Molecular cloning of heat shock protein 60 from Marsupenaeus japonicus and its expression profiles at early developmental stages and response to heat stress

Water temperature is an essential environmental factor in aquaculture that affects many aspects of organism, and a rise in water temperature stresses aquatic animals. HSP60 is a major component of the chaperone system and plays an important role in stress response. In this study, the full‐length cDNA sequence of HSP60 from Marsupenaeus japonicus was cloned for the first time, and the tissue distributions and expression profiles of MjHSP60 at the early developmental stages of M. japonicus and under heat stress were verified by qRT‐PCR. The full‐length cDNA sequence of MjHSP60 was 2,303 bp with the deduced amino acid (AA) sequence of 579 AA. MjHSP60 was expressed in all eight tested tissues of M. japonicus and was mainly expressed in the hepatopancreas under normal condition. MjHSP60 was expressed in all of the early developmental stages examined from the fertilized egg to the post‐larval stage, and the expression level of MjHSP60 peaked at zoea II and reached a trough at the transition periods between each two stages (N and M1) and showed a relatively stable expression level at the post‐larval stage. The expression level of MjHSP60 was significantly up‐regulated in the gills, muscle, heart, hepatopancreas and stomach at 3 hr post‐heat‐stress and showed varied sensitivity to heat stress. The expression profile of MjHSP60 in the hepatopancreas and gill showed a time‐dependent manner post‐heat‐stress and peaked at 3 hr post‐heat‐stress. The present study provided a basis for further studies for elucidating the function of MjHSP60 in the heat stress response and the early developmental stages of M. japonicus.     

Metabolic responses of matrinxã, Brycon amazonicus (Spix & Agassiz, 1829), exposed to environmental nitrite

Nitrite is usually found in aquatic environments where nitrification process occurs. This ion can cause several injuries to aquatic species, particularly fish. Nitrite reacts with haemoglobin yielding the non‐functional methaemoglobin, which leads to many physiological consequences such as functional anaemia and supposed hypoxia. Metabolism of the freshwater teleost matrinxã, Brycon amazonicus, exposed to environmental 0.6 mg L^?1 of nitrite N‐NO₂^? for 96 h was studied, and the fermentative/oxidative preference was gauged. Concentrations of glycogen, glucose, lactate, pyruvate and ammonia, plus the activities of lactate, glutamate and malate dehydrogenase were assayed. The exposure resulted in a metabolic profile that allowed inferring the continuity of oxidative metabolism. Catabolism of amino acids prevailed or was apparently exacerbated by inferred branchial injury and consequent impairment of nitrogen excretion. Moreover, the studied enzymes glutamate dehydrogenase, lactate dehydrogenase and malate dehydrogenase from brain and heart were little affected by nitrite. The expected fermentative metabolism due to the high methaemoglobin formation was not observed.     

Response of facilitative glucose transporter 1 to salinity stress and dietary carbohydrate nutrition in white shrimp Litopenaeus vannamei

Facilitative glucose transporter 1 (GLUT1) is a transporter protein for glucose transport via the plasma membrane of the cells to provide energy through carbohydrate metabolism. GLUT1 cDNA from Litopenaeus vannamei was obtained and analysed in this study. Full‐length GLUT1 cDNA is 2062 bp long and contained a 1506‐bp ORF encoding a 502 amino acid protein, a 270‐bp 5′UTR and a 284‐bp 3′UTR. When shrimp were under acute low salinity stress, the expression in hepatopancreas, muscle, gill and eyestalk was all up‐regulated at 12 h (P < 0.05) and 96 h (P < 0.05), while the expression in the four tissues was all down‐regulated at 6 h (P < 0.05) and 48 h (P < 0.05) . The expression in the muscle of shrimp at water salinity of 3 was lower than that at water salinity of 30 independent of dietary carbohydrate levels, while expression in hepatopancreas, gill and eyestalk was up‐regulated at 200 and 300 g kg^?1 carbohydrate levels. The expression in all tissues fed glucose was up‐regulated when compared to the expression in shrimp held at a water salinity of 30. This study suggests that GLUT1 is a conserved protein in L. vannamei, and changes in expression due to environmental salinity and dietary carbohydrate level and source.     

Effects of dietary supplementation with freeze‐dried powder of Ampithoe sp. on the growth performance,energy metabolism,and ammonia‐nitrogen tolerance of the Pacific white shrimp,Litopenaeus vannamei

This study evaluated the influence of dietary supplementation with freeze‐dried powder of Ampithoe sp. (FDPA) on the growth, energy metabolism, and resistance to ammonia‐nitrogen stress in Litopenaeus vannamei. There were four treatment groups: a 0% group (no FDPA addition), a 33% group, a 66% group (33% and 66% of the shrimp diet, respectively, replaced with FDPA), and a 100% group (only FDPA). The results of this study suggested a positive effect of FDPA supplementation on shrimp survival: the supplemented groups had significantly higher survival than the 0% group (p < 0.05). The body length, body weight, and specific growth rate (SGR) of the 33% group were higher than those of the other groups and were significantly higher than that of the 100% group (p < 0.05). FDPA feeding had a negative effect on carbohydrate metabolism pathways and energy consumption due to decreases in pyruvate kinase (PK), phosphofructokinase (PFK), succinate dehydrogenase (SDH), lactic dehydrogenase (LDH), and respiratory electron transport system (ETS) activity in shrimp fed FDPA during the culture period. The shrimp in the 33% group exhibited good resistance to ammonia‐nitrogen stress. Additionally, the glycolysis pathway and energy consumption of shrimp in the 33% group were enhanced during the ammonia‐nitrogen stress period. Consequently, it was inferred that FDPA supplementation could improve the resistance of shrimp to ammonia‐nitrogen stress (in the 33% group), which might be related to the effects of the supplement on energy metabolism pathways, particularly in terms of enhancing glycolysis to provide sufficient energy for the stress response.     

Metabolic enzyme activities in relation to crowding stress in the wedge sole (Dicologoglossa cuneata)

In this work, we have assessed the effects of different stocking densities on the biometry, survival and physiological status of the wedge sole (Dicologoglossa cuneata), focusing on changes in the stress system and intermediate metabolism, with the aim of determining a stress indicator for chronic‐stress situations in this species. Wedge sole were kept at three different stocking densities (0.2, 0.4 and 0.8 kg m^?3) for 197 days, and survival, growth, plasma metabolites, cortisol and enzymatic activities were assessed. Survival rates were the highest at low density, though growth did not vary significantly among treatments. Enzymatic activities, mainly in muscle, differed depending on stocking density. Liver hexokinase activity at low stocking density was the highest, while no differences were detected for the other enzymes assessed. In muscle, all enzymes significantly increased in activity with stocking density. We concluded that long‐term high stocking density culture significantly changed enzyme activities (hexokinase, glutamate dehydrogenase, glyceraldehyde 3‐phosphate dehydrogenase, and lactate dehydrogenase) in wedge sole muscle, although liver enzymes, plasma cortisol and metabolites did not vary significantly. Therefore, muscle enzymes, hexokinase and glutamate dehydrogenase, could be considered stress indicators for this species in chronic‐stress situations.     

iTRAQ‐based quantitative proteomic analysis of Sargassum fusiforme in response to high temperature stress

Global warming increases seawater temperature, causing high temperature stress to marine organisms, including algae. This study aimed to explore the global proteomic response of Sargassum fusiforme under high temperature stress. Sargassum fusiforme seedlings were cultured in natural seawater for 24 hr and subjected to different temperatures (22°C, control group; 27°C and 32°C, high temperature stress group) for 1, 3, 5 and 7 days. Changes in their membrane lipid peroxidation after high temperature stress were investigated. Proteomic changes in the air bladders of S. fusiforme were analysed using isobaric tags for relative and absolute quantification, along with liquid chromatography–tandem mass spectrometry. Data were analysed using bioinformatics methods. Results showed that high temperature stress destroyed the cell membrane of the air bladders. Further, 28 and 53 differentially expressed proteins (DEPs) were found in the 27°C and 32°C treatment groups respectively. These DEPs were mainly involved in glycolysis, single‐organism catabolism, purine nucleoside diphosphate metabolism and carbohydrate catabolism. In addition, DEPs were significantly enriched in 10 pathways, such as glycolytic process, biosynthesis of antibiotics, ribosome, biosynthesis of secondary metabolites and biosynthesis of amino acids. Proteomics analyses indicated that proteins associated with synthesis, folding, degradation, photosynthesis and energy and carbohydrate metabolism are differentially expressed under high temperature stress and normal conditions.     

采用Illumina MiSeq测序技术比较日本鲭和大黄鱼冷藏期间的腐败特性

为比较日本鲭和大黄鱼肌肉中微生物和代谢功能的变化及其与鱼肉腐败特性之间的关系，本研究检测了2种鱼在冷藏过程中的理化指标和菌落总数的变化，利用Illumina Miseq测序技术分析细菌群落变化，并利用皮尔森相关性分析检验微生物与鱼肉腐败及组胺产生相关性，结合功能预测分析细菌群落组成与代谢功能之间的关系。结果显示，冷藏期间日本鲭和大黄鱼的pH、挥发性盐基氮、组胺、菌落总数等均呈上升趋势，且日本鲭上升较快；冷藏末期2种鱼TVB-N值和组胺含量分别达到76.34、59.98和59.92、3.11 mg/100 g；日本鲭肌肉中细菌丰富度和多样性先增加后减少，大黄鱼则整体呈下降趋势；2种鱼肌肉中的优势腐败菌均为希瓦氏菌属；日本鲭体内与TVB-N产生相关的菌共12种，其中10种与组胺产生具有显著相关性；大黄鱼体内与TVB-N产生相关的菌共7种，但未检测出与组胺产生具有相关性的细菌；冷藏过程中氨基酸代谢和碳水化合物代谢为最主要的代谢通路，日本鲭样品组氨酸、精氨酸、脯氨酸等氨基酸代谢相关基因和丁酸丁酯代谢、丙酸酯代谢及丙酮酸代谢丰度均显著高于同一时期的大黄鱼，本实验从微生物代谢水平解释了日本鲭比大黄鱼更易腐败的原因，为不同水产品腐败特性的研究提供新思路。     

A global view of hepatopancreas and intestinal reveals the potential influencing mechanism of aflatoxin B1 on nutrition and metabolism in Litopenaeus vannamei

The present study was to determine the time‐dependent alterations in growth performance, histomorphology, digestive enzymes activities and the genes expression related to target of rapamycin (TOR) signalling pathway involved in hepatopancreas and intestine of Litopenaeus vannamei at 3, 6, 12, 18, 24 and 30 days after Aflatoxin B1 (AFB1) challenge. The results manifested that AFB1 could induce a significant decrease in growth performance and visible variations of the hepatopancreas and intestine structures in shrimp. Meanwhile, the digestive enzymes activities and genes expression level were significantly lower in the experimental group than in the control group and showed a time‐dependent decrease (p < 0.05). The expression levels of tor and s6k were significantly higher in the experimental group than in the control group from days 6 to 24 and showed a tendency to increase first and then decrease as a whole (p < 0.05). Meanwhile, we analysed candidate pathways involved in hepatopancreas and intestine of shrimp in metabolic response to AFB1. The mainly disturbed pathways related to metabolism in hepatopancreas were involved in pyrimidine, glycosylphosphatidylinositol, glycosaminoglycan, vitamin and amino acid biosynthesis and metabolism, while the mainly disturbed metabolic pathways in intestine were glycosaminoglycan, vitamin and amino acid biosynthesis and metabolism.     

Utilization of dietary carbohydrates by sea cucumber Apostichopus japonicus (Selenka) as indicated by carbon stable isotope analysis

Carbon stable isotopes were used as trophic markers to investigate the utilization of dietary terrestrial‐source carbohydrates by sea cucumber Apostichopus japonicus. Sea cucumbers were fed by five different types of diets with the ingredients containing Sargassum muticum either without starch or with one of the four starches including corn starch, sweet potato starch, tapioca starch and potato starch. After the 70‐day feeding trial, the carbon isotopic compositions of A. japonicus appeared to reflect those of corresponding dietary components. The average contribution of corn starch (22.0%) to the growth of A. japonicus was slightly higher than the expected contribution (20%). While the proportional contributions of sweet potato, tapioca and potato starches (6.0%, 7.0% and 4.0%, respectively) were all considerably lower relative to the expected contributions. These results indicated that A. japonicus could utilize corn starch more efficiently than sweet potato, tapioca or potato starch. Moreover, A. japonicus fed diet containing corn starch showed the highest specific growth rates which were significantly higher than those fed diets containing potato or tapioca starch. The results of the present study suggested that the corn starch could be used as dietary carbohydrate source in the artificial feeds for A. japonicus farming.