In order to verify the synthesis pathway of linoleic acid(LA)to generate arachidonic acid(ARA),the functions ofΔ6 FAD and Elovl 5 in Apostichopus japonicus were tested by heterologous expression in Pichia pastoris.A ...In order to verify the synthesis pathway of linoleic acid(LA)to generate arachidonic acid(ARA),the functions ofΔ6 FAD and Elovl 5 in Apostichopus japonicus were tested by heterologous expression in Pichia pastoris.A 60-day feeding experiment was conducted to evaluate the effects of dietary LA and ARA on growth,polyunsaturated fatty acids(PUFA)biosynthesis and im-mune function of A.japonicus.Seven diets containing graded levels of LA or ARA were formulated,and one diet without PUFA was applied as a control.The results confirmed thatΔ6 FAD from A.japonicus has a double desaturation ability ofΔ6 andΔ5 for PUFA,and it works together with Elovl 5 enzyme play important roles in biosynthesis of ARA from LA.With increasing dietary LA from 4.9 to 12.1 g/kg,the expressions ofΔ6 FAD and Elovl 5,levels of LA,ARA and EPA in tissues increased,and the activities of ACP,AKP and CAT enzymes in intestine initially increased and then decreased.Additionally,by increasing dietary ARA from 0.7 to 3.8 g/kg,the contents of ALA,EPA and DHA decreased,while theΔ6 FAD expression and the activities of ACP,AKP and CAT in intes-tine increased significantly.Therefore,appropriate dietary levels of LA(12.1 g/kg)or ARA(3.8 g/kg)build a reasonable composi-tion of n-3/n-6 PUFA in A.japonicus,and improved its growth,antioxidant capacity and immunological defenses.展开更多
Sea cucumber Apostichopus japonicus is a crucial aquatic species known for its nutritional value.However,the genetic basis and regulatory mechanisms underlying its nutritional quality remain underexplored.This study i...Sea cucumber Apostichopus japonicus is a crucial aquatic species known for its nutritional value.However,the genetic basis and regulatory mechanisms underlying its nutritional quality remain underexplored.This study investigates the nutritional quality of A.japonicus from different geographical regions and identifies genetic markers associated with these traits through a genome-wide association study(GWAS).We observed significant regional variations in the nutritional content of A.japonicus.Samples collected from Nanhuangcheng Island displayed the highest levels of saponins,whereas those from Laizhou exhibited the highest concentrations of glycosaminoglycans.Lingshan Island samples were the richest in amino acids,while samples from Rizhao contained the highest levels of polyunsaturated fatty acids.Through GWAS,265 candidate genes and related single nucleotide polymorphisms(SNPs)were identified as being significantly associated with essential nutritional traits,including genes like ubiquitin domain-containing protein 1(UBTD1),inactive pancreatic lipase-related protein 1,protein arginine N-methyltransferase 5(PRMT5)and GDP-fucose protein O-fucosyltransferase 1(POFUT1).This study advanced our knowledge of the genetic mechanisms underlying the nutritional quality of A.japonicus.The genetic markers identified herein o ffer crucial insights for breeding initiatives aimed at optimizing the nutritional profile of sea cucumbers.展开更多
In order to control the spawning quality and improve the economic benefit of Apostichopus japonicus Selenka, the indoor temperature and light control methods were employed for artificial maturation, spawning and hatch...In order to control the spawning quality and improve the economic benefit of Apostichopus japonicus Selenka, the indoor temperature and light control methods were employed for artificial maturation, spawning and hatching of A. japonicus, and the larval survival rate, disease incidence rate and economic benefit were analyzed. The results showed that compared with those of conventional temperature-reared larvae, the survival rate of raised temperature-reared larvae was increased by 15.8%, the disease incidence rate was reduced by 10.2%, and the economic benefit was improved by 21.6%. These results will provide theoretical basis and technical support for the artificial breeding of A. japonicus.展开更多
The effect of water temperature on gut mass and digestive enzyme activity in sea cucumber Apostichopusjaponicus, including relative gut mass （RGM）, amylase, lipase, pepsin and trypsin activities were studied at temp...The effect of water temperature on gut mass and digestive enzyme activity in sea cucumber Apostichopusjaponicus, including relative gut mass （RGM）, amylase, lipase, pepsin and trypsin activities were studied at temperatures of 7, 14, 21, and 28℃ over a period of 40 days. Results show that RGM significantly decreased after 40 days at 21 ℃ and markedly decreased over the whole experiment period at 28℃; however, no significant effect of duration was observed at 7 or 14℃. At 14℃, trypsin activity significantly decreased over 10 and 20 days, then increased; amylase and trypsin activity significantly decreased after 40 days at 28℃. However, no significant effect of duration was found on amylase, pepsin or trypsin activities in the other temperature treatment groups. At 28℃, 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℃ continuously increased over the whole experimental period. Principle component analysis showed that sea cucumbers on day 40 in the 21℃ group and in the previous 20 days in the 28℃ group were in the prophase of aestivation. At 28℃, sea cucumbers aestivated at 30-40 days after the start of the experiment. It is concluded that the effect of temperature on the digestion ofA. 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.展开更多
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 （Apostichop...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.展开更多
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...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), ly-sozyme (LSZ) activity and superoxide dismutase (SOD) activity in the coelomic fluid were analyzed on the 0th, 5th, 10th and 15th 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 15th day, PC, ISAP and LSZ activity returned to levels similar to those on the 0th 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.展开更多
This study investigated the seasonal changes in carbon (C) and nitrogen (N) stable isotope values of several typical food sources of Apostichopus japonicus in a farm pond, including particulate organic matter (POM), m...This study investigated the seasonal changes in carbon (C) and nitrogen (N) stable isotope values of several typical food sources of Apostichopus japonicus in a farm pond, including particulate organic matter (POM), macroalgae, benthic microalgae and animals such as nematode and copepod. The stable isotope technique was used to quantify relative contributions of various sources to the food uptake by A. japonicus. The results showed that significant changes occurred in the C and N stable isotope values of sea cucumber food sources due to the seasonality of micro-or macroalgae prosperity and the fluctuation of environmental conditions. The sea cucumber A. japonicus exhibited corresponding alterations in feeding strategy in response to the changes in food conditions. Calculation with a stable isotope mixing model showed that macroalgae was the principal food source for A. japonicus throughout the 1-yr investigation, with the relative contribution averaging 28.1% - 63.2%. The relative contributions of other food sources such as copepod and nematode, POM, benthic microalgae to the total food uptake by sea cucumber averaged 22.6% - 39.1%, 6.3% - 22.2%, 2.8% - 6.5%, and 2.8% - 4.2%, respectively. Together these results indicated that the seasonal changes in food sources led to the obvious temporal differences in the relative contribution of various food sources utilized by A. japonicus. Such findings provide the basic scientific information for improving the aquaculture techniques of A. japonicus, particularly for optimizing the food environment of A. japonicus culture in farm ponds.展开更多
In this study, the influences of immersion bathing in different concentrations of Bacillus velezensis DY-6 on the body weight gain rate and non-specific immune enzyme activities of the coelom fluid of sea cucumber (Ap...In this study, the influences of immersion bathing in different concentrations of Bacillus velezensis DY-6 on the body weight gain rate and non-specific immune enzyme activities of the coelom fluid of sea cucumber (Apostichopus japonicus) were determined in order to obtain the optimum bacterial concentration. The gut microbiota change in A. japonicus was then analyzed through high-throughput sequencing during the immersion bathing in B. velezensis DY-6 at the optimum concentration for 49 d. The results illustrate that the body weight growth rate of all bathing groups was higher than that of the control. The highest growth rate (25.3%) was achieved when the bacterial concentration was 1×10^3 CFU/mL. The activities of non-specific immune enzymes (ACP, AKP, SOD and LZM) of all bathing groups increased, and the activities of the enzymes of groups bathed with the bacterium at 1×10^3 and 1×10^4 CFU/mL reached the highest on day 21 and day 28. Taking the growth rate and economic cost into consideration, the optimum concentration of B. velezensis DY-6 was 1×10^3 CFU/mL. The influences of immersion bathing in B. velezensis DY-6 at 1×10^3 CFU/mL on the gut microbiota of A. japonicus were then evaluated through 16S rDNA sequencing analysis. Results showed that the gut microbiota changed with the addition of B. velezensis DY-6, and the richness and diversity of the gut microbiota peaked twice on day 14 and day 21, respectively. In association with the non-specific immune enzyme activities and if day 28 was selected as the dividing point, the community structure of the gut microbiota could be obviously divided into two types. The correlation analysis revealed that the non-specific immune enzyme activities were correlated significantly to some gut bacteria (in the phyla Firmicutes, Proteobacteria, and Bacteroidetes) after immersion bathing in B. velezensis DY-6. Our findings will provide the theoretical foundation for probiotic application in sea cucumber farming.展开更多
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 ...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℃(159.26±3.28).This study clearly indicated that at low temperature(<24℃),metamorphic rate was remarkably higher than at higher temperature(>26℃).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℃ 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.展开更多
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 phosp...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.展开更多
Different amounts of vitamin C were added to diets fed to juveniles （2.5＋0.15 g） of sea cucumber Apostichopus japonicus （Selenka） in an attempt to reduce the stress response of specimens exposed to nitrite stress...Different amounts of vitamin C were added to diets fed to juveniles （2.5＋0.15 g） of sea cucumber Apostichopus japonicus （Selenka） in an attempt to reduce the stress response of specimens exposed to nitrite stress. A commercial feed was used as the control diet and three experimental diets were made by supplementing 1 000, 1 500, or 2 000 mg vitamin C/kg diet to control diet separately in a 45-day experiment. Sea cucumbers were exposed to three different levels （0.5, 1.0, and 1.5 mg/L） of nitrite stress for 4, 8, and 12 h at four time intervals （0, 15, 30, and 45 d）. Growth of the animals was recorded during the experiment. Reactive oxygen species （ROS） （i.e. hydroxyl free radical （-OH）, malondialdehyde （MDA） and total antioxidant capacity （T-AOC）） and antioxidant enzyme activities （i.e., superoxide dismutase （SOD） and eatalase （CAT）） were measured. Response surface methodology （RSM） was used to analyze the effect of multiple factors on ROS indices and enzyme activities. Weight gain （WG） and special growth rate （SGR） of vitamin C supplementation groups were significantly higher than those of control group （P〈0.05）. The levels of-OH and MDA increased under exposure time extending and nitrite concentration increasing, whereas T-AOC level decreased. SOD and CAT activities increased at 4 h and 8 h and decreased at 12 h. During the days in which the animal consumed experimental diets, the levels of-OH and MDA decreased and that of T-AOC increased. This result suggests that diets containing vitamin C could reduce the nitrite stress response in the animals and increase their antioxidant capacity. The multifactor regression equation of growth performance, ROS indices, and duration of feeding results suggest that vitamin C supplementation of 1 400-2 000 mg/kg diet for 29-35 days could reduce effectively the effects of nitrite exposure.展开更多
The sea cucumber, Apostichopusjaponicus, undergoes aestivation to improve survival during periods of high-temperature. During aestivation, the metabolic rate is depressed to reduce the consumption of reserved energy. ...The sea cucumber, Apostichopusjaponicus, 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 l, 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.展开更多
Microflora in the intestinal tract and on the intestinal wall of both cultured and wild Apostichopus japonicus was studied in this paper. The screening for probiotics was performed based on enzyme producing and hemoly...Microflora in the intestinal tract and on the intestinal wall of both cultured and wild Apostichopus japonicus was studied in this paper. The screening for probiotics was performed based on enzyme producing and hemolytic analysis. The results showed that the number of bacteria in the intestinal wall and tract of wild Apostichopus japonicus was（3.30 ± 0.41） ×107cfu/g and（6.39 ± 0.32） ×10~7cfu/g, respectively. The number of bacteria in the intestinal wall and tract of cultured group was（2.83 ± 0.31） ×10~7cfu/g and（5.67 ± 0.53） ×10~7cfu/g, respectively. The dominant species in the intestinal tract of wild group was Vibrio and the Pseudomonas and Shewanella were the secondary dominant species. The dominant species in the cultured group was Vibrio and Pseudomonas. In 224 strains of bacteria, a total of160 strains of bacteria produced enzyme with a ratio of 71.43%. Among these bacteria, 114 strains could produce protease, 114 strains could produce amylase, and108 strains could produce lipase. The percentages were 50.89%, 50.89%, and48.21%, respectively. A total of 23 strains of bacteria could produce hemolytic toxin in 99 strains of bacteria, which accounts for 23.23% of the total bacterial population.Through the comprehensive analysis of test data, we selected 6 strains of bacteria as intestinal potential probiotic strains of Apostichopus japonicus, which were HS1（Pseudomonas）, HS5（Bacillus）, HS7（Shewanella）, HS8（Vibrio）, HS10（Vibrio）, and HS11（Vibrio） respectively.展开更多
Quantitative real-time reverse transcription-polymerase chain reaction （qRT-PCR） is a technique that is widely used for gene expression analysis, and its accuracy depends on the expression stability of the internal ...Quantitative real-time reverse transcription-polymerase chain reaction （qRT-PCR） is a technique that is widely used for gene expression analysis, and its accuracy depends on the expression stability of the internal reference genes used as normalization factors. However, many applications of qRT-PCR used housekeeping genes as internal controls without validation. In this study, the expression stability of eight candidate reference genes in three tissues （intestine, respiratory tree, and muscle） of the sea cucumber Apostichopus japonicus was assessed during normal growth and aestivation using the geNorm, NormFinder, delta CT, and RefFinder algorithms. The results indicate that the reference genes exhibited significantly different expression patterns among the three tissues during aestivation. In general, the β-tubulin （TUBB） gene was relatively stable in the intestine and respiratory tree tissues. The optimal reference gene combination for intestine was 40S ribosomal protein S18 （RPSI 8）, TUBB, and NADH dehydrogenase （NADH）; for respiratory tree, it was β-actin （ACTB）, TUBB, and succinate dehydrogenase cytochrome B small subunit （SDHC）; and for muscle it was α-tubulin （TUBA） and NADH dehydrogenase [ubiquinone] 1α subcomplex subunit 13 （NDUFA13）. These combinations of internal control genes should be considered for use in further studies of gene expression in A.japonicus during aestivation.展开更多
基金supported by the Natural Sci-ence Foundation of Shandong(Nos.ZR2022MC086 and ZR2023MC162).
文摘In order to verify the synthesis pathway of linoleic acid(LA)to generate arachidonic acid(ARA),the functions ofΔ6 FAD and Elovl 5 in Apostichopus japonicus were tested by heterologous expression in Pichia pastoris.A 60-day feeding experiment was conducted to evaluate the effects of dietary LA and ARA on growth,polyunsaturated fatty acids(PUFA)biosynthesis and im-mune function of A.japonicus.Seven diets containing graded levels of LA or ARA were formulated,and one diet without PUFA was applied as a control.The results confirmed thatΔ6 FAD from A.japonicus has a double desaturation ability ofΔ6 andΔ5 for PUFA,and it works together with Elovl 5 enzyme play important roles in biosynthesis of ARA from LA.With increasing dietary LA from 4.9 to 12.1 g/kg,the expressions ofΔ6 FAD and Elovl 5,levels of LA,ARA and EPA in tissues increased,and the activities of ACP,AKP and CAT enzymes in intestine initially increased and then decreased.Additionally,by increasing dietary ARA from 0.7 to 3.8 g/kg,the contents of ALA,EPA and DHA decreased,while theΔ6 FAD expression and the activities of ACP,AKP and CAT in intes-tine increased significantly.Therefore,appropriate dietary levels of LA(12.1 g/kg)or ARA(3.8 g/kg)build a reasonable composi-tion of n-3/n-6 PUFA in A.japonicus,and improved its growth,antioxidant capacity and immunological defenses.
基金Supported by the Key Research and Development Program of Shandong(Nos.2021LZGC029,2023LZGC019)the National Natural Science Foundation of China(No.42076093)+1 种基金the Special Funds for the Central Government to Guide Local Science and Technology Development(No.YDZX2023043)the Taishan Scholars Program(No.tsqn202306279)。
文摘Sea cucumber Apostichopus japonicus is a crucial aquatic species known for its nutritional value.However,the genetic basis and regulatory mechanisms underlying its nutritional quality remain underexplored.This study investigates the nutritional quality of A.japonicus from different geographical regions and identifies genetic markers associated with these traits through a genome-wide association study(GWAS).We observed significant regional variations in the nutritional content of A.japonicus.Samples collected from Nanhuangcheng Island displayed the highest levels of saponins,whereas those from Laizhou exhibited the highest concentrations of glycosaminoglycans.Lingshan Island samples were the richest in amino acids,while samples from Rizhao contained the highest levels of polyunsaturated fatty acids.Through GWAS,265 candidate genes and related single nucleotide polymorphisms(SNPs)were identified as being significantly associated with essential nutritional traits,including genes like ubiquitin domain-containing protein 1(UBTD1),inactive pancreatic lipase-related protein 1,protein arginine N-methyltransferase 5(PRMT5)and GDP-fucose protein O-fucosyltransferase 1(POFUT1).This study advanced our knowledge of the genetic mechanisms underlying the nutritional quality of A.japonicus.The genetic markers identified herein o ffer crucial insights for breeding initiatives aimed at optimizing the nutritional profile of sea cucumbers.
文摘In order to control the spawning quality and improve the economic benefit of Apostichopus japonicus Selenka, the indoor temperature and light control methods were employed for artificial maturation, spawning and hatching of A. japonicus, and the larval survival rate, disease incidence rate and economic benefit were analyzed. The results showed that compared with those of conventional temperature-reared larvae, the survival rate of raised temperature-reared larvae was increased by 15.8%, the disease incidence rate was reduced by 10.2%, and the economic benefit was improved by 21.6%. These results will provide theoretical basis and technical support for the artificial breeding of A. japonicus.
基金Supported by the Science Fund for Creative Research Groups (No.40821004)National Natural Science Foundation of China (No.40576073)+1 种基金Breeding Project of Shandong Province (China),National Key Technology Research and Development Program of China (No. 2006BAD09A02)the National High Technology Research and Development Program of China (863 Program) (No.2006AA100304 /2006AA10A411)
文摘The effect of water temperature on gut mass and digestive enzyme activity in sea cucumber Apostichopusjaponicus, including relative gut mass （RGM）, amylase, lipase, pepsin and trypsin activities were studied at temperatures of 7, 14, 21, and 28℃ over a period of 40 days. Results show that RGM significantly decreased after 40 days at 21 ℃ and markedly decreased over the whole experiment period at 28℃; however, no significant effect of duration was observed at 7 or 14℃. At 14℃, trypsin activity significantly decreased over 10 and 20 days, then increased; amylase and trypsin activity significantly decreased after 40 days at 28℃. However, no significant effect of duration was found on amylase, pepsin or trypsin activities in the other temperature treatment groups. At 28℃, 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℃ continuously increased over the whole experimental period. Principle component analysis showed that sea cucumbers on day 40 in the 21℃ group and in the previous 20 days in the 28℃ group were in the prophase of aestivation. At 28℃, sea cucumbers aestivated at 30-40 days after the start of the experiment. It is concluded that the effect of temperature on the digestion ofA. 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.
文摘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.
基金supported by grant No.2006AA100313 from the National High Technology Research and Development Project of P.R. China (863 Project)
文摘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), ly-sozyme (LSZ) activity and superoxide dismutase (SOD) activity in the coelomic fluid were analyzed on the 0th, 5th, 10th and 15th 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 15th day, PC, ISAP and LSZ activity returned to levels similar to those on the 0th 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.
基金funded by the National Natural Science Foundation of China (Grant Nos.31172426 and 30871931)the Ministry of Science and Technology of China (Grant Nos.2011BAD13B03 and JQ201009)+1 种基金the State Oceanic Administration of China (Grant No.200905020)supported by the Program for New Century Excellent Talents in University
文摘This study investigated the seasonal changes in carbon (C) and nitrogen (N) stable isotope values of several typical food sources of Apostichopus japonicus in a farm pond, including particulate organic matter (POM), macroalgae, benthic microalgae and animals such as nematode and copepod. The stable isotope technique was used to quantify relative contributions of various sources to the food uptake by A. japonicus. The results showed that significant changes occurred in the C and N stable isotope values of sea cucumber food sources due to the seasonality of micro-or macroalgae prosperity and the fluctuation of environmental conditions. The sea cucumber A. japonicus exhibited corresponding alterations in feeding strategy in response to the changes in food conditions. Calculation with a stable isotope mixing model showed that macroalgae was the principal food source for A. japonicus throughout the 1-yr investigation, with the relative contribution averaging 28.1% - 63.2%. The relative contributions of other food sources such as copepod and nematode, POM, benthic microalgae to the total food uptake by sea cucumber averaged 22.6% - 39.1%, 6.3% - 22.2%, 2.8% - 6.5%, and 2.8% - 4.2%, respectively. Together these results indicated that the seasonal changes in food sources led to the obvious temporal differences in the relative contribution of various food sources utilized by A. japonicus. Such findings provide the basic scientific information for improving the aquaculture techniques of A. japonicus, particularly for optimizing the food environment of A. japonicus culture in farm ponds.
基金Supported by the Special Scientific Research Funds for Central Non-profit Institutes,Yellow Sea Fisheries Research Institute,Chinese Academy of Fishery Sciences(No.20603022016008)the National Key R&D Program of China(No.2018YFD0901603)the Agriculture Seed Improvement Project of Shandong Province(No.2017LZGC010)
文摘In this study, the influences of immersion bathing in different concentrations of Bacillus velezensis DY-6 on the body weight gain rate and non-specific immune enzyme activities of the coelom fluid of sea cucumber (Apostichopus japonicus) were determined in order to obtain the optimum bacterial concentration. The gut microbiota change in A. japonicus was then analyzed through high-throughput sequencing during the immersion bathing in B. velezensis DY-6 at the optimum concentration for 49 d. The results illustrate that the body weight growth rate of all bathing groups was higher than that of the control. The highest growth rate (25.3%) was achieved when the bacterial concentration was 1×10^3 CFU/mL. The activities of non-specific immune enzymes (ACP, AKP, SOD and LZM) of all bathing groups increased, and the activities of the enzymes of groups bathed with the bacterium at 1×10^3 and 1×10^4 CFU/mL reached the highest on day 21 and day 28. Taking the growth rate and economic cost into consideration, the optimum concentration of B. velezensis DY-6 was 1×10^3 CFU/mL. The influences of immersion bathing in B. velezensis DY-6 at 1×10^3 CFU/mL on the gut microbiota of A. japonicus were then evaluated through 16S rDNA sequencing analysis. Results showed that the gut microbiota changed with the addition of B. velezensis DY-6, and the richness and diversity of the gut microbiota peaked twice on day 14 and day 21, respectively. In association with the non-specific immune enzyme activities and if day 28 was selected as the dividing point, the community structure of the gut microbiota could be obviously divided into two types. The correlation analysis revealed that the non-specific immune enzyme activities were correlated significantly to some gut bacteria (in the phyla Firmicutes, Proteobacteria, and Bacteroidetes) after immersion bathing in B. velezensis DY-6. Our findings will provide the theoretical foundation for probiotic application in sea cucumber farming.
基金Supported by the National High Technology Research and Development Program of China(863 Program)(No.2006AA10A411)Shandong Agriculture Seed Stock Projects
文摘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℃(159.26±3.28).This study clearly indicated that at low temperature(<24℃),metamorphic rate was remarkably higher than at higher temperature(>26℃).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℃ 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.
基金supported by the Natural Science Funds for Distinguished Young Scientists of Shandong Province (No.JQ201009)the National Great Project of Scientific and Technical Supporting Programs (No.2011-BAD13B03)+2 种基金the National Natural Science Foundation of China (No.30771661)the Public Science and Technology Research Funds Projects of Ocean, State Oceanic Administration of China (No.200905020)the 111 Project of China Ministry of Education (No.B08049)
文摘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.
基金Supported by the Main Direction Program of Knowledge Innovation of Chinese Academy of Sciences(No.KZCX2-EWQ215)
文摘Different amounts of vitamin C were added to diets fed to juveniles （2.5＋0.15 g） of sea cucumber Apostichopus japonicus （Selenka） in an attempt to reduce the stress response of specimens exposed to nitrite stress. A commercial feed was used as the control diet and three experimental diets were made by supplementing 1 000, 1 500, or 2 000 mg vitamin C/kg diet to control diet separately in a 45-day experiment. Sea cucumbers were exposed to three different levels （0.5, 1.0, and 1.5 mg/L） of nitrite stress for 4, 8, and 12 h at four time intervals （0, 15, 30, and 45 d）. Growth of the animals was recorded during the experiment. Reactive oxygen species （ROS） （i.e. hydroxyl free radical （-OH）, malondialdehyde （MDA） and total antioxidant capacity （T-AOC）） and antioxidant enzyme activities （i.e., superoxide dismutase （SOD） and eatalase （CAT）） were measured. Response surface methodology （RSM） was used to analyze the effect of multiple factors on ROS indices and enzyme activities. Weight gain （WG） and special growth rate （SGR） of vitamin C supplementation groups were significantly higher than those of control group （P〈0.05）. The levels of-OH and MDA increased under exposure time extending and nitrite concentration increasing, whereas T-AOC level decreased. SOD and CAT activities increased at 4 h and 8 h and decreased at 12 h. During the days in which the animal consumed experimental diets, the levels of-OH and MDA decreased and that of T-AOC increased. This result suggests that diets containing vitamin C could reduce the nitrite stress response in the animals and increase their antioxidant capacity. The multifactor regression equation of growth performance, ROS indices, and duration of feeding results suggest that vitamin C supplementation of 1 400-2 000 mg/kg diet for 29-35 days could reduce effectively the effects of nitrite exposure.
基金Supported by the National Key Technology Research and Development Program(No.2006AA10A411)the Agricultural Seed Project of Shandong Province
文摘The sea cucumber, Apostichopusjaponicus, 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 l, 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.
基金Supported by Spark Program(2012GA651002)the 863 Program(2012-AA10A412)Liaonning Key Program for Science and Technology Development(2012003003)~~
文摘Microflora in the intestinal tract and on the intestinal wall of both cultured and wild Apostichopus japonicus was studied in this paper. The screening for probiotics was performed based on enzyme producing and hemolytic analysis. The results showed that the number of bacteria in the intestinal wall and tract of wild Apostichopus japonicus was（3.30 ± 0.41） ×107cfu/g and（6.39 ± 0.32） ×10~7cfu/g, respectively. The number of bacteria in the intestinal wall and tract of cultured group was（2.83 ± 0.31） ×10~7cfu/g and（5.67 ± 0.53） ×10~7cfu/g, respectively. The dominant species in the intestinal tract of wild group was Vibrio and the Pseudomonas and Shewanella were the secondary dominant species. The dominant species in the cultured group was Vibrio and Pseudomonas. In 224 strains of bacteria, a total of160 strains of bacteria produced enzyme with a ratio of 71.43%. Among these bacteria, 114 strains could produce protease, 114 strains could produce amylase, and108 strains could produce lipase. The percentages were 50.89%, 50.89%, and48.21%, respectively. A total of 23 strains of bacteria could produce hemolytic toxin in 99 strains of bacteria, which accounts for 23.23% of the total bacterial population.Through the comprehensive analysis of test data, we selected 6 strains of bacteria as intestinal potential probiotic strains of Apostichopus japonicus, which were HS1（Pseudomonas）, HS5（Bacillus）, HS7（Shewanella）, HS8（Vibrio）, HS10（Vibrio）, and HS11（Vibrio） respectively.
基金Supported by the National Nature Science Foundation of China(Nos.41176139,31201972)
文摘Quantitative real-time reverse transcription-polymerase chain reaction （qRT-PCR） is a technique that is widely used for gene expression analysis, and its accuracy depends on the expression stability of the internal reference genes used as normalization factors. However, many applications of qRT-PCR used housekeeping genes as internal controls without validation. In this study, the expression stability of eight candidate reference genes in three tissues （intestine, respiratory tree, and muscle） of the sea cucumber Apostichopus japonicus was assessed during normal growth and aestivation using the geNorm, NormFinder, delta CT, and RefFinder algorithms. The results indicate that the reference genes exhibited significantly different expression patterns among the three tissues during aestivation. In general, the β-tubulin （TUBB） gene was relatively stable in the intestine and respiratory tree tissues. The optimal reference gene combination for intestine was 40S ribosomal protein S18 （RPSI 8）, TUBB, and NADH dehydrogenase （NADH）; for respiratory tree, it was β-actin （ACTB）, TUBB, and succinate dehydrogenase cytochrome B small subunit （SDHC）; and for muscle it was α-tubulin （TUBA） and NADH dehydrogenase [ubiquinone] 1α subcomplex subunit 13 （NDUFA13）. These combinations of internal control genes should be considered for use in further studies of gene expression in A.japonicus during aestivation.
