Levels of Metabolic Enzymes and Nitrogenous Compounds in the Swimming Crab Portunus trituberculatus Exposed to Elevated Ambient Ammonia-N

This study investigated the effects of ammonia-N(control and 1 and 5 mg L~(-1)) on the changes of metabolic enzymes activities and nitrogenous compounds levels in the gills, hepatopancreas, muscles, and hemolymph of the swimming crab Portunus trituberculatus. Results showed that the levels of hemolymph ammonia increased significantly at 6 h and then decreased to control level at 12 h. The activities of key enzymes involved in biosynthesis pathways of glutamine(Gln) and urea, such as glutamate dehydrogenase(GDH), glutamine synthetase(GS), and arginase(ARG), increased remarkably in different tissues of P. trituberculatus. Gln level was the highest in muscles, while urea content was the largest in hepatopancreas. The m RNA expression level of xanthine dehydrogenase(XDH) was downregulated sharply in gills but upregulated in the hepatopancreas and muscles. A small amount of uric acid was detected in the hemolymph. This result suggests that Gln in muscles may be used as an intermediary nitrogen repository, while urea may be primarily synthesized in the hepatopancreas and then transferred to other tissues via the hemolymph. Uric acid could be synthesized by the anabolism of purine nucleotides, in additon to the inhibitory activities of gills. In addition, the levels of Gln, urea and uric acid exhibited dose-and time-dependent effects with ammonia content. Therefore, P. trituberculatus could possibly convert ammonia into Gln, urea, or uric acid to detoxify ammonia during high ambient ammonia-N stress.     

Phospholipid Compositions in Portunus trituberculatus Larvae at Different Developmental Stages

Phospholipids are used to improve the growth and survival of Portunus trituberculatus,a widely cultured crab species in China.However,only total phospholipids or several classes are applied in crab diets.In this study,we employed a targeted lipidomic method to investigate the comprehensive phospholipid composition in P.trituberculatus larvae and reveal the changing phospholipid profile over the larval development.Results showed that P.trituberculatus larvae contain 112 phospholipid species belonging to 10 phospholipid classes,in which phosphatidylcholine(PC)and phosphatidylethanolamine(PE)species are the most abundant,and PC,PE,phosphatidic acid,and phosphatidylserine(PS)are with high concentrations.The levels of all phospholipids significantly changed with larval development,which was highlighted by the downward parabolic changes in PE,phosphatidylglycerol,phosphatidylino-sitol,PS,lysophosphatidic acid,and sphingomyelin levels.In addition,nearly all phospholipid species were depleted at the M stage,which probably contributed to the mass mortality of crab larvae.These findings on the composition and alterations of phospholipids in P.trituberculatus larvae provide novel perspectives for the targeted supplementation of phospholipids in crab diets.Our work also highlights the use of targeted UHPLC-MS lipidomics in understanding the changes of phospholipids during crab development.     

Production of Pacific White Shrimp Polycultured with Swimming Crab at Different Densities,and Nutrient Budget in the Enclosure System

This study was to evaluate the effect of stocking densities on the nitrogen and phosphorus budgets for a polyculture of the crab Portunus trituberculatus and the Pacific white shrimp Litopenaeus vannamei.The shrimps(initial weight,0.012 g)were cultured at a density of 45 shrimp m^-2,and juvenile crabs(initial weight,0.024 g)were cultured at five densities of 0,3,6,9,and 12 crabs m^-2.The treatments were grouped as C0S45,C3S45,C6S45,C9S45,and C12S45,respectively.Water quality parameters,growth of shrimp and crabs,and nitrogen and phosphorus budget were measured.The results indicated that the shrimp performances in polyculture treatments C3S45 and C6S45 were superior to those in the crab-free treatment(C0S45).The crab in treatments C3S45 and C6S45 exhibited a significantly higher final mean weight and carapace width/length than those in treatments C9S45 and C12S45.The final size and survival of crabs had a negative correlation with the increasing crab stocking density.The contents of total phos-phorus and total nitrogen and the comprehensive contamination index values were higher in the C9S45 and C12S45 treatments than in the other treatments.The conversion ratios of nitrogen for crab and shrimp growth in treatment C3S45 were significantly higher than those in the crab-free treatment.These findings indicate that polyculturing shrimp with crabs at suitable densities can improve productivity,profitability,nutrient utilization,and the environmental quality.From the 60-day treatments,the optimal culture densi-ties were 3-6 crabs m^-2 and 45 shrimps m^-2.     

Impacts of sampling design on the abundance estimation of Portunus trituberculatus using crab pots

Fishery-independent surveys can provide high-quality data and support fishery assessment and management.Optimization of sampling design is crucial to increase the quality of fishery surveys.Crab pots are important fishing gears used to catch crabs.We analyzed the impacts of sampling design of crab pots on the abundance of Portunus trituberculatus in the Changjiang(Yangtze) River estuary to the Hangzhou Bay and its adjacent waters in East China Sea.The crab pots were cylindrical,240 mm in height and 600 mm in diameter of the iron ring.Our sampling designs(including fixed-station sampling,simple random sampling,stratified fixed-station sampling,and stratified random sampling),three number of stations(9,16,and 24),and three numbers of crab pots(500,1 000,and 3 000) were simulated and compared with the "true" abundance that obtained from bottom trawl survey s in the study area in 2007.The scenarios with16 stations were set in stratification as a control group for comparison with unstratified designs.Results show that simple random sampling can obtain more stable results than fixed-station sampling in the abundance estimation of P.trituberculatus.In addition,stratified sampling resulted in more accurate abundance than unstratified sampling.The accuracy of the simulated results improved with the increase of the number of stations.No remarkable differences in the re sults were found among the scenarios of different number of crab pots at each station.However,resource-intensive areas exerted great impacts on simulation results.Thus,prior information or pre-survey results about resource abundance and density distribution are necessary.This study may serve as a reference for future sampling designs of crab pots of P.trituberculatus and other species.     

Identification of the Major Allergen in Greasy-Back Shrimp （Metapenaeus ensis） by MALDI-TOF-MS

In order to better understand shrimp allergen,some basic characters of the major allergen of greasy-back shrimp (Metapenaeus ensis)were investigated.The major allergen was extracted and separated,and its peptide mass fingerprint(PMF) was analyzed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF-MS).After screening in the NCBI database with Mascot searching engine,the results indicated that the major allergen of greasy-back shirmp was muscle tropomyosin.Database matching search showed that the top protein matched,i.e.the tropomyosin from giant tiger prawn(Penaeus monodon),had a Mowse value of 268.In addition,there were 27 queries matched with the allergen in greasy-back shirmp with an amino acid sequence coverage value of 65%.The matching scores and the sequence coverage values were also high with tropomyosins of other invertebrates,including Tyrophagus putrescentiae and Lepisma saccharina.These results indicated that the allergen of Metapenaeus ensis had high homology with other crustacean allergens,and provided molecular explanations for the high cross-reactivity of the major allergens between crustaceans and some other invertebrates.     

Using iTRAQ-Based Quantitative Proteomics Analysis to Identify Differentially Expressed Proteins Related to Larval Development of Portunus trituberculatus

The swimming crab Portunus trituberculatus is an economically important marine crustacean species. Here isobaric tags for relative and absolute quantitation(iTRAQ) analysis were used to identify proteins that are differentially expressed during larval development of P. trituberculatus to elucidate the underlying mechanisms. In comparison with the first zoea stage(Z1), 3980 proteins were identified from 32789 peptides, which were matched with 115497 spectrums. A total of 241 proteins were screened with significantly differential expressions in all development stages. These 241 proteins are involved in various biological processes, such as cytoskeleton organization, protein synthesis, energy production and substance metabolism, physiological activities, and transport.Cluster analyses of the 241 differentially expressed proteins led to the generation of four protein clusters based on the overall similarity in protein expression patterns. Exactly 54, 70, 36, and 45 proteins clustered in profiles 10(0, 0, 0,-1, 0, 0), 15(0, 1, 0, 1, 0, 1), 18(0, 1, 2, 2, 1, 0), and 19(0, 1, 2, 3, 4, 5), respectively. Muscle development and exoskeleton renewal were important processes throughout the development stages. In addition, protein synthesis, degradation, and digestion actively occurred, especially at the Z4 stage. These results provide novel insights into the mechanisms underlying larval development of swimming crab and can assist in larval rearing.     

Influence of Ultrasonic Treatment on the Allergenic Properties of Shrimp （Penaeus vannamei ） Allergen

The present study was undertaken to determine whether high intensity ultrasound could reduce the allergic properties of shrimp allergens. Reducing the allergenic properties of these allergens will be beneficial to allergic individuals. Samples of shrimp protein extract and shrimp muscle were treated by high-intensity ultrasound with water bathing at 0 ℃ or 50 ℃ for different time periods. The treated and untreated samples were then analyzed by SDS-PAGE, Western blots and competitive inhibition ELISA （Ci-ELISA） to determine the shrimp allergenicity. The results show that high-intensity ultrasound has no effect on allergenicity when the extracts were treated at 0℃. However, a significant decrease was observed in the level of the major shrimp allergen, Pen a 1, when the samples were treated at 50 ℃. In the determination of allergenicity with CiELISA, a reduction in IgE binding was also observed.     

A Comparative Study of Genetic Variation betwccn Chinese Mitten Crab Eriocheir Sinensis and Hepu Mitten Crab E. Hepuensis

Horizontal starch gel electrophoresis was used to investigate the biochemical genetic structure of Chinese mittencrab Eriocheir sinensis and Hepu mitten crab E. hepuensis. Sixteen putative enzyme-coding loci were examined for forty-nineChinese mitten crabs and thirty-eight Hepu mitten crabs. Nine loci, AAT-1 *, AAT-2 *, G3PDH*, GPI*, IDHP-1 *, IDHP-2 *,MDH-1 *, MDH-2 * and PGM * , are polymorphic in Chinese mitten crab, and seven, AAT-1 *, AAT-2 *, GPI *, IDHP-1 *,MDH-1 *, MDH-2 * and PGM * , are polymorphic in Hepu mitten crab. The proportion of polymorphic loci and the expectedheterozygosity are 0.562 5 and 0.080 3 for Chinese mitten crab, and 0.437 5 and 0.075 4 for Hepu mitten crab. The Nei' s ge-netic distance between two species is 0.002 4.     

Monthly Variation in Crustacean Assemblage(Decapod and Stomatopod) and Its Relationships with Environmental Variables in Laizhou Bay,China

In this study,we investigated the community structure of crustaceans(decapod and stomatopod) inhabiting the sandy mud bottoms of Laizhou Bay(northeastern China) monthly from May 2011 to April 2012.Investigation was stopped from December 2011 to February 2012 because of the extreme weather and sea ice.A total of 205,057 specimens belonging to 31 species(shrimp,15;crab,15;and stomatopod,1) were collected in 148 hauls.From 2011 to 2012,Oratosquilla oratoria was the dominant biomass species(47.80%),followed by Charybdis japonica(15.49%),Alpheus japonicas(12.61%),Portunus trituberculatus(6.46%),and Crangon spp.(4.19%).Crangon spp.was the most dominant species by individual(32.55%).O.oratoria was the most-frequently encountered species(81.76%),followed by Palaemon gravieri(70.95%),C.japonica(65.54%),A.japonicas(62.16%),and P.trituberculatus(54.73%).The biomass density increased from August to September 2011 and decreased from March 2012 to April 2012.The dynamics of the ecological indices evolve in a similar manner,with high values of diversity and evenness and rich species from May to June 2011 and low values from September to October 2011.O.oratoria,C.japonica,and P.trituberculatus differed by biomass data between groups I(samples obtained from September to October 2011) and II(samples in other months).These species contributed more than 70% to the similarity of the crustacean community structure.Furthermore,the subsets of environmental variables that best matched the crustacean-assemblage structure were as follows:water depth(WD) in summer(June to August);sea surface temperature(SST),dissolved oxygen(DO),and WD in autumn(September to November);and DO,salinity,and WD in spring(March to May).The calculated correlation coefficients and significance level were higher in the period of July to August 2011 than in other months.Comparing 2011 to 2012 with 1982 to 1983,the species composition remained stable.However,the dominant species changed significantly.High value and large species,such as F.chinensis,P.trituberculatus,and T.curvirostris,have been replaced by low value and small species(i.e.,Crangon spp.,P.gravieri,and C.japonica).     

Dietary effects of A zolla pinnata combined with exogenous digestive enzyme(Digestin~(TM)) on growth and nutrients utilization of freshwater prawn,Macrobrachium rosenbergii(de Man 1879)

The present study was conducted to evaluate the effect of either individual or combined wheat bran(WB) replacement with Azolla pinnata supplemented with Digestin~(TM) in the diet of freshwater prawn, Macrobrachium rosenbergii Postlarvae(PL) on growth performance, nutrient utilization, chemical body composition and survival(%). Experimental diets were a wheat bran-soybean based diet with no Azolla and Digestin~(TM)(control, T_1), and diets containing 17% Azolla supplemented with Digestin~(TM) 0%(T_2), 1%(T_3), 2%(T_4) and 3%(T 5). Each experimental diet was allocated into three tanks(6 m~3/tank) fed for 12 wks. Each tank was subdivided into three equal pens by nets(2 m~3) and stoked with 84 PL/m~2. The experimental diets were readily consumed by prawns PLs where both high growth and good feed efficiency were achieved for all diets. The results showed that the diets containing A. pinnata supplemented with Digestin? at the level up to 3% have the higher growth and better nutrient utilization than the control diet. No differences were observed for moisture and protein content among the experimental diets. However, the highest protein content was observed on prawns fed on diets T_1 and T 5 respectively, while the lowest value was recorded for T_4 diet. The results also show that prawn PLs fed the diets contain A. pinnata and supplemented with Digestin~(TM) recorded the highest values of body lipid content compared to the control diet. Feed efficiency and economic conversion rate(ECR) values show that economic performance and the cost-effectiveness of the A. pinnata supplemented with up to 3% Digestin~(TM) recorded the highest net return, and therefore it is recommended for prawn, M. rosenbergii PL's. These results are clearly indicating that A. pinnata have a good potential for use in prawn diets at reasonable levels than other conventional diets.     

提取蚀变信息时TM影像的最佳波段组合研究

多光谱数据的最佳波段选择直接影响图像的目视解译和信息提取。在分析TM影像各波段间的标准差、相关系数和最佳指数因子内在联系的基础上,提出采用最佳指数因子与蚀变信息光谱特征相结合的方法选择遥感影像的最佳波段组合。研究表明,最佳指数因子与蚀变信息光谱特征相结合是多光谱数据最佳波段选择的理想方法;TM4+TM5+TM7波段组合获取的合成图像构造清晰,岩性差异显著,最有利于蚀变信息的提取。     

Analysis of DNA Methylation Level of Portunus trituberculatus Subjected to Low Salinity with Methylation-Sensitive Amplification Polymorphism

In this study, the methylation-sensitive amplification polymorphism(MSAP) was used to compare the genomic DNA methylation level of muscle, gill and hepatopancreas of Portunus trituberculatus subjected to salinity 12 for 30 days to illustrate the epigenetic mechanism of osmoregulation. Thirty primers were used to analyze the difference of methylation level of different tissues. The results showed that the baseline methylation level of muscle, hepatopancreas and gill was 47.31%, 22.94% and 17.69%, respectively. After exposed to low salinity stress, the methylation epiloci changed in the three tissues. Both demethylation and methylation processes occurred under low salinity stress. The methylation ratio decreased in muscle and gill but increased in hepatopancreas. These results indicated that DNA methylation is tissue-specific when P. trituberculatus responds to low salinity.     

洪涝灾情快速反应的星载SAR与TM数据的融合处理

本文以1998年鄱阳湖地区发生洪涝灾害为研究实例,应用灾中的Radarsat-SAR和灾前的TM数据,以不规则三角网(TIN)方法实现了高精度几何配准。然后,通过波段相关性和信息熵值分析,选取最佳波段。最后,对不同融合模型得出的图像进行对比,得出结论:以灾中的SAR和灾前TM的一个近红外(TM5或TM7)以及一个可见光波段(TM3最佳),分别赋RGB色合成的图像效果很好,既能识别区分洪涝淹没的绝产区、土壤滞水减产区和未受灾区,并且操作简便,能满足洪涝灾情快速反应要求。     

EVALUATION ON NUTRITIVE VALUE OF PORTUNUS TRITUBERCULATUS

MODUon0NP.trituberculatushasanupswellingshuttiedsecephalothoraxcrust0nWhosescheearemanydiversefinegranuleswhichbererneroughandconcentIatalinthegillaas.IthasthIeewartybulgesandtwosharpodhonitSforeheadPaiAiy-U,l986).Thechelipedisdeveloped,itSmerusisshuuleshaped,thefotirthpairofpereiopodsareuseoars,itScarpopoditiswideandshort,thedaCty0poditisslab-shaped,eachjoint'sedgehasshorthahaforswirTuning,themalebodyisbluishgreen,andthebolebodyisadeeppmple.Maleanddrileglands')locateatthedorsalordorsola…     

Effects of Several Immunostimulants on Phenoloxidase and Hemocytes of the Crab Charybdis japonica

To investigate the stimulating effects of immunostimulants on the autogenous immunocompetence of crabs and the possible mechanisms involved, the immunostimulating effects of β-1,3-glucan, lipopolysaccharide (LPS), inactivated Vibrio harveyi and Vibrio anguillarum on phenoloxidase (PO) and hemocytes of Charybdis japonica were investigated in this study. It was found that the yields and the enzymatic activities of purified PO in C. japonica increased significantly after the crabs were treated with immunostimulants, while the unit enzymatic activities remained almost the same. After treatment with β-1,3-glucan and LPS, the amount of rough endoplasmic reticulum (RER) and the number of mitochondria in both semigranular cells and granular cells increased greatly, and the number of cytoplasmic granules decreased but with enlarged volume. However, the corresponding characteristics of hyaline cells remained almost the same. On the other hand, the number of granules in semigranular cells decreased greatly, and the number of mitochondria of hyaline cells increased greatly, after treatment with inactivated vibrios. It may be concluded that the effect of polysaccharide immunostimulants on the innate immune system of C. japonica is different from that of inactivated vibrio immunostimulants. The immunity-enhancing mechanism of polysaccharides in crab autogenous immunocompetence is probably accomplished by the increased yields of PO and total PO activities, while that of inactivated vibrios is probably accomplished by the partially increased yields of PO and total PO activities as well as the significantly improved phagocytotic abilities of semigranular cells and hyaline cells.     

EFFECTS OF FISHES ON THE GROWTH AND SURVIVAL OF CHINESE MITTEN CRAB, ERIOCHEIR SINENSIS

This experiment on the effects of fishes (Pelteobagrus fulvidraco, Channa argus, Cyprinus carpio) on growth and survival of juvenile Chinese mitten crab,Eriocheir sinensis was conducted in cages placed in Bao'an Lake. T-tests showed that effects of fishes on the carapace width growth were not significant as compared with the control (no fish). But the effects of these fishes on crab survival were negative. The causes of negative effects of fishes on crab survival were simply discussed on the basis of food habits. This research was financially supported by the Department of Agriculture (96-008-02-03), P. R. China, and Chinese Academy of Sciences (Key Project: KZ951-A1-102).     

Comparative analysis of mitochondrial genome of a deepsea crab Chaceon granulates reveals positive selection and novel genetic features

Deep-sea organisms survive in an extremely harsh environment.There must be some genetic adaptation mechanisms for them.We systematically characterized and compared the complete mitochondrial genome(mitogenome) of a deep-sea crab(Chaceon granulates) with those of shallow crabs.The mitogenome of the crab was 16 126 bp in length,and encoded 37 genes as most of a metazoan mitogenome,including 13 protein-coding genes(PCGs),22 transfer RNA(tRNA) genes,and 2 ribosomal RNA(rRNA) genes.The gene arrangement and orientation was conserved in the crabs.However,a unique mitogenome element regulator,the origin of light-strand replication(O L),was firstly predicted in the present crab mitogenome.In addition,further positive selection analysis showed that two residues(33 S in ND3 and 502 I in ND5) in C.granulates mitogenome were positively selected,indicated the selective evolution of the deep-sea crab.Therefore,the mitogenome of deep-sea C.granulates showed a unique O_L element and positive selection.These special features would influence the mitochondrial energy metabolism,and be involved in the adaptation of deepsea environment,such as oxygen deficits and low temperatures.     

Studies on culture condition and extracellular hydrolase of psychrophilic bacteria from Arctic sea ice

Arctic sea ice in the polar region provides a cold habitat for microbial community. Arctic sea ice microorganisms are revealed to be of considerable importance in basic research and potential in biotechnological application. This paper investigated the culture condition and extraceIlular hydrolase of 14 strains of different Arctic sea ice bacteria. The results showed that optimal growth temperature of strains is 15 ℃ or 20 ℃. The optimal pH is about 8.0. They hardly grow at acid condition. 3 % NaCl is necessary for better growth. These strains have different abilities in producing amylase, protease, eellulase and lipase. Pseudoalteronomas sp. Bsi429 and Pseudoalteronomas sp. Bsi539 produced both cellulose, protease and lipase. These results provide a basis for further developing and exploiting the cold adapted marine enzyme resources.     

三种海水蟹类血淋巴细胞的比较

采用Wright’s-Geimsa混合染料对三疣梭子蟹、锈斑蟳和锯缘青蟹的血淋巴细胞进行染色,光镜下对这三种海水蟹的血淋巴细胞进行了观察和比较。根据血淋巴细胞的核质比、细胞整体着色特点、细胞质中颗粒的存在与否、颗粒的数量和分布等,三种海水蟹的血淋巴细胞均可分为颗粒细胞、半颗粒细胞和透明细胞三种主要类型,三种蟹的同类血淋巴细胞在染色形态上稍有差别。三种蟹的颗粒细胞大小差异较大,而半颗粒细胞和透明细胞大小相近。三种蟹的血淋巴细胞均以半颗粒细胞为主,透明细胞在血淋巴细胞中所占比例最小。三种蟹的血淋巴细胞密度均存在明显的雌雄差异,锈斑蟳和锯缘青蟹血淋巴细胞分类计数也存在明显的雌雄差异。     

Enhanced Biodegradation of High-Salinity and Low- Temperature Crude-Oil Wastewater by Immobilized Crude-Oil Biodegrading Microbiota

High salt and low temperature are the bottlenecks for the remove of oil contaminants by enriched crude-oil degrading microbiota in Liaohe Estuarine Wetland(LEW),China.To improve the performance of crude-oil removal,microbiota was further immobilized by two methods,i.e.,sodium alginate(SA),and polyvinyl alcohol and sodium alginate(PVA+SA).Results showed that the crude oil was effectively removed by the enrichment with an average degrading ratio of 19.42-31.45 mg(L d)^?1.The optimal inoculum size for the n-alkanes removal was 10%and 99.89%.Some members of genera Acinetobacter,Actinophytocola,Aquabac-terium,Dysgonomonas,Frigidibacter,Sphingobium,Serpens,and Pseudomonas dominated in crude-oil degrading microflora.Though the removal efficiency was lower than free bacteria when the temperature was 15℃,SA and PVA+SA immobilization im-proved the resistance to salinity.The composite crude-oil degrading microbiota in this study demonstrated a perspective potential for crude oil removal from surface water under high salinity and low temperature conditions.