Abstract:The introduced Japanese scallop has been farmed in China, primarily along the coasts of the Bohai Sea and the North Yellow Sea, since the 1980s. Following the adaptation and localization of in China, specific problems have appeared after several decades of aquafarming: especially low survival and slower grow. This can be associated with an effectively small population that experiences inbreeding, eventually leading to unknown degrees of decline in its genetic diversity after several generations. Consequently, it is important to carry out genetic diversity research on in its main mariculture area of China. In this study, mitochondrial cytochrome-b (Cyt ) gene-sequencing techniques were used to determine genetic diversity and variations among six stocks of : the bottom-sowing cultured stocks of Changdao, Haiyangdao and Zhangzidao; the natural stocks of Lüshun and Japan; and the artificial selected stock of Zhangzihong. A total of 20 haplotypes were found among 120 sequences of Cyt The minimum number of haplotypes was found in the Zhangzihong artificial breeding stock, and the maximum number occurred in the Japan Aomori Mutsu Bay stock, with a haplotype diversity () of 0.10000 and 0.88400, respectively. Analysis of molecular variance (AMOVA) showed the percentage of variation between the Japan and China groups was 15.34%, which was significantly higher than when measured as one genetic pool. For the China group, 83.41% of the genetic variation come from within the stocks, and only 1.52% of the variation was among the stocks in the group, indicating that genetic variation between individuals of the China group was higher than between the stocks. analysis showed a moderate level of genetic variance among the Japan stock and China stocks (0.07455-0.17895, P. yessoensis was calculated based on the Tamura-Nei model; the maximum genetic distance was found between the Japan Aomori Mutsu Bay stock and the Haiyangdao bottom-sowing cultured stock; the minimum genetic distance was found between the Zhangzidao bottom-sowing cultured stock and the Zhangzihong artificial breeding stock. The UPGMA method was used to construct a phylogenetic tree based on genetic distances among the six stocks of ; the tree shows that the five China stocks clustered into a genetic branch, and then these clustered together with the Japan group. These results demonstrate obvious genetic differentiation between the China stocks and the Japan stock of this scallop. Moreover, genetic diversity was lower in the China stocks as compared with the Japanese stock. The findings of this study could be usefully applied to scientific strategies for the protection, sustainable development and commercial utilization of scallop germplasm resources.

Abstract:For this study, a double haploid population of Japanese flounder was produced using mitotic gynogenetics. Body weight and 7 morphological traits (i.e., total length, dorsal-fin length, pelvic-fin length, body height, caudal peduncle width, head width and trunk length) were measured. After normalizing the measures, we acquired a data set that could explain 90.4% of these principal-component analysis traits (PPCTs), and accordingly we denoted these PPCTs. Next, by modifying conditional probabilities of quantitative trait loci (QTL) genotypes on the distorted flanking markers, Bayesian model sel­ection was used to dissect the genetic arc­hitecture of the PPCTs with four genetic linkage maps, created with the software JoinMap4, MapDisto, JoinMap-DistortedMap and MapDisto-DistortedMap. The different methods used to construct the genetic maps and QTL mapping of the PPCTs produced variable results. Comparing the map produced with JoinMap4, the rel­ative positions and the genetic map distances of the partial separation markers differed from the map produced with MapDisto. Furthermore, five separation markers were not located at the corresponding linkage groups. Comparing the maps based on JoinMap 4 and MapDisto and after correction by DistortedMap, the positions were not changed, but the genetic map distances of the partial separation markers were changed. In the overall maps, three additive-effect QTL were detected in linkage groups 6, 9 and 22, of which a negative effect could account for 12.95%, 14.85%, 11.56% and 11.76% in linkage group 6; a negative effect could account for 13.86%, 13.27%, 11.17% and 11.25% in linkage group 9; and a positive effect could account for 5.68%, 4.36%, 4.97% and 3.58% in linkage group 22. At the same time, 28, 19, 29 and 20 pairs of additive-additive interactions were identified, mainly distri­buted in linkage groups 6, 7, 9, 17, 20 and 22, and these interactions could account for approximately 2.19%-17.62%, 2.40%-22.26%, 2.08%-26.0%, and 3.16%-22.05% of the variance in the PPCTs, respectively. We believe that the results of QTL mapping were more accurate in the linkage map using DistortedMap based on JoinMap4.

Abstract:The is a commercially important anadromous species distributed mainly in the Yangtze River of China and in coastal waters of China, Korea and Japan. The species is popularly known locally as one of the three most delicious fishes in the Yangtze River, along with the obscure pufferfish; additionally, is the main source of income for many Yangtze River fishermen. migrates to sea for growth and so experiences a wide range of salinities, making osmoregulation especially important for the species during this migration. The protein aquaporin-1 (AQP1), a member of the aquaporin family of integral membrane proteins, functions in transporting water as well as ammonia, urea and glycerin. Accordingly, it is important for hypertonic and hypotonic regulation and maintaining osmotic homeostasis in an organism. To investigate the osmoregulatory role of AQP1 in , we used rapid amplification of complementary DNA (cDNA) ends (RACE method) to clone, for the first time, aquaporin-1 from this species. The full-length cDNA covered 1 299 base pairs (bp) with a 777 bp open reading-frame (ORF) within the 5' untranslated region (UTR) of 107 bp and the 3'UTR of 458 bp. The ORF encoded 258 amino acids, with a calculated molecular mass of 27.1 kDa, and theoretical isoelectric point of pH 6.13. The results show that AQP1 of possesses typical conserved domains of the AQP family, including six transmembrane helices and two NPA (Asn-Pro-Ala) motifs, and one binding site for mercury (II) chloride (HgCl₂), which is an AQP1 inhibitor. Next, we analyzed the homology and phylogenetic systematics of the AQP1 sequence from . The highest homology and similarity (reaching 93%) occurred between the AQP1 of . The AQP1 of displayed an evolutionary relationship the furthest from AQP1 of mammals. We performed real-time quantitative PCR (qPCR) to study the expression of AQP1 in various tissues of , namely gill, brain, mid-kidney, anterior intestine, posterior intestine, spleen, liver, heart and muscle. The results showed that it expressed in many of the tissues (including gill, mid-kidney, anterior intestine, posterior intestine, liver and muscle), but the level of expression was highest in the brain. Lastly, we performed a high-salinity stress test to discover the osmoregulatory function of AQP1 in under conditions of hypertonic stress. The expression of AQP1 in gill, mid-kidney, anterior intestine, and posterior intestine significantly differed from that in tissues of the controls (<0.05). These results demonstrate that AQP1 indeed plays an important role in osmoregulation in , and the study provides a theoretical reference for further research on the metabolic costs of osmoregulation in .

Abstract:Ras-related nuclear protein (Ran) is a small GTPase with many functions, such as hydrolysis of GTP, control of cell development, replication of DNA, and RNA transcription. In this study, the cDNA-encoding ) was cloned using expressed sequence tag (EST) analysis and a rapid amplification of cDNA ends (RACE) approach. The full-length cDNA of was 1191 bp, comprising a 5' untranslated region of 218 bp, a 3' untranslated region of 405 bp, and an open reading frame of 648 bp. The deduced protein had 215 amino acid residues with a molecular mass of 24.6 kD, and 7.13 point of theoretical isoelectric. Ran belongs to the P-loop NTPase super family. Ran has a PTZ00132 model that crosses multiple domains. The members of P-loop NTPase super family have extremely conservative nucleotide sequences. Phylogenetic analyses indicate evolution of Ran proteins within the animal kingdom is very conservative, with that of . Quantitative real-time RT-PCR analysis revealed the gene was expressed in testis, ovary, brain, muscle, eyestalk, abdominal nerve, heart and gill tissues. The ovary has the highest level of expression and the eyestalk has the lowest level of expression ( gene expression of ovary is seven-eight times higher than that of testis, and the expression level of gene increased with the development of ovary. After ovulation in ovarian regression period, the expression level of gene was at a low level. After RNA interference (RNAi), expression of gene in an experimental group of adult females was significantly lower than in the control group (<0.05). After RNA interference (RNAi) in the mature female prawns, the expression of gene in experimental group (injected dsRNA solution into the shrimp's pericardial cavity) was significantly lower than in the control group (injected equal amount of DEPC water into the pranw's pericardial cavity) ( gene changes with the development of ovary. The expression of gene increased from the early stage of ovarian development to the mature stage and decreased rapidly after ovulation in the ovary. Expression of in ovarian tissues in the experimental group was also significantly lower than that of control group (<0.05), indicating RNA interference was effective. Expression of vitellogenin was significantly affected by RNA interference, with expression in the experimental group significantly lower than in the control group ( gene plays a regulatory role in expression of the gene is involved in female ovary development.

Abstract:Epigenetic regulation of the related genes DNA methyltransferase (DNMT1), histone deacetylase (HDAC3), and histone methyltransferase (MLL5) were used to analyze gene-expression quantity in the respiratory tree, intestine, coelomic fluid, and muscle tissue of the sea cucumber under high-temperature stress conditions. The housekeeping gene of the sea cucumber was used for internal calibration, and the relative quantification 2^{-ΔΔmethod was employed using real-time fluorescent quantitative PCR. Sea cu­­cumber kept at <15℃ were used as a control group, and the target gene-expression quantity of this group was used as benchmark-1. Increased expression of the DNMT1 gene was insignificant in the coelomic fluids at 18℃, but the expression rose significantly between 18℃ and 21℃, thereafter the increase was less acute, until the ex­pres­sion quantity reached a maximum of 3.26 at 30℃. The expression quantity of the MLL5 gene and the HDAC3 gene were significant at 2℃, and then the expression quantity increased slightly as the temperature increased, reaching a maximum of 2.90 and 3.19, respectively, at 30℃. Overall, the expression quantity of the three genes increased with increasing temperatures, and this trend was obvious, reaching a maximum at 30℃. Comparisons of different tissues showed diverse expression quantities of the three genes. Expression of the genes was prominent in the coelomic fluids, at >2.5, above 24℃, while the expression quantity of the three genes in muscle tissue was <2 in the whole spectrum of high temperatures. Under the stress conditions of 30℃, the gene-expression quantity of the three genes was detected at 0 h, 3 h, 6 h, 9 h, 12 h and 15 h, and the expression quantity of every gene at 0 h was used as benchmark-1. The data showed that the expression quantities of the three genes were characterized by increasing expression with the extension of stress time during 0 to 9 h (<0.05), except expression of HDAC3 in intestine stabilized at about 6 h. The expression quantity was either not obviously or only slightly lower after 9 h. In addition, the trend in expression quantity was consistent in the different tissues, yet the maximum expression quantity of every gene was highest in the coelomic fluids than in any of the other tissues. This expression analysis of epigenetic-related genes under high temperatures lays a foundation for interpreting the mechanism of response to high-temperature stress in sea cucumber from the viewpoint of epigenetic regulation.}

Abstract:Artificially induced gynogenesis is a major technique used for controlling the sex ratio of aquatic animals as well as for culturing selectively bred colonies. In this study, cryopreserved sperm of Japanese seabass was UV-irradiated and used to fertilize eggs of starry flounder and diploid gynogenesis was induced using cold and hydrostatic pressure shocks. Meanwhile, sperm but without cold or hydrostatic pressure shock were used as the haploid group; sperm were used for hybridization; and sperm were used as a control group. The results showed that cell division started at a similar point in time in the control group as in the gynogenetic haploids and hybridized embryos. However, as compared with the controls, embryonic development among each of the three experimental groups was retarded from the late blastula stage. All the hybridized embryos died before formation of an embryonic body; all the haploid embryos died before the heart started to beat. Thus, none of the embryos in these two groups could be successfully hatched. Embryonic development of a small portion of the gynogenetic diploids followed a pattern resembling that of the controls, although with less success: (58.01±5.30)% of embryos in the control group were hatched out at 100 h 10 min, with (35.11±6.19)% malformed; in comparison, only (0.11±0.01)% of the gynogenetic diploids were hatched out at 104 h 50 min, with (53.59±0.36)% malformed. Although the gynogenetic diploids had a very low hatching rate and a relatively high rate of malformation, we were still able to construct a gynogenetic colony of with normally developing larvae. Hence, we propose that this study uncovers a promising technique for gynogenesis, and the results provide abundant cytobiological evidence contributing to information on the developmental biology of haploid, hybridized, and gynogenetic embryos of this species.

Abstract:The marine bivalve mollusk is mainly distributed in tropical and subtropical waters of the southwestern Pacific Ocean, including coastal areas of China, Japan, Philippines, India, Indonesia, Australia and New Zealand. The species is relatively new to shellfish mariculture, having potentially high economic value and good prospects for development. This study used biological productivity methods to investigate the species' reproductive cycle, gonad development, fatness, breeding season, embryo development and larval development in the period from August 2013 to July 2014. In the examination of its breeding cycle, the gonadogenesis of presented a unimodal gametogenic cycle, with one spawning season annually. The gonad development of the species could be divided into five stages: proliferating stage, growing stage, maturing stage, spawning stage and resting stage. This shellfish's mode of reproduction is oviparity, with a slightly greater gonad development among males than female. The species displays gonochorism, with a female to male ratio of 1:1. The spawning season was recorded as from October to March. Peaks in spawning occurred in March, October and November, at water temperatures of 20.3-24.6℃. Fatness peaked at 33.43% in August, and the lowest measure of fatness was in April at ~18.31%. The cultivation of embryos and larvae revealed demersal, elliptically shaped eggs, and the egg diameter ranged from 70 to 80 μm. Under the conditions of water temperatures of 24-26℃ and salinities of 28-32, the fertilized eggs developed into D-shaped larvae, 90-110 μm in length, within 16 hours of hatching. After 7-8 days, the D-shaped larvae then developed into late-umbo larvae, 210-230 μm in length. Thereafter, the late-umbo larvae developed into juvenile mollusks within 4-5 days. These observations provide a foundation for the artificial breeding and protection of genetic resources of .

Abstract:The dynamic energy budget (DEB) theory is a mechanistic theory of metabolism that captures the flows of energy and matter throughout the entire lifecycle of an organism. A DEB model of an individual organism describes the rates at which the organism assimilates and utilizes energy for maintenance, growth and reproduction, as functions of the state of the organism and its environment. The purpose of the present paper is to setup a DEB model for Japanese scallop , the main mariculture species of northern China, in order to understand the energy distribution characteristics of the scallop and to analysis the influence of temperature and food availability on its growth and reproduction. The model depends on seawater temperature and food density (chlorophyll-a concentration), as forcing variables. The key DEB parameter values for the scallop were calculated based on previous experiments in our laboratory. We calibrated and validated with stage data for mariculture efforts in Sungo Bay from May to December 2013, and for small-, middle-and large-sized scallop from Changhai, China, from August 2007 to July 2008. The DEB model developed here showed good growth simulations and provided an extensive description of the energetic allocation of throughout its growth span. Meanwhile, it was demonstrated that, in summer, seawater temperature was the main limiting factor for scallop growth. Especially in Sungo Bay, at days 45-125 (nearly 85 days' duration), the value of T-dependence was very low, as scallop growth was limited by water temperature, and the scallop almost ceased to grow at this stage. In Changhai, at days 40-320, food limitation was stronger than the temperature restriction, which proved that mariculture densities might become overloaded. The results of the sensitivity analysis showed the model was relatively sensitive to changes in _AM increased by 10%, the growth of the scallop could be increased by 13%. Therefore, the sensitivity of these parameters can have a great impact on the results of the model, thus it is crucial to have accurate measurements of the parameters.

Abstract:Hybridization is the most effective thus widely used technique in the artificial breeding of grouper. Salinity is one of the most crucial environmental variables affecting embryonic development in marine fishes. To study the effect of salinity on fertilized eggs of the hybrid grouper () in terms of egg diameter, hatching rate, morphology and survival of the yolk-sac larvae, fertilized eggs were reared in seawater with different salinities, and the effects on total egg diameter, oil globule diameter, hatching rate, abnormality rate, as well as the morphology and survival of the yolk-sac larvae, were observed. The diameter of the fertilized eggs increased with decreases in salinity, whereas the diameter of the oil globule appeared unaffected by the choice of salinity in the test range. Larvae hatched out in the salinity range 18 to 36, but the rates of hatching and abnormalities were significantly affected by salinity level (<0.05). With increases in salinity, the hatching rate first increased at a salinity up to 75.5%, and then decreased at salinity of 30, whereas the rate of abnormalities followed an opposite pattern. The yolk sac was almost completely absorbed within 3 days of hatching at a salinity between 24 and 36, and there was no significant difference between the groups. Within the same salinity range, the survival activity index (SAI) of newly hatched larvae first increased and then decreased with increases in salinity. The newly hatched larvae had an SAI greater than 5; by 5 days after hatching they were starved to death, with half the deaths at 3.5 days after hatching. The fertilized eggs of this hybrid grouper can be hatched out in a salinity range of 24-36, with the optimal salinity at 30-33.

Abstract:The effects of replacement of a dietary microalgae with a microencapsulated diet on gene expressionrelated to pearl production traits, growth and biomineralization were studied in the pearl oyster . Three experimental groups were set up: EG1 was fed the marine green alga ; EG2 was fed a mixed diet of the microcapsules and the algae; and EG3 was fed only the microcapsules. The expression level of the growth-related genes in the adductor muscle (A), and the biomineralization-related genes in the mantle central (MC) and mantle edge (ME), were detected and compared among the three treatments after a 170-day indoor farming period. The results showed that survival rate, retention rate, the pearl nacre thickness, and average mass of the harvested pearls did not significantly differ (EGFR, I in the A did not significantly differ among the three treatments (N19 in the MC did not significantly differ among the three treatments ( was higher in EG2 and EG3 than in EG1, and the expression level of <0.05). No significant differences were detected in the relative expression levels of in the ME among the three treatments ( was higher in EG2 and EG3 than in EG1 (<0.05). These results suggest that the microencapsulated diet can replace part of the microalgae diet, which may be helpfully applied in future studies of artificial feed development for pearl oyster .

Abstract:The green mud crab has been widely cultured in coastal areas of southern China for many years, with production reaching approximately 14.1 tons in 2015. The successful farming of mud crabs presently depends on conventional diets, comprising mainly trash fish, mollusk meat and animal viscera, but the demand for these foods by the crab industry is difficult to satisfy. Compared with the traditional feeds, compound feeds offer better-balanced nutrition and have better storage qualities. Studies of the nutritional requirements of cultured mud crab are important to developing cost-effective, environmentally friendly and nutritionally balanced artificial diets. Dietary protein, a major macronutrient in aquatic animal diets, provides the essential and non-essential amino acids needed to synthesize body protein and energy. As protein represents the most expensive component in feeds, it is important to determine the optimal level for a cultured species. This study investigated the effects of dietary protein level on growth performance, body composition, and digestive enzyme activities in mud crab () juveniles. Six isoenergetic diets were formulated with varying protein content (i.e., 31.45%, 36.37%, 41.55%, 46.13%, 51.72% and 56.86%) and fed to juvenile mud crabs for three weeks. Triplicate groups of 80 juvenile crabs, with an average body weight of 11.86±0.11 mg, were randomly assigned to each of the six diets. Each crab was raised in a bucket (20 cm diameter × 25 cm height) provided with a 5 cm layer of sand. The results showed that the dietary protein level did not influence crab survival rate in any of the treatments (<0.05). The weight gain ratio and specific growth rate of the juvenile crabs significantly increased as dietary protein was increased from 31.45% to 51.72% (<0.05), and then the indicators showed a slight decrease with a further increase to 56.86% protein; the gains were highest for crabs fed 51.72% protein. Dietary protein level significantly affected the whole-body crude protein, lipid and ash contents (<0.05), but did not show any significant influence on the moisture content of the crabs (>0.05). Values for crude protein content of crabs raised on one of the four higher protein diets (41.55%-56.86%) were significantly higher than that for crabs fed a lower protein diet (31.45% and 36.37%) (<0.05), while the whole-body lipid content significantly decreased as the dietary protein content was increased (<0.05). The trend in whole-body ash content was similar to that observed for weight gain ratio, which significantly increased at first and then decreased as the dietary protein content was increased (<0.05). With an increasing level of dietary protein, the protease activity significantly increased (<0.05). The level of dietary protein also affected lipase activity in the crabs. Under the present experimental conditions, juvenile green mud crabs fed 41.55%-56.86% protein exhibited the best growth performance, and regression analysis showed that the optimal level was 49.03% protein. These results will be helpful for formulating pelleted diets for commercial mud crab culture. Future studies of protein metabolism mechanisms and the interactions of proteins with other nutritional components are recommended for this species.

Abstract:Biofloc technology (BFT) is a zero-water exchange, environmentally friendly aquaculture system that is now widely used since it is proven to enhance the growth and immunity of farmed fish. Gibel carp (also known as Prussian carp) is a major aquaculture species in China, especially around Yancheng in Jiangsu Province, and it is also grown in mudflats. Developing an effective aquaculture system for gibel carp is also helpful for exploring mudflats usage in fish aquaculture. However, sudden outbreaks of disease have become a great and common threat to carp-farming operations, as this causes huge economic losses. Since 2009, an epizootic with severe mortality rates caused by cyprinid herpes virus 2 (CyHV-2) has emerged among cultured gibel carp in China, especially in the northern part of Jiangsu Province. The disease develops fast, spreads widely to almost all main areas of gibel carp, and mortalities may reach 90%-100%. Thus, disease becomes a limiting factor for aquaculture, yet effective methods to prevent or control CyHV-2 are lacking. In order to discuss the practical usage of BFT in mudflat fish culture in terms of disease prevention, this study subjected the representative species (gibel carp) to the disease, using fish cultured in two different systems: in a BFT system without any water exchange, and in a common recirculating system with 1/4-1/3 water exchange daily (as control), to analyze the effect of BFT on growth performance and disease resistance in this carp. Growth factors, digestive enzymes, and immune-related enzyme activities were detected by common methods using commercial kits; heat-shock protein 70 (HSP70) mRNA relative expression was analyzed with qRT-PCR; and disease resistance against bacterial infection was evaluated by challenge tests. The results demonstrated the following: (1)The rates of weight gain, specific growth, and survival of the gibel carp in the BFT group were significantly higher than occurred for fish in the control group (<0.01); however, the condition factor and viserosomatic ration did not significantly differ between the two groups (>0.05). (2)The activity of digestive enzymes (i.e., amylase, lipase and protease) in the BFT group was significantly enhanced (by 53.10%, 28.10% and 17.99%, respectively) in comparison with the control group (<0.05). (3)The super oxide dismutase activity in skin mucus, total antioxidant capacity in liver, and lysozyme in serum were significantly induced in the BFT group (<0.01). (4)HSP70 mRNA relative expression in the BFT group was up-regulated 1.29-, 1.34-, 1.87-and 1.68-fold in the spleen, kidney, liver and gill, respectively. (5)Disease resistance against bacterial infection was improved in the gibel carp eventually challenged by . The cumulative mortalities of gibel carp cultured in the BFT system after experimental injection with bacterial concentrations of 4.2×10⁷ CFU/mL, 4.2×10⁶ CFU/mL and 4.2×10⁵ CFU/mL, were 62.5%, 25%, and 0 within 7 days, respectively. These values were much higher than those detected among fish in the control system (i.e., 75%, 37.5% and 12.5%). This study demonstrated that BFT can improve growth performance, digestive enzyme activity, and disease resistance of gibel carp, and that BFT could be successfully more often used in mudflat fish aquaculture. We suggest that future studies with gibel carp cultured in BFT systems should investigate the optimum concentrations of total suspended solids, prebiotics specialized for mudflat fishes, and relevant molecular immune mechanisms.

Abstract:We investigated the effects of water temperature, salinity, and dissolved oxygen (DO) content on the swimming ability of juvenile tiger puffer , cm/s) and maximum swimming speed (was determined under different temperatures (15.2℃, 20.2℃, 25.6℃ and 30.4℃), salinities (0, 10, 20, 32 and 40), and DO content (2.14 mg/L, 4.10 mg/L, 5.81 mg/L and 7.36 mg/L). The both increased as water temperature ( increased from 13.12 to 29.07 cm/s; increased from 17.51 to 37.38 cm/s. The relationship between could be interpreted with quadratic models, as: ²=0.995 (T-12.27, U_max also increased as DO was increased from 2.14 to 7.36 mg/L: increased from 16.43 to 33.65 cm/s. The relationship between DO and could be interpreted with quadratic models, as: =0.995 (+11.10DO-3.919, <0.01). The salinity challenges had no significant effects upon of the juvenile tiger puffer (>0.05). Temperature is known to strongly influence the contractile properties of muscles. According to "compression of the recruitment order theory", recruiting all of a fish's aerobic fibers at a lower speed results in reduced sustainable performance. The effects of temperature on the physical properties of water may also affect fish swimming ability. The viscosity of water decreases as temperature increases, and this may increase the swimming efficiency of fish in warmer waters. The swimming ability of juvenile tiger puffer might be limited by the cardiorespiratory system in response to hypoxia as a result of impaired functioning of the oxygen transport and exchange system. Furthermore, as another response to hypoxia, increases in respiratory frequency to improve the absorbance of oxygen must evoke an additional energy cost to the animal. The absence of any significant changes in the of the juvenile tiger puffer following the various salinity challenges could be a direct result of this species' exceptional ability for maintaining plasma homeostasis and water balance in the tissues. In natural environments, lower temperatures and hypoxia would reduce the swimming ability of juvenile tiger puffer and thereby weaken their predation and/or predator-avoidance capacity. These findings increase our understanding of the biology and ecology of the tiger puffer, and may be useful for enhancing stocks of this near-threatened species.

Abstract:This research assessed the effect of five tagging methods: namely, PIT marking, fin amputation marking, fluorescent marking, ultrasound marking and T-type marking, on the critical swimming speed of grass carp . Grass carp with a body length of 15-25 cm (standard length, SL) were divided into three size groups: short body length (15-18 cm), medium body length (18-21 cm) and long body length (21-25 cm). Critical swimming speed was measured for 144 fish in an annular flume with a water temperature of (16±1)℃. SPSS17.0 software was used for data processing. The results showed that the absolute critical swimming speed of the grass carp tended to increase, while the relative critical swimming speed tended to decrease along with an increasing body length. Ultrasonic tagging showed a significant impact (<0.01) on the swimming ability of the fish in all size groups (body length 15-25 cm). The absolute critical swimming speed of the grass carp in each of the three size groups (short, medium, large) decreased by 18.72%, 16.40% and 23.15%, respectively, after ultrasound tagging, and the relative critical swimming speed of the three groups decreased by 18.95%, 17.78% and 21.86%, respectively. T-type tagging also showed a significant impact (<0.01) on the swimming ability of grass carp in the short body length group: their absolute critical swimming speed decreased by 8.35%, whereas their relative critical swimming speed decreased by 9.30%. No significant impact of T-type tagging was detected on the critical swimming speed of grass carp with a medium or long body length (18-25 cm). Finally, no significant impact on swimming speed was detected for fish with a PIT tag, fin amputation marking or fluorescent tag.

Abstract:Numerous commercial fish populations have declined in recent years owing to the impacts of overfishing, climate change, environmental pollution, and other factors. However, the cephalopod fishery has continued to develop, including increasing harvests of Japanese squidL. japonica plays an important role in the food web of the Haizhou Bay ecosystem, where it is a primary catch in bottom trawls and some net fishing. The species spawns in the bay and adjacent waters annually in early May, and it uses these waters as a nursery ground. Despite the ecological and economic importance of the species, few studies of its spatio-temporal patterns of distribution are available, and spatial autocorrelation of data is often lacking in the studies. Analysis of temporal and spatial heterogeneity in the distribution of in Haizhou Bay and adjacent waters has significance for sustainable utilization and scientific management of this fishery. To study this, we used the survey data of bottom trawls in Haizhou Bay and adjacent waters, collected in 2011, 2013, 2014 and 2015, for spatial analysis of center of gravity and semivariogram plots to investigate temporal and spatial distribution of resource density and average body size. The average mantle length of was larger in spring than in autumn for each of the four sample years, and the average mantle lengths did not significantly differ (>0.05) for spring, but did significantly differ for autumn, except in 2011 and 2013, 2013 and 2015. The population was distributed throughout Haizhou Bay and its adjacent waters, and local distribution showed no correlation with size. In both spring and autumn, the dominant size class was 20-50 mm mantle length. The resource density of <0.05) between spring and autumn, except in 2013, and was greater in autumn than in spring. Center of gravity of the resource density changed in opposite directions between spring and autumn: squid were more concentrated in the autumn than in the spring of each year; yet in both seasons, the resource density was relatively centered in the middle of Haizhou Bay and adjacent waters; differences in center of gravity among the years and seasons were not significant. Spatial heterogeneity in resource density of appeared to be caused less by randomness than by structural variations. The spatial distribution of in the study area displayed a well-organized structure, with medium to strong spatial autocorrelation in 20-30 km.

Abstract:Purple sea urchin is an important benthic species in the western Pacific Ocean. In recent years, environmental disruption and over-exploitation have damaged the resources of this species. Stable isotope analysis has become a powerful tool for studying the feeding habits of marine animals, and this method can overcome some of the limitations associated with traditional stomach contents analysis. Carbon (δ¹³C) and nitrogen (δ¹⁵N) were used as tools to evaluate the stable isotopic characteristics, trophic levels, shell diameter, and feeding habits of , from Daya Bay on the south coast of Guangdong Province, using data collected in August 2015. In the present study, the trophic levels of 15N. Average δ¹³C of N was (9.14±0.38)‰, and the trophic levels ranged from 2.09 to 2.13 (mean 2.11); no significant correlation was found between the different shell diameters of the 15N (C values of the potential food sources were between-(9.93±1.59)‰ and-(20.76±1.42)‰, and their average δ¹⁵N values were between (-0.16±1.34)‰ and 14.99‰; the trophic levels ranged from 1.34 to 3.77. Cluster analysis showed that the food web among major species of Daya Bay could be classified into four trophic groups: suspended organic matter, primary producers and primary consumers, secondary consumers, and top predators. There were two main channels in the overall food chain: one was the grazing food chain, and the other was the detritus food chain. have a tendency to feed on detritus in August, when corals are scarce, macroalgae density is low, and the terrigenous materials present as particulate organic matter (POM) into Daya Bay follow a period of heavy rains. The main food resources of were: POM, sediment organic matter (SOM), the brown macroalgae , bacillariophyceae (diatoms), zooplankton and phytoplankton Among these, POM was the most important food item for , accounting for 67.3% of its food consumption; the other food categories accounted for 9.7%, 9.3%, 6.7%, 3.7% and 3.3% of its food composition, respectively. Considerable competition exists between and the benthos, which suggests urgent efforts are needed to improve this ecological environment as well as to increase the seaweed biomass. Variations in the trophic levels were related to differences in the isotope baseline, and temporal and spatial variations. There was no significant correlation between shell diameter of and its feeding habits, perhaps because its food source is relatively singular. Thus, it is evident that feeding habits play an important role in population resource recovery and ecological niches of the system.

Abstract:Human activity has put great pressure on the sustainable utilization of fishery resources. In order to quantify human activities based on the degree of interference, a model of the marine ecological footprint was calculated in order to understand the condition of Jiangsu coastal areas from 1978 to 2014. The system dynamics approach used a dynamic nutrition model and marine ecological footprint (MEF) analysis to build a system dynamics model of the marine ecological footprint of Jiangsu coastal areas. It is feasible to verify the model by way of historical fisheries data from 2006 to 2014, simulating the ecosystem development of Jiangsu coastal waters from 2006 to 2022 under different fishing intensities. The simulation results show that from 1978 to 2014, the marine ecological footprint of Jiangsu coastal waters increased from 2.41×10⁶ hm² to 6.61×10⁶ hm², with a mean value of 5.38×10⁶ hm², which is higher than the ecological carrying capacity of 3.45×10⁶ hm², by approximately 1.5 times as much. This means that development in Jiangsu coastal waters has been at an unsustainable level according to the model simulation analysis; and assuming that the fishing intensity will only strengthen, the marine ecological footprint will fall sharply, and eventually to a zero value. The footprint drops faster and the time of tending to a zero value becomes shorter with increasing development. Based on the calculation of threshold catch, we need to reduce the fishing intensity by 50%, from now forward, to allow the fishery biomass to bounce back with smooth reductions in the ecological deficit, which might prevent a collapse in the fishery ecosystem. The research results are a worthwhile reference for making decisions on sustainable development in Jiangsu Province, and especially for forecasting ecological effects along the Jiangsu coast.

Abstract:, with several varieties, is one of numerous species of unicellular free-swimming marine green algae that are widely used for intensive rearing of a variety of marine animal larvae. is a dinoflagellate capable of producing powerful biotoxins and it is commonly found in shrimp ponds. Chief physical and chemical concerns of the commercial growing environment of microalgae are pH and the nitrogen-phosphorus (N/P) ratio. To identify the optimum growth conditions of Prorocentrum minimum in shrimp ponds, we tested for interspecies competition at various nutrient concentrations after determining the optimum pH for each species. Prior to the start of each test, cultures were maintained for an adaption period of approximately 5 generations. Thereafter, the experiments were conducted in batch cultures and continued up to 10 days, and the microorganisms were grown until either late exponential phase or early stationary phase. The pH levels and N/P ratios chosen took into account the common environmental conditions found in shrimp ponds. With this in mind, a culture experiment was first carried out using four different pH levels (7.5, 8.0, 8.5 and 9.0) and a constant N/P ratio of 24, so as to determine the optimum growth rate for . After deciding the best pH conditions, the N/P ratio was selected considering the concentrations of DIN and DIP found in shrimp ponds. The next test thus used four different N/P ratios: 3:2, 6:1, 24:1 and 96:1. The results showed that the maximum carrying capacity ( was lower than that in the environments of individual cultivation. This finding indicated that the inhibition parameter of interspecies competition between these two algae could be significantly influenced by pH conditions. The maximum rate of competition inhibition of occurred at pH 8.5, with an inhibition parameter value of 5.7447, and the value of inhibition of was 3.4886, also at pH 8.5. Therefore, the nutrient limitation study was set at a constant pH of 8.5 against the four N/P ratios. The results of that experiment showed that the inhibition parameter of interspecies competition could also be significantly influenced by N/P ratio. Maximum inhibition of occurred at the N/P ratio 96:1, with an inhibition value of 9.2063, whereas the inhibition value for was 3.4886; hence, the inhibition effect of was stronger than that of . Thus, we propose that the optimum growth conditions of can be achieved at pH 8.5 and N/P ratio 96:1. In shrimp culture, the pH level and N/P ratio of the culture water can be accordingly adjusted to control algal growth; this approach can be applied for the purposes of controlling the production process, to assist with accurate water technology research, or to control eutrophication the culture water.

Abstract:In July 2014, the widely cultured pond crab (commonly called Chinese swimming crab) had a disease outbreak along the coasts of Tianjin and Hebei provinces, China, resulting in massive mortalities. Symptoms of two different illnesses appeared in the crabs. One was white muscle disease (WMD), with its symptoms being anorexia, slowed movement, and the muscles turning opaque white and finally emulsifying into a toothpaste-like consistency. Mortality among the crabs displaying WMD was 60%-70%. The other was milky disease (MD), which displayed as a softened carapace, cessation of feeding, slowed movement, and the muscles similarly turning opaque white but then becoming watery (‘milky') and finally shrinking; the coelomic fluid and the deteriorated tissue leaked from the body and had not coagulated. Mortality among the crabs displaying MD exceeded 90%. According to the characteristics of the epidemic, and histopathological and ultrastructural analyses, the supposed cause of WMD was infection with a microsporidian. The parasite was ellipsoid, measured (1544±250) nm, and had the following structure: pole filament (8 or 9 laps), spore wall [thickness (130±22) nm], cilium (length 120-500 nm), polaroplast [(726±200) nm], pole tube, anchoring disk, plasma membrane and nucleus. The MD outbreak was caused by sp., a monad, sized 8-10 μm, with an oval shape, and wrinkled and raised surface; its basic structure was a cell wall, multiple nuclei, cilium and other structures. Histopathological examination showed that the two kinds of diseased crabs had significantly different muscle lesions. The crabs associated with WMD had many microsporidia adhering to their muscle fibers, causing the fibers to break, collapse and necrosis, finally reducing them to muscle bundles. SEM observations showed that the muscle cells became fused and acquired a large number of vacuoles, while the mitochondria and endoplasmic reticulum disintegrated and disappeared. In comparison, the gap in the muscle bundles was bigger in cases of MD, in which numerous parasites occupied or adhered to the muscle fibers, resulting in the fibers rupture, disintegration, and resolvable necrosis. Electron microscopy also confirmed that the two different parasites could infect the same tissue in the same crab, and both parasites could enter the disintegrated cytoplasm after muscle cell fusion. Since the initial epidemic of these diseases, other occurrences of WMD and MD have tended to cause a higher level of mortality, causing concern as significant diseases currently effecting the crab-farming industry.

Abstract:Streptococcosis in cultured tilapia ( has increased during the past decade in China. In this study, a total of 248 strains were isolated from diseased tilapia, collected in the major tilapia-culturing area of China, during 2007-2015. The aim of the study was to analyze the characteristics of the 248 tilapia strains. In molecular characterization assays, four genotypic categories comprised of molecular serotype, multilocus sequence typing (MLST), virulence genes profiling patterns and prophage genotype, were used to analyze the genotypic diversity of these S. agalactiae strains were of molecular serotype Ia, and the remaining 19 were of type Ib. MLST revealed that all the serotype Ia strains were ST7, and all the serotype Ib strains were ST261. The results of virulence genes detection indicated that the 229 Ia-ST7 strains have the same genotype of virulence genes (V1), which carried 9 virulence genes (9/11: 81.8%), except that virulence genes were negative for them. The 19 Ib-ST261 strains have the same genotype of virulence genes (V2), which possessed 6 virulence genes (6/11: 54.5%) detected by PCR. Prophage detection suggested that the 229 Ia-ST7 strains could be divided into two genotypes: type P1 (36 strains) and P2 (193 strains). All the Ib-ST261 strains were negative for 10 prophage genes, and they belonged to type P3. According to the four molecular typing methods, all 248 strains could be divided into three genotypes: namely, Ia-ST7-V1-P1, Ia-ST7-V1-P2, and Ib-ST261-V2-P3. Notably, the predominant strains of in China had shifted from type Ia-ST7-V1-P1 to type Ia-ST7-V1-P2 during 2010-2011. In addition, the type Ia-ST7-V1-P1 strain was predominant before 2011, and the type Ia-ST7-V1-P2 strain predominated thereafter. In conclusion, the strains collected from tilapia in China showed low genetic diversity. Through analyses of the characteristics of the strains isolated from deceased tilapia in the major tilapia-culturing area of China, we can deduce that the transmission of relies on the import of either new tilapia stock or culture water.

Abstract:White spot syndrome and infectious hypodermal and haematopoietic necrosis are primary viral diseases of prawns. Outbreaks of white spot syndrome virus (WSSV) and infectious hypodermal and haematopoietic necrosis virus (IHHNV) arise worldwide and have caused serious economic losses in recent years, including in China. Some researchers found that, given the same conditions, WSSV-infected prawn display lower levels of mortality than IHHNV-infected prawn. Our laboratory previously demonstrated that IHHNV capsid protein (CP) has the ability to combine with NM23 protein in prawn gill membrane. NM23, a kind of nucleoside diphosphate kinase (NDPK), has various biological functions and is located mainly in the cytoplasm and nucleus, but is also evident in the cell membrane. We conducted an experiment to investigate whether WSSV and IHHNV compete with NM23 protein receptor sites on the cell membrane of shrimp gill tissue. First, we purified the total WSSV proteins using sucrose gradient centrifugation, and then we used a virus overlay protein binding assay (VOPBA) to detect protein-protein interaction between the total proteins and NM23. Next, three suspected positive proteins were selected by the combined LC-MS/MS technique: WSSV013, Wsv497 and Wsv035. Wsv035, also known as VP110, is one of the capsule membrane proteins of WSSV and is encoded by . It contains a membrane structure domain and a RGD locus (namely, the Arg-Gly-Asp motif). Some scholars, using fluorescence assay and competitive inhibition tests, found Wsv035 could stick to the host cell: Wsv035 RGD can form RGDT peptides at the cell surface and it plays a role in WSSV infection. There have been few studies of Wsv497 and WSSV013 proteins, however, and thus their distribution, structure and functions are not yet clear. Therefore, we constructed prokaryotic expression vectors of these three proteins, for purifying WSSV013, Wsv497 and Wsv035 in order to interact with NM23 protein, using VOPBA and co-immunoprecipitation, respectively. The results showed that Wsv497 and Wsv035 have the ability to interact with NM23, while WSSV013 showed an invalid effect on NM23. Accordingly, we surmise that WSSV and IHHNV have the ability to compete with NM23 protein receptors on the cell membrane of prawn gill tissue. This study adds to theories about the mechanism of competition between WSSV and IHHNV for receptor sites on the cell membrane of prawn gill tissue, and lays a foundation for further studies of protein interactions in the context of WSSV and prawn culture.

Abstract:Paralytic shellfish toxins (PSTs) are potent neurotoxins produced by a variety of marine dinoflagellates, such as . PSTs can accumulate and be passed into the food chain through the filter feeding of toxin-producing algae by bivalve mollusks. The consumption of contaminated shellfish can cause poisoning in humans, with symptoms including nausea, vomiting, diarrhea, numbness or tingling in the extremities, and even death as a result of respiratory failure. is widely distributed along the coast of China, and often forms large algal blooms in the waters surrounding Nanhuangcheng Island, the Zhoushan area, and the port city of Xiamen. Many controlled feeding experiments with shellfish have investigated the accumulation and fate of PSTs toxins in the marine food web. Comparative studies of PST profiles often found significant differences between the causative dinoflagellates and contaminated bivalve mollusks, thus indicating active metabolism of the toxin in some shellfish. The scallops are known to become much more toxic for consumption than mussels grown under the same environmental conditions, and several months are required until the toxicity drops to an acceptably safe level. Conversely, levels of toxicity in the short-necked clam and oysters are generally low, and the toxicity can decline as quickly as in the mussel. The metabolic interconversion of incorporated toxins achieved by enzymatic and chemical reactions in the tissues of bivalves, and different uptake and depuration kinetics of the individual toxins, may each contribute to differences in the PST profiles.In this study, we exposed scallop directly to toxin-producing algae in order to characterize the formation of PSTs in . Furthermore, we followed PST accumulation and metabolic biotransformation in different organs of the scallop using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). The results showed that the AT5-3 strain of in separate areas of the South China Sea were stable both in their growth and toxin production. The composition of the toxin consisted mainly of gonyautoxin-1 and-4, with a cell toxin-production of 7.95 fmol/cell. Rapid accumulation of the PST was observed, while total accumulation efficiency reached 84.4%. The highest accumulation concentration was 1903 μg STX.2HCl-eq/kg. The toxin-accumulation ability of the different tissues was ordered as viscera>gonad>pallium>muscle. Moreover, the viscera showed the strongest PST metabolic ability and was also the target organ. In addition, the proportion of PST in bivalves differed from the profile of the dinoflagellate that produces the toxins. has a strong ability to perform biological transformation. The main transformation pathway was: N-sulfocarbamoy­lgonyautoxin-2 (C1) → gonyautxin-2 (GTX2)/decarbamoylgonyautoxin-2 (dcGTX2), and N-sulfocarbamoylgony­autoxin-3 (C2) → gonyautxin-3 (GTX3). In this experiment, showed characteristically rapid PST-accumulation and slow PST-metabolism; meanwhile the high ratio of the toxin component GTX1 and 4 and NEO resulted in significantly increased risk of PSTs in this scallop. This study contributes to the scientific evaluation of the formation mechanisms of PSTs, thereby providing scientific support for the monitoring of PSTs in China.