Use of antioxidant enzymes of clam Ruditapes philippinarum as biomarker to polycyclic aromatic hydrocarbon pollution

The typical organic pollutant polycyclic aromatic hydrocarbon(PAH) anthracene was selected as a contaminant to investigate its eff ects on the activities of superoxide dismutase(SOD), catalase(CAT) and glutathione peroxidase(GSH-Px) in the clam Ruditapes philippinarum. The results show that SOD, CAT and GSH-Px had diff erent induction and inhibition reactions to anthracene stress, and that three diff erent organs in R. philippinarum(visceral mass, muscle tissue and mantle) had diff erent sensitivities to anthracene stress. This study suggest that SOD activities of the visceral mass, CAT activitities of the mantle and the visceral mass, and GSH-Px activity of the muscle tissue could be used as sensitive indicators of anthracene stress in R. philippinarum.     

Effects of ocean acidification on the metabolic rates of three species of bivalve from southern coast of China

Oceanic uptake of anthropogenic carbon dioxide results in a decrease in seawater pH, a process known as "ocean acidification". The pearl oyster Pinctada fucata, the noble scallop Chlamys nobilis, and the green-lipped mussel Perna viridis are species of economic and ecological importance along the southern coast of China. We evaluated the effects of seawater acidification on clearance, respiration, and excretion rates in these three species. The animals were reared in seawater at pH 8.1 (control), 7.7, or 7.4. The clearance rate was highest at pH 7.7 for P. fucata and at pH 8.1 for C. nobilis and P. viridis. The pH had little effecton the respiration rate of P. fucata and P. viridis. In contrast, the respiration rate was significantly lower atpH 7.4 in C. nobilis. The excretion rate was significantly lower at pH 7.4 than pH 8.1 for all species. Theresults indicate that the reduction in seawater pH likely affected the metabolic process (food intake, oxygenconsumption, and ammonia excretion) of these bivalves. Different species respond differently to seawateracidification. Further studies are needed to demonstrate the exact mechanisms for this effect and evaluateadaptability of these bivalves to future acidified oceans.     

Manila clam Venerupis philippinarum as a biomonitor to metal pollution

The Manila clam Venerupis philippinarum is a good biomonitor/bioindicator to marine metal pollution and is frequently used in aquatic toxicology. Two dominant pedigrees (white and zebra) of clam are distributed in the Bohai Sea; however, little attention has been paid to potential biological differences between these two pedigrees. In this study, we tested the sensitivity of both pedigrees to marine metal (cadmium and zinc) pollution biomonitoring and marine environmental toxicology. Results demonstrate significant biological differences in gills of white and zebra clams based on metabolic profiles and antioxidant enzyme activities. In addition, we found that hypotaurine, malonate and homarine were relatively high in white clam gills, while alanine, arginine, glutamate, succinate, 4-aminobutyrate, taurine and betaine were high in zebra clam gills. Zebra clam gills were also more sensitive to a mixture of Cd and Zn, as shown by antioxidant enzyme activities and metabolic profiles, but white clam gills could accumulate more Zn. Therefore, we suggest that the white pedigree can be used as a biomonitor to marine Zn pollution, whereas the zebra pedigree can be used for toxicology studies on Cd and Zn mixed pollution.     

Observations on shell growth and morphology of the bivalve Ruditapes philippinarum

The bivalve Ruditapes philippinarum is not only one of the main mollusc species in the west Pacific Ocean, but also one of the main species for aquaculture in China. Knowledge of growth of the R. philippinarum shell will improve our understanding of the shell as an environmental archive. It is also useful for the aquaculture of R. philippinarum. In this research, a hanging box culture method was introduced in the culture of R. philippinarum. The bivalves were cultured for 126 days, from March 31 to August 3, 2002. The average growth rates of shell width, height, and thickness were 0.069, 0.046, and 0.032 mm/d, respectively. The mean increase of average individual wet mass was 0.028 g/d. The largest growth rates of both shell and average individual wet mass occurred in June, indicating that water temperature and bivalve reproduction were 2 important factors. The shell morphology underwent significant changes with shell growth. The ratio of height to thickness (value of B/C) shifted 1.58 in the first 60 days to 1.54 in the last 30 days, which was resulted from the change in major shell growth direction. Periodic changes in the B/C ratio led to corrugated shell form, which could be used to determine the age of the shell.     

Use of antioxidant enzymes of clam <Emphasis Type="Italic">Ruditapes philippinarum</Emphasis> as biomarker to polycyclic aromatic hydrocarbon pollution

The typical organic pollutant polycyclic aromatic hydrocarbon (PAH) anthracene was selected as a contaminant to investigate its effects on the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) in the clam Ruditapes philippinarum. The results show that SOD, CAT and GSH-Px had diff erent induction and inhibition reactions to anthracene stress, and that three diff erent organs in R. philippinarum (visceral mass, muscle tissue and mantle) had diff erent sensitivities to anthracene stress. This study suggest that SOD activities of the visceral mass, CAT activitities of the mantle and the visceral mass, and GSH-Px activity of the muscle tissue could be used as sensitive indicators of anthracene stress in R. philippinarum.     

Mass Selection for Fast Growth in the Third Generation of the Orange Line of the Manila Clam Ruditapes philippinarum

The Manila clam Ruditapes philippinarum is natively distributed along the Pacific coast of Asia, where it is one of the most important bivalve species for local fisheries. This species has been intentionally introduced due to its high adaptability to various coastal environments and is currently widely cultivated in Europe and North America. We have initiated a selective breeding program to improve the growth rate of the Manila clam since 2007, and a full-sib family with orange shell color was selected. This family features high survival but slow growth. Thereafter, two generations of mass selection were conducted in this family by 10% upward selection for faster growth. In 2011, three types of lines were produced by selecting the breeder from the second-generation of selected lines. These lines are SS(subject to three generations of selection for growth in shell length), SC(selected for the second but not the third generation), and C(randomly sampled individuals from the national population). The genetic parameters of the shell length including hR2(realized heritability), SR(selection response), and GG(genetic gain) were analyzed. Results showed that the shell length of the SS line was significantly larger than those of the SC and C lines at all developmental stages(P 0.05). For the larval stage, the values of hR2, SR, and GG were 0.42, 0.73, and 6.66, respectively. For the juvenile stage, the values of hR2, SR, and GG were 0.40, 0.69, 21.76, respectively. For grow-out stage, the values of hR2, SR, and GG were 0.48, 0.83, and 18.22, respectively. The results of hR2 indicated the good potential of the SS line in selective breeding. The level of GG improvement was encouraging and consistent with the previous expectations. No inbreeding depressions in the shell length were observed in the SS and SC lines at all sampling days. All of the results indicate that maintaining selection pressure in successive generations may be effective in the selective breeding program of this family.     

Selection of diet for culture of juvenile silver pomfret, Pampus argenteus

Juvenile silver pomfret, Pampus argenteus, was grown in culture tanks for 9 weeks on four different diets, and their effects on fish growth, digestive enzyme activity, and body composition were assessed. The feeding regime was as follows: Diet 1: fish meat; Diet 2: fish meat+artificial feed; Diet 3: fish meat+artificial feed+Agamaki clam meat; Diet 4: fish meat+artificial feed+Agamaki clam+copepods. The greatest weight gain was associated with Diet 4, while the lowest weight gain was associated with Diet 1. No significant difference was observed in weight gain between fish receiving Diet 2 and Diet 3. Specific growth rate followed similar trends as weight gain. The feed conversion ratio (FCR) of fish fed Diet 1 was significantly higher than the other fish groups, but no significant differences were observed in FCRs of fish fed Diet 2, Diet 3 or Diet 4. There was also no significant difference in the hepatosomatic index (HSI) between the four diets. For fish that received Diets 2-4, containing artificial feed, higher protease activities were detected. A higher lipid content of the experimental diets also significantly increased lipase activities and body lipid content. No significant differences in amylase activity or body protein content were found between Diets 1-4. In conclusion, a variety of food components, including copepods and artificial feed, in the diet of silver pomfret significantly increased digestive enzyme activity and could improve growth performance.     

Growth performance of larval and juvenile manila clam (<Emphasis Type="Italic">Ruditapes philippinarum</Emphasis>) from divergently selected individuals of a full-sib family

In this study, the method of divergent selection was employed to test the larval and juvenile growth performance within a full-sib family of Manila clam Ruditapes philippinarum. The 10% largest and 10% smallest clam individuals (on the basis of shell length) of a full-sib family were selected as parents for the fast and slow growing lines, respectively. The difference in shell length was significant among the three lines (fast, control, and slow) tested. The sequence of shell length were fast line > control line > slow line. The responses to selection, realized heritability, and genetic gain were 0.06%–0.81%, 0.04%–0.47% and 0.58%–18.89% in the fast direction, respectively; and were 0.14%–1.27%, 0.08%–0.73%, and 0.31%–49.03% in the slow direction, respectively. The results suggested that there was a large portion of additive genetic variance affecting the growth in the full-sib family. Selection in the fast direction within the full-sib family would greatly improve the growth of R. philippinarum.     

Isozyme analysis on the populations ofRuditapes philippinarum

Horizontal starch gel electrophoresis was used to investigate the genetic structure of six populations of Ruditapes philippinarum in the coast of China. Seven enzymes revealed eleven putative loci, and seven of them were polymorphic, PGM^＊, MDH-1^ ＊, MDH-3^＊, LAP-1^＊, LAP-2^＊, MPI-1^＊ and MPI-2^＊. The proportions of polymorphic loci of R. philippinarum populations varied from 0.454 5 to 0.636 4. The values of observed and expected heterozygosities were from 0.039 4 to 0.154 5 and from 0.111 1 to 0.238 5, respectively. Nei＇s genetic distance varied from 0.003 3 to 0.025 3 with an average of 0.013 5. It is suggested that the genetic diversity of R. philippinarum was high.     

3种贝类糖胺聚糖的粗提及含量比较

以企鹅珍珠贝、菲律宾蛤仔和锯齿巴非蛤全脏器为原料,经酶解、离心去蛋白、醇沉、分离、洗涤、干燥,得糖胺聚糖(GAG)粗制品,分别测定并比较3种贝类的GAG的含量和得率。结果表明,企鹅珍珠贝的GAG质量分数为22.35%,菲律宾蛤仔为18.69%,锯齿巴非蛤为20.36%,由此得出企鹅珍珠贝的GAG的含量比其他两种贝类高,是较好的GAG来源。     

Inbreeding depression on growth and survival of full-sib family of Manila clam (<Emphasis Type="Italic">Ruditapes philippinarum</Emphasis>)

In present study, the inbreeding depression (ID) of growth and survival of Manila clam (Ruditapes philippinarum) was investigated at larval and juvenile stages. Nine inbred families (A ₂, B ₂, C ₂, D ₂, E ₂, F ₂, G ₂, H ₂ and I ₂) were established by mating within nine full-sib families with expected inbreeding coefficient of 0.25. Inbred families showed significant differences in shell length and hatching rate of D-larvae (straight-hinged larvae). The larvae of the nine inbred families grew slower than those of control group (CG), and their ID value ranged from 0.81% ± 6.09% to 16.10% ± 1.49%. The ID value of larval survival rate varied between 27.47% ± 9.36% and 70.50% ± 13.66%. The ID was also detected for juvenile growth in A ₂, B ₂, C ₂, and D ₂, which ranged from 4.60 ± 2.21 to 17.71 ± 7.73. The A ₂ family maintained the highest juvenile survival rate, whereas the other inbred families exhibited ID values varying between 62.79% ± 4.54% and 96.14% ± 0.87%. The linear relationship of estimated ID between growth and survival was negatively correlated (R = ?0.434, P < 0.05). The results of this study suggested that the ID of growth was common at the larval stage but was less prevalent at juvenile stage. In contrast, the ID of survival increased from larval to juvenile stage. A better understanding of the effect of inbreeding may aid to selective breeding of Manila clam.     

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.     

Food sources of the Manila clam Ruditapes philippinarum in intertidal areas: evidence from stable isotope analysis

Based on stable isotope analysis, we characterized the dietary regime of the Manila clam Ruditapes philippinarum inhabiting intertidal areas along the Liaodong Peninsula, Northern China. Samples, including particulate organic matter (POM; n =30), benthic microalgae (BMI; n =30) and R. philippinarum (n =60), were collected from six sampling sites displaying the same ecological conditions. Of the two primary food sources, POM was more depleted in δ ¹³C (?20.61‰ to ?22.89‰) than BMI was (?13.90‰ to ?16.66‰). With respect to ¹⁵N, BMI was more enriched (2.90‰ to 4.07‰) than POM was (4.13‰ to 5.12‰). The δ ¹³C values of R. philippinarum ranged from ?18.78‰ to ?19.35‰ and the δ ¹⁵N values from 7.96‰ to 8.63‰, which were intermediate between the POM and BMI values. In a two-source isotope mixing model, we estimated the relative contributions of POM and BMI to the diet of R. philippinarum to be 74.2% and 25.8%, respectively. We conclude that R. philippinarum feeds mainly on POM, and BMI is also an important supplemental food source in intertidal areas.     

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.     

持续高温对吉富罗非鱼幼鱼消化酶和溶菌酶活力的影响

通过人工控温,对吉富罗非鱼（GIFT Oreorhromis niloticus）幼鱼实施34℃持续高温处理,研究持续高温对吉富罗非鱼幼鱼消化酶和溶菌酶活力的影响。结果表明：持续高温对吉富罗非鱼幼鱼消化酶活力的影响有统计学意义（P〈0．05）,对溶菌酶活力影响不具统计学意义（P〉0.05）;随着时间的推进,脂肪酶、淀粉酶活力增加,差异有统计学意义（P〈0．05）,溶菌酶活力增加,但差异无统计学意义（P〉0.05）。表明持续高温能一定程度地提高罗非鱼的消化能力和免疫功能。     

Annual variations of biogenic element contents of manila clam (Ruditapes philippinarum) bottom-cultivated in Jiaozhou Bay,China

Manila clam(Ruditapes philippinarum) was monthly sampled from its benthic aquaculture area in Jiaozhou Bay from May 2009 to June 2010. The annual variations of major elemental composition, organic content, fatness and element ratio of Manila clam were examined. The element removal effect of clam farming in Jiaozhou Bay was analyzed based on natural mortality and clam harvest. The results indicated that the variation trend of carbon content in shell(Cshell) was similar to that in clam(Cclam). Such a variation was higher in summer and autumn than in other seasons, which ranged from 9.10 ± 0.13 to 10.38 ± 0.09 mmol g-1 and from 11.28 ± 0.29 to 12.36 ± 0.06 mmol g-1, respectively. Carbon content of flesh(Cflesh) showed an opposite variation trend to that of shell in most months, varying from 29.42 ± 0.05 to 33.64 ± 0.62 mmol g-1. Nitrogen content of shell(Nshell) and flesh(Nflesh) changed seasonally, which was relatively low in spring and summer. Nshell and Nflesh varied from 0.07 ± 0.009 to 0.14 ± 0.009 mmol g-1 and from 5.46 ± 0.12 to 7.39 ± 0.43 mmol g-1, respectively. Total nitrogen content of clam ranged from 0.50 ± 0.003 to 0.76 ± 0.10 mmol g-1 with a falling tend except for a high value in March 2010. Phosphorus content of clam(Nclam) fluctuated largely, while phosphorus content of shell(Pshell) was less varied than that of flesh(Pflesh). Pshell varied from 0.006 ± 0.001 to 0.016 ± 0.001 mmol g-1; while Pflesh fluctuated between 0.058 ± 0.017 and 0.293 ± 0.029 mmol g-1. Pclam ranged from 0.015 ± 0.002 to 0.041 ± 0.006 mmol g-1. Carbon and nitrogen content were slightly affected by shell length, width or height. Elemental contents were closely related to the reproduction cycle. The removal amounts of carbon, nitrogen and phosphorus from clam harvest and natural death in Jiaozhou Bay were 2.92×104 t, 1420 t and 145 t, respectively. The nutrient removal may aid to reduce the concentrations of nitrogen and phosphorus, the main causes of eutrophication, and to maintain the ecosystem health of Jiaozhou Bay.