new technique has been summarized in detail, which focuses on its principle, application and development. Based on these, its practicability in aquatic animal is also introduced. It would provide a new point of view on the innovation of simple and fast detection of disease in aquaculture.

Abstract:The bacterial community composition in the intestine and aquaculture pond of Apostichopus japonicus cultured at Zhuanghe, Dalian was primarily analyzed with 16S rDNA PCR-DGGE fingerprint. The results showed that 41, 31 and 41 bands were observed in the PCR-DGGE fingerprint of intestine, sediment and seawater, with 12, 9 and 10 dominant bands respectively. 20 bands different in abundance were shared by intestine, sediments and seawater, and 4 of them were dominant bands in each sample. 5, 2 and 13 special bands appeared in each sample, in which band 34, 42 from intestine and band 25, 26 from water were with higher amount than the others. Similarity between the intestine and sediment was 77.88%, while 65.9% was defined between the intestine and seawater and 556% between the sediment and seawater.