Effect of water temperature on digestive enzyme activity and gut mass in sea cucumber Apostichopus japonicus （Selenka）, with special reference to aestivation

The effect of water temperature on gut mass and digestive enzyme activity in sea cucumber Apostichopus japonicus, including relative gut mass (RGM), amylase, lipase, pepsin and trypsin activities were studied at temperatures of 7, 14, 21, and 28°C over a period of 40 days. Results show that RGM significantly decreased after 40 days at 21°C and markedly decreased over the whole experiment period at 28°C; however, no significant effect of duration was observed at 7 or 14°C. At 14°C, trypsin activity significantly decreased over 10 and 20 days, then increased; amylase and trypsin activity significantly decreased after 40 days at 28°C. However, no significant effect of duration was found on amylase, pepsin or trypsin activities in the other temperature treatment groups. At 28°C, lipase activity peaked in 20 days and then markedly decreased to a minimum at the end of the experiment. On the other hand, pepsin activity at 28°C continuously increased over the whole experimental period. Principle component analysis showed that sea cucumbers on day 40 in the 21°C group and in the previous 20 days in the 28°C group were in the prophase of aestivation. At 28°C, sea cucumbers aestivated at 30–40 days after the start of the experiment. It is concluded that the effect of temperature on the digestion of A. japonicus is comparatively weak within a specific range of water temperatures and aestivation behavior is accompanied by significant changes in RGM and digestive enzyme activities.     

Growth, metabolism and physiological response of the sea cucumber, Apostichopus japonicus Selenka during periods of inactivity

The growth, metabolism and physiological response of the sea cucumber, Apostichopus japonicus, were investigated during periods of inactivity. The body weight, oxygen consumption rate (OCR), activities of acidic phosphatase (ACP), alkaline phosphatase (AKP), catalase (CAT) and superoxide dismutase (SOD), and content of heat shock protein 70 (Hsp70) in the body wall and coelomic fluid of A. japonicus were measured during starvation, experimental aestivation and aestivation. The results showed that the body weight of sea cucumber in the three treatments decreased significantly during the experimental period (P < 0.05). The OCR of sea cucumber reduced in starvation and experimental aestivation treatments, but increased gradually in natural aestivation treatment. The activities of ACP and AKP of sea cucumber decreased gradually in all treatments, whereas those of SOD and CAT as well as Hsp70 content decreased in the starvation and experimental aestivation treatments and increased in natural aestivation treatment. The sea cucumber entered a state of aestivation at 24℃. To some extent, the animals in experimental aestivation were different from those in natural aestivation in metabolism and physiological response. These findings suggested that the aestivation mechanism of A. japonicus is complex and may not be attributed to the elevated temperature only.     

Relation of body weight and food consumption to metabolic rate of juvenile Japanese sea bass,Lateolabrax japonicus

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Effects of Hanseniaspora opuntiae C21 on the growth and digestive enzyme activity of juvenile sea cucumber Apostichopus japonicas

The effects of a diet containing Hanseniaspora opuntiae C21 on growth and digestive enzyme activity were estimated in juvenile Apostichopus japonicus. Groups of sea cucumbers were fed diets containing H. opuntiae C21 at 0(control), 10 4, 10 5, and 10 6 CFU(colony-forming units)/g feed. Results showed that after 45 d the specific growth rate(SGR) of sea cucumbers fed a C21-supplemented diet at 10 4 CFU/g feed was significantly higher than that of the control( P 0.05). Intestinal trypsin and lipase activities were significantly enhanced by C21 administration at 10 4 and 10 5 CFU/g feed compared with the control( P 0.05). After feeding for 23–42 d, C21 was demonstrated by denaturing gradient gel electrophoresis to be present in the intestine of sea cucumbers. In addition, after feeding the C21-supplemented diets for 15 d, the sea cucumbers were switched to an unsupplemented diet and C21 was confirmed to be capable of colonizing the intestine for at least 31 d after cessation of feeding. In conclusion, C21 was shown to successfully colonize the intestine of juvenile A. japonicus via dietary supplementation, and improve growth and digestive enzyme activity.     

Optimization of stocking density for the sea cucumber, Apostichopus japonicus Selenka, under feed-supplement and non-feed-supplement regimes in pond culture

Optimal stocking densities were investigated for the sea cucumber Apostichopus japonicus Selenka under feed-supplement and non-feed-supplement regimes in net enclosures for 333 d. Substantial weight loss occurred during the aestivation phase (AE). Decreased growth rates were also observed during the winter phase (WT). In contrast, sea cucumbers showed rapid growth during the spring (SP) and autumn (AU) phases. Feeding regimes considerably influenced the growth performance, i.e., sea cucumbers grew faster under feed-supplement regime than under non-feed-supplement regime (P < 0.05). The average survival rates of sea cucumbers under feed-supplement regime were higher than those under non-feed-supplement regime for both the autumn phase and spring phase, but the differences were only significant for the latter phase (P < 0.05). The fitted B-N curves showed that the optimal stocking densities, in terms of net production, were 22.3 ind. m-2 for feed-supplement regime and 14.1 ind. m-2 for non-feed-supplement regime.     

Effect of Bacillus baekryungensis YD13 supplemented in diets on growth performance and immune response of sea cucumber (Apostichopus japonicus)

The effect of a potential probiotic on the growth performance and immune response of sea cucumber(Apostichopus japonicus) was investigated. Bacillus baekryungensis YD13 isolated from sea cucumber culturing ponds was added to sea cucumber basal feed as a probiotic in different doses(0, the control; 1×104(YD134), 1×106(YD136) and 1×108(YD138) CFU g-1 of diet), and administered orally to A. japonicus(initial mean wet weight 5.44 g ± 0.17 g). The sea cucumbers were fed in 20 aquaria, 5 each treatment, for 60 d. At the end of growth trial, 20 sea cucumbers from each treatment were challenged with Vibrio splendidus. A. japonicus in YD134 and YD136 exhibited significantly better growth performance than control(P 0.05). Five non-specific immune parameters including lysozyme, acid phosphatase, alkaline phosphatase, superoxide dismutase and catalase in coelomic fluid were measured to evaluate the immune response of A. japonicus to the probiotics. Results showed that all parameters were significantly improved when YD11 was supplemented in the dose of 1×106 CFU g-1(P 0.05). The cumulative incidence and mortality after the Vibrio splendidus challenge decreased significantly in sea cucumbers of YD136. Accordingly, 1×106 CFU g-1 of YD13 in diet was recommended for the growth promotion and immune enhancement of A. japonicus.     

Effects of dietary β-glucan and glycyrrhizin on non-specific immunity and disease resistance of the sea cucumber (<Emphasis Type="Italic">Apostichopus japonicus</Emphasis> Selenka) challenged with <Emphasis Type="Italic">Vibrio splendidus</Emphasis>

Sea cucumbers, Apostichopus japonicus Selenka, were fed diets containing non-immunostimulant (basal diet), 0.2% β-glucan and 0.02% glycyrrhizin in a recirculatory water system for 45 days, and subsequently challenged with Vibrio splendidus by injection at 1.0×10⁸ cfu / sea cucumber for 15 days. Phagocytic capacity (PC), intracellular superoxide anion production (ISAP), lysozyme (LSZ) activity and superoxide dismutase (SOD) activity in the coelomic fluid were analyzed on the 0^th, 5^th, 10^th and 15^th days after injection. Results showed that after the 45-day feeding period, PC, ISAP, LSZ activity and SOD activity in sea cucumbers fed with dietary β-glucan or glycyrrhizin were significantly higher than in those fed with the basal diet. On the 5th day after infection, all the immune parameters examined in the sea cucumbers injected with V. splendidus decreased in value significantly. On the 15^th day, PC, ISAP and LSZ activity returned to levels similar to those on the 0^th day. For the sea cucumbers injected with saline, there were no significant differences in all the immune parameters examined and in the cumulative morbidity during the 15-day challenging trial. After injecting with V. splendidus, the cumulative morbidity of sea cucumbers fed with the basal diet was significantly higher than those fed with dietary β-glucan or glycyrrhizin when challenged with V. splendidus challenged sea cucumber fed with the basal diet was significantly higher than those fed with dietary β-glucan or glycyrrhizin. There was no significant difference in cumulative morbidity between the dietary β-glucan and glycyrrhizin treatments over time.     

Analysis of fatty acid composition of sea cucumber Apostichopus japonicus using multivariate statistics

Fatty acids （FAs） provide energy and also can be used to trace trophic relationships among organisms. Sea cucumber Apostichopus japonicus goes into a state of aestivation during warm summer months. We examined fatty acid profiles in aestivated and non-aestivated A. japonicus using multivariate analyses （PERMANOVA, MDS, ANOSIM, and SIMPER）. The results indicate that the fatty acid profiles of aestivated and non-aestivated sea cucumbers differed significantly. The FAs that were produced by bacteria and brown kelp contributed the most to the differences in the fatty acid composition of aestivated and non- aestivated sea cucumbers. Aestivated sea cucumbers may synthesize FAs from heterotrophic bacteria during early aestivation, and long chain FAs such as eicosapentaenoic （EPA） and docosahexaenoic acid （DHA） that produced from intestinal degradation, are digested during deep aestivation. Specific changes in the fatty acid composition of A. japonicus during aestivation needs more detailed study in the future.     

Transcriptional changes in epigenetic modifiers associated with gene silencing in the intestine of the sea cucumber, Apostichopus japonicus (Selenka), during aestivation

The sea cucumber, Apostichopus japonicus, undergoes aestivation to improve survival during periods of high-temperature. During aestivation, the metabolic rate is depressed to reduce the consumption of reserved energy. We evaluated the role of epigenetic modification on global gene silencing during metabolic rate depression in the sea cucumber. We compared the expression of epigenetic modifiers in active and aestivating sea cucumbers. The expression of three genes involved in DNA methylation and chromatin remodeling (DNA (cytosine-5)-methyltransferase 1, Methyl-CpG-binding domain protein 2), and Chromodomain-helicase-DNA-binding protein 5) was significantly higher during aestivation (Days 20 and 40). Similarly, we observed an increase in the expression of genes involved in histone acetylation (Histone deacetylase 3) and Histone-binding protein RBBP4) during the early (Days 5 and 10) and late phases (Days 20 and 40) of aestivation. There was no change in the expression of KAT2B, a histone acetyltransferase. However, the expression of histone methylation associated modifiers (Histone-arginine methyltransferase CARMER and Histone-lysine N-methyltransferase MLL5) was significantly higher after 5 d in the aestivating group. The results suggest that the expression of epigenetic modifiers involved in DNA methylation, chromatin remodeling, histone acetylation, and histone methylation is upregulated during aestivation. We hypothesize that these changes regulate global gene silencing during aestivation in A. japonicus.     

Ecological effects of co-culturing the sea cucumber <Emphasis Type="Italic">Apostichopus japonicus</Emphasis> with the Chinese white shrimp <Emphasis Type="Italic">Fenneropenaeus chinensis</Emphasis> in an earthen pond

Using net enclosures in an earthen pond, we established three culture treatments with the sea cucumber Apostichopus japonicus and the Chinese white shrimp Fenneropenaeus chinensis: monoculture of sea cucumbers (C), monoculture of shrimp (S), and co-culture of the two species (CS). We measured levels of suspended particulate matter in the water column; total organic matter, total organic carbon, total nitrogen, and carbon/nitrogen ratios in both settling particles and the sediment; and chlorophyll a levels in the sediment. We then compared these variables between the three treatments. We also examined growth, survival, and yield of the two species in the different treatments. From June to September, the mean monthly suspended particulate matter sedimentation rates in the CS and S treatments were significantly (P<0.05) greater than those in the C treatment. From August to November, the mean monthly total organic matter, total organic carbon, total nitrogen, and chlorophyll a contents in the sediment in the CS and S treatments were significantly ( P <0.05) greater than those in the C treatment. Final wet weight, specific growth rate, survival rate, and total yield of sea cucumbers in co-culture were all significantly greater than those of sea cucumbers in monoculture. There were no significant differences among any of these variables for shrimp reared in the two systems. The bioturbation of the sediment and fecal production of the shrimp likely supplied natural food for the sea cucumbers. Co-culture of the two species is a viable option for increasing yield per unit area, maximizing use of the water body, and diversifying crop production.     

Effects of rearing temperature and density on growth, survival and development of sea cucumber larvae, Apostichopus japonicus (Selenka)

In laboratory conditions, effects of rearing temperature and stocking density were examined on hatching of fertilized egg and growth of auricularia larvae of Apostichopus japonicus respectively. Data series like larval length and density, metamorphic time, and survival rate of the larvae were recorded. Statistics showed that for A. japonicus, survival rate (from fertilized egg to late auricularia) decreased significantly with the increasing rearing temperature (P<0.05). At different temperatures SGR was statistically significant as well (P<0.05) from day 1, and maximal SGR was found on day 9 at 24°C (159.26±3.28). This study clearly indicated that at low temperature (<24°C), metamorphic rate was remarkably higher than at higher temperature (>26°C). Hatching rate was significantly different between 0.2–5 ind./ml groups and 20–50 ind./ml groups. Rearing larvae at the higher density had the smaller maximal-length, whereas needed longer time to complete metamorphosis. This study suggested that 21°C and 0.4 ind./ml can be used as the most suitable rearing temperature and stocking density for large -scale artificial breeding of A. japonicus’s larvae.     

Rheological and structural properties of sea cucumber Stichopus japonicus during heat treatment

Changes in tissue structure, rheological properties and water content of raw and heated sea cucumber meat were studied. Sea cucumber Stichopus japonicus was heated at 25°C , 70°C and 100°C water for 5 min. The structural changes were observed using a light microscope and the rheological parameters (rupture strength, adhesive strength and deformation) determined using a texture meter. Microscopic photograph revealed that the structural change of heated meat was greater than that of raw meat. The rupture strength, adhesive strength and deformation of raw meat were smaller than those of the heated meat. Meanwhile, rheological parameters showed positive correlation with heating temperature. These changes are mainly caused by thermal denaturation and gelatinization of collagen during heating. These changes were also evidenced in observations using a light microscope and differential scanning calorimetry.     

Stress Responses to Rapid Temperature Changes of the Juvenile Sea Cucumber （Apostichopusjaponicus Selenka）

Activities of hexokinase （HK）, pyruvate kinase （PK）, superoxide dismutase （SOD） and catalase （CAT） and Hsp70 level were measured to evaluate the response of the commercially important sea cucumber （Apostichopus japonicus Selenka） to rapid temperature changes in laboratory. Animals were subjected to a higher temperature （from 10 to 20℃） （Tinc treatment） or to a lower temperature （from 20 to 10℃） （Tddec treatment） for 72 h. At 1, 3, 12, 24, 72 h of exposure, animals were removed and prepared for further analysis. Results showed that the effect of acute temperature changes on enzyme activities was significant. In Tinc treatment, activities of SOD and CAT increased immediately. The significant enhancement of SOD and CAT activities suggested that oxidative stress increases significantly when ambient temperature increasing from 10 to 20℃. The up-regulation of Flsp70 in Tinc and Tdec treatments indicated that Hsp70 was a bioindicator of thermal stress in the sea cucumber, and the expression pattern depended on the thermal treatment.     

Effect of temperature under unfavourable conditions on oxygen uptake of larval herring,Clupea harengus

Oxygen uptake under starvation and short periods of sudden temperature change was measured for larval herring (Clupea harengus L.) reared at average temperature of 7.3, 11 and 12.9°C. Larval stages between first feeding and premetamorphosis were used. For comparison, the routine oxygen uptake (ROU) was also investigated and followed the relationshipQ=0.974+0.174tW ^−0.210, whereQ is in μg/(mg·h),W is dry body weight in mg andt is temperature in °C. The oxygen uptake under starvation (SOU, deprived of food for 24 h) was different from the routine when the larval dry weight was less than 0.6–0.8 mg; it increased with temperature and body weight giving the relationshipQ=1.568+0.110tW ^0.380; if the larval dry weight was more than 0.6–0.8 mg, it reverted to the norm (Q=1.704+0.078tW ^−0.349). The oxygen uptake was tested in short periods (3 h) of sudden temperature changes in six groups: 7.3 to 11, 7.3 to 12.9, 11 to 7.3, 11 to 12.9, 12.9 to 7.3 and 1.29 to 11°C. The oxygen uptake in the 7.3 to 12.9 and 12.9 to 7.3°C groups varied in a way similar to that of the larvae under starvation. The other four groups' oxygen uptake were more or less normal.     

Effect of culture methods on individual variation in the growth of sea cucumber Apostichopus japonicus within a cohort and family

There is substantial individual variation in the growth rates of sea cucumber Apostiehopus japonicus individuals. This necessitates additional work to grade the seed stock and lengthens the production period. We evaluated the influence of three culture methods （free-mixed, isolated-mixed, isolated-alone） on individual variation in growth and assessed the relationship between feeding, energy conversion efficiency, and individual growth variation in individually cultured sea cucumbers. Of the different culture methods, animals grew best when reared in the isolated-mixed treatment （i.e., size classes were held separately）, though there was no difference in individual variation in growth between rearing treatment groups. The individual variation in growth was primarily attributed to genetic factors. The difference in food conversion efficiency caused by genetic differences among individuals was thought to be the origin of the variance. The level of individual growth variation may be altered by interactions among individuals and environmental heterogeneity. Our results suggest that, in addition to traditional seed grading, design of a new kind of substrate that changes the spatial distribution of sea cucumbers would effectively enhance growth and reduce individual variation in growth of sea cucumbers in culture.     

Starvation metabolism of two common species of chironomids in Biandantang Lake

1 INTRODUCTION Chironomid larvae are main groups in most aquatic ecosystems, playing a crucial ecological role in decomposition of detritus and material ex- changes between water column and the sediment (Liang et al., 1995a, b; Chen, et al., 1982). They a…     

Microsatellite DNA polymorphisms and the relation with body weight in sea cucumber <Emphasis Type="Italic">Apostichopus japonicus</Emphasis>

The relationship between microsatellite polymorphism and body weight of captive bred Chinese sea cucumber Apostichopus japonicus was investigated in two local populations in Dalian. Among ten loci discovered, nine show changes except for AJ07 loci. Seven loci were found highly polymorphic in both populations. For each locus in two populations, the average number of alleles is 6.428 6 and 6.285 7, the average observed heterozygosity at 0.225 7 and 0.245 9, the expected heterozygosity at 0.776 8 and 0.748 8, the polymorphism information content (PIC) at 0.709 2 and 0.674 6, respectively. Further analysis show significant correlation between A. japonicus body weight and occurrence markers AJ02 and AJ04. The findings of the relation may be helpful for molecular breeding, as well as the marker-assisted selection of sea cucumbers. Supported by the National High Technology Research and Development Program of China (863 Program, No. 2006AA10A411), the Natural Science Foundation of Liaoning Province (No.20072139), the Grant of Dalian Fisheries University, the Key Laboratory Foundation of the Educational Department of Liaoning Province (No.2009S024)     

Oxygen consumption and ammonia-N excretion of Meretrix meretrix in different temperature and salinity

Effects of temperatures and salinities on oxygen consumption and ammonia-N excretion rate of clamMeretrix meretrix were studied in laboratory from Oct. 2003 to Jan. 2004. Two schemes were designed in incremented temperature at 10, 15, 20, 25°C at 31.5 salinity and in incremented salinity at 16.0, 21.0, 26.0, 31.5, 36.0, and 41.0 at 20°C, all for 8–10 days. From 10 to 25°C, both respiration and excretion rate were increased. One-way ANOVA analysis demonstrated significant difference (P<0.01) in physiological parameters in this temperature range except between 15 and 20°C. The highestQ ₁₀ thermal coefficient value (12.27) was acquired between 10 and 15°C, and about 1 between 15 and 20°C, indicatingM. meretrix could well acclimate to temperature changes in this range. Salinity also had significant effects on respiration and excretion rate (P<0.05). The highest values of respiration and excretion rate ofM. meretrix were recorded at 16.0 salinity (20°C). These two physiological parameters decreased as salinity increased until reached the minimumQ ₁₀ value at 31.5 (20°C), then again, these parameters increased with increasing salinity from 31.5 to 41.0.M. meretrix can catabolize body protein to cope with osmotic pressure stress when environmental salinity is away from its optimal range. No significant difference was observed between 26.0 and 36.0 in salinity (P>0.05), suggesting that a best metabolic salinity range for this species is between 26.0 and 36.0. This work is supported by National High-Tech R & D Program of China. (863 Program) (2002AA603014).     

Metabolism of polychaete Neanthes japonica Izuka： relations to temperature, salinity and body weight

Polychaete Neanthes japonica is a species geographically specific in China and Japan with important scientific implication and commercial value. In this study, the relations of body weight, salinity and temperature to oxygen consumption and ammonia excretion of N. japonica were determined. Three different groups in body weight (large: 2.34±0.36 g, middle: 1.50±0.21 g and small: 0.62±0.12 g) were set for all experiments. Results show that the body weight is negatively related to the rates of oxygen consumption and ammonia excretion; and the relationship is significant. The oxygen consumption and ammonia excretion at 24°C decreased at salinity from 5 to 30 and increased above 30, indicating that both lower and higher salinity are adverse and certain degree of salinity stress is necessary for enhancing the energy demand. At salinity 30, rising temperature from 18°C to 30°C, the oxygen consumption increased before 27°C and then decreased. However, the relation of ammonia excretion and temperature seems more complex. Two-way ANOVA shows that salinity, temperature and body weight all have a significant effect on the oxygen consumption and ammonia excretion of the worm. Moreover, interaction between salinity/temperature and body weight is also significant. O:N (oxygen/nitrogen) ratio varies greatly in this case from 5.97 to 463.22, indicating that N. japonica can regulate the type of metabolic substrate against environment changes. Supported by the National Natural Science Foundation of China (No. 30490233) and the Three-Gorge Project and the Estuarine Environment (No. SX2004-018) from the Three Gorges Project Construction Committee     

Antibacterial activity of extracts from <Emphasis Type="Italic">Zostera marina</Emphasis> against pathogens of <Emphasis Type="Italic">Apostichopus japonicus</Emphasis> skin ulceration disease

The purpose of this study was to investigate the antibacterial activity of extracts from Zostera marina against the pathogens of Apostichopus japonicus skin ulceration disease. When 95% ethanol (v/v) solvent was used to extract Zostera marina at 50°C, aqueous extract (ZA) showed obvious bacteriostatic effects on the tested bacterial strains (inhibition halo diameters between 8.23 mm and 13.62 mm), whereas the ethyl acetate extract (ZE) was almost inactive. The minimal inhibitory concentration (MIC) of ZA against four pathogens were homogeneous at 12.8 g L⁻¹. ZA components were analyzed by thin layer chromatography (TLC) assay and six fractions were obtained. In another study, the six fractions showed inhibitory effects against the tested bacteria while their functions seemed to counteract the ZA activity.