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An 8-week feeding trial was conducted in Pacific white shrimp (Litopenaeus vannamei) to evaluate the effects of dietary choline supplementation on choline transport and metabolism, hepatopancreas histological structure and fatty acid profile, and regulation of lipid metabolism. Six isonitrogenous and isolipidic diets were formulated to contain different choline levels of 2.91 (basal diet), 3.85, 4.67, 6.55, 10.70 and 18.90 g/kg, respectively. A total of 960 shrimp (initial weight, 1.38 ± 0.01 g) were distributed randomly into twenty-four 250-L cylindrical fiber-glass tanks, with each diet assigned randomly to 4 replicate tanks. The results indicated that dietary choline significantly promoted the deposition of choline, betaine and carnitine (P < 0.05). The diameters and areas of R cells, total lipid and triglyceride contents in hepatopancreas, and triglyceride and non-esterified fatty acid contents in hemolymph were negatively correlated with dietary choline level. The contents of functional fatty acids in hepatopancreas, the activity of acetyl-CoA carboxylase (Acc), and the mRNA expression of fas, srebpand accwere highest in shrimp fed the diet containing 4.67 g/kg choline, and significantly higher than those fed the diet containing 2.91 g/kg, the lowest level of choline (P < 0.05). The number of R cells, content of very low-density lipoprotein (VLDL), activities of carnitine palmitoyl-transferase (Cpt1), lipoprotein lipase and hepatic lipase, and the mRNA expression levels of cpt1, fabp, fatp, ldlr, and ampkin hepatopancreas increased significantly as dietary choline increased (P < 0.05). In addition, hepatopancreas mRNA expression levels of ctl1, ctl2, oct1, badh, bhmt, ck, cept, and cctwere generally up-regulated as dietary choline level increased (P < 0.01). In conclusion, dietary choline promoted the deposition of choline and its metabolites by up-regulating genes related to choline transport and metabolism. Moreover, appropriate dietary choline level promoted the development of hepatopancreas R cells and maintained the normal accumulation of lipids required for development, while high dietary choline not only promoted hepatopancreas lipid export by enhancing VLDL synthesis, but also promoted fatty acid β-oxidation and inhibited de novo fatty acid synthesis by activating the Ampk/Srebp signaling pathway. These findings provided further insight and understanding of the mechanisms by which dietary choline regulated lipid metabolism in L. vannamei.

The physiological processes involved in adaptation to osmotic pressure in euryhaline crustaceans are highly energy demanding, but the effects of dietary lipids (fat) on low salinity adaptations have not been well evaluated. In the present study, a total of 120 mud crabs (Scylla paramamosain, BW = 17.87 ± 1.49 g) were fed control and high-fat (HF) diets, at both medium salinity (23‰) and low salinity (4‰) for 6 wk, and each treatment had 3 replicates with each replicate containing 10 crabs. The results indicated that a HF diet significantly mitigated the reduction in survival rate, percent weight gain and feed efficiency induced by low salinity (P < 0.05). Low salinity lowered lipogenesis and activated lipolysis resulting in lipid depletion in the hepatopancreas of mud crabs (P < 0.05). Thus, HF diets enhanced the process of lipolysis to supply more energy. In the gills, low salinity and the HF diet increased the levels of mitochondrial biogenesis markers, the activity of mitochondrial complexes, and the expression levels of genes related to energy metabolism (P < 0.05). Consequently, the positive effects of the HF diet on energy metabolism in mud crabs at low salinity promoted osmotic pressure regulation. Specifically, significantly higher haemolymph osmotic pressure and inorganic ion content, as well as higher osmotic pressure regulatory enzyme activity in gills, and gene and protein expression levels of NaK-ATPase were observed in crabs fed the HF diet at low salinity (P < 0.05). In summary, high dietary lipid levels improved energy provision to facilitate mitochondrial biogenesis, which increased ATP provision for osmotic pressure regulation of mud crabs. This study also illustrates the importance of dietary lipid nutrition supplementation for low salinity adaptations in mud crabs.

The objective of the present study was to evaluate the effects of replacing fish meal (FM) with spray-dried blood meal (SDBM) on growth performance, antioxidant capacity, digestive enzyme activity, tissue Fe content, gene expression related to inflammation, and the TOR pathway in juvenile swimming crab (Portunus trituberculatus). Four isonitrogenous (47.0% protein) and isolipidic (9.0% lipid) diets were formulated to contain different levels of SDBM, with FM replaced by SDBM at 0, 10%, 20%, and 30%, respectively. The results showed that crabs fed diet SDBM30 exhibited the lowest survival among all treatments, and crabs fed the SDBM20 diet had the highest final weight, percent weight gain, specific growth rate and feed efficiency among all treatments. In addition, the protein content of the crab hepatopancreas decreased significantly and the lipid content increased significantly as SDBM replaced FM in the diets. Compared to other diets, crabs fed the SDBM30 diet had the lowest activities of antioxidant enzymes and the highest concentration of malondialdehyde in the hepatopancreas and hemolymph. The Fe contents in hepatopancreas and gill increased significantly, and the color of the hepatopancreas got darker as replacement of FM with SDBM increased from 0% to 30%. The expression of genes related to TOR (tor, akt, s6, and s6k1) and antioxidant (gpx) pathways in the hepatopancreas were down-regulated significantly with increased replacement of dietary FM with SDBM. In contrast, expression levels of genes related to protein translation (4ebp1, eif4e1a, eif4e2, and eif4e3) and inflammation pathways (myd88, traf6, relish, and irak4) were up-regulated significantly in crabs fed diets with SDBM supplementation. In summary, replacing FM with less than 20% of SDBM can promote the growth, but can darken the hepatopancreas, negatively affect antioxidant capacity, and induce an inflammatory response.

The eggshell color of avian species is an important trait that is predominantly determined by the pigments biliverdin and protoporphyrin. Various factors affect eggshell pigment deposition and coloration; however, the underlying mechanisms remain unclear. We analyzed the hepatic transcriptomes and metabolomes of Changshun green-shell hens laying dark green and light green eggs to investigate the potential role of the liver in regulating the intensity of the green eggshell color. In total, 350 differentially expressed genes and 211 differentially altered metabolites were identified. Gene set enrichment analysis revealed that the enriched pathways and Gene Ontology (GO) terms were mainly associated with energy, immunity, and nutrient metabolism. Metabolite set enrichment analysis revealed that the enriched pathways were mainly associated with amino acid, vitamin, bile acid, and lipid metabolism. Moreover, gene–metabolite interaction network analysis revealed 1 subnetwork. Most genes and metabolites in this subnetwork were determined to be related to melanin metabolism and transport. In conclusion, our results suggest that hepatic melanin metabolism and transport are critical for eggshell coloration. Six candidate genes (CDKN2B, DDC, PYCR1, ABCG5, SLC3A1, and P2RX2) and 7 candidate metabolites (serotonin, 5-hydroxyindoleacetic acid, ornithine, acetylcholine, L-tryptophan, D-ornithine, and ADP) were suggested to play important roles in this process. Meanwhile, this study suggests that changes in hepatic energy metabolism, immune status, antioxidation activity, nutrient availability, and bile acid synthesis can impair eggshell coloration.

The intestinal function of hibernating animals undergoes seasonal changes, during which they are in a fasting state, which also affects the morphology, structure, and function of intestine. The aim of this study was to analyse the histological differences in the intestinal structure, goblet cells, and mucosa of the initial (ISI), anterior (ASI), middle (MSI) and posterior (PSI) of small intestine of Pelodiscus sinensisduring the whole hibernation process and to investigate the role of the key enzyme creatine kinase (CK) in regulating energy homeostasis during hibernation. The results of HE staining showed that the intestinal mucosa of the P. sinensisduring hibernation presented detachment and atrophy of villus, and narrowing of gaps while the villous length of the ISI, ASI, MSI, and PSI were significantly decreased than those in normal stage. As hibernation progresses, the intestinal crypt of the ISI first increased and then decreased significantly. The AB-PAS staining results revealed that as hibernation progressed, the number of type Ⅰ goblet cells in the ISI significantly increased, while the number of type Ⅲ goblet cells significantly decreased. The immunohistochemical results of CK-MB revealed that CK-MB was highly expressed in the intestinal villus of P. sinensis. During hibernation, the expression of CK-MB in P. sinensiscontinued to increase in the ISI and decrease in the PSI. In summary, the results of this study suggested that hibernation process could alter the morphological structure of the ISI and ASI of P. sinensis, making them more vulnerable. Meanwhile, the ISI may protect the mucosal tissue during hibernation by increasing type Ⅲ goblet cells and decreasing type I goblet cells. During this process, the continuously increasing expression of CK in the ISI may participate in the adaptation of P. sinensisto intestinal hibernation by regulating energy metabolism.

Salinity has been demonstrated to influence the biosynthesis of long-chain (C20–24) polyunsaturated fatty acids (LC-PUFAs) in teleost fish. Since LC-PUFAs are essential nutrients for vertebrates, it is central to understand how fish cope with an acute change in salinity associated with natural events. We herein report on the cloning and functional characterization of two elongation of very-long-chain fatty acid (Elovl)4 proteins, namely, Elovl4a and Elovl4b, and study the roles that these enzymes play in the biosynthesis of LC-PUFAs and very-long-chain (>C24) polyunsaturated fatty acids (VLC-PUFAs) in marine teleost Pampus argenteus. The P. argenteusElovl4 displayed all of the typical features of Elovl-like enzymes and have eyes and brain as major sites through which they exert their functions. Moreover, functional studies showed that the P. argenteusElovl4 can effectively elongate C18–22substrates to C36VLC-PUFA. Because both P. argenteusElovl4 are able to produce 24:5n– 3 from shorter precursors, we tested whether the previously reported Δ6 Fads2 from P. argenteuswas able to desaturate 24:5n– 3 to 24:6n– 3, a key step for docosahexaenoic acid (DHA) synthesis. Our results showed that P. argenteuscan indeed bioconvert 24:5n– 3 into 24:6n– 3, suggesting that P. argenteushas the enzymatic capacity required for DHA biosynthesis through the coordinated action of both Elovl4 and Fads2. Furthermore, an acute salinity test indicated that low-salinity stress (12 ppt) upregulated genes involved in LC-PUFA biosynthesis, with 12 ppt salinity treatment showing the highest hepatic LC-PUFA content. Overall, our results unveiled that the newly characterized Elovl4 enzymes have indispensable functions in LC- and VLC-PUFA biosynthesis. Moreover, acute salinity change influenced the biosynthesis of LC-PUFA in P. argenteus. This study provided new insight into the biosynthesis of LC- and VLC-PUFAs in vertebrates and the physiological responses that teleosts have under acute salinity stress.

An eight-week feeding trial was conducted to determine the optimal dietary histidine requirement of juvenile swimming crab (Portunus trituberculatus). Six isonitrogenous and isolipidic diets (45.0% crude protein and 8.0% crude lipid) were formulated to contain 0.43%, 0.68%, 0.97%, 1.24%, 1.45% and 1.79% histidine, respectively. Each diet was randomly assigned to triplicate groups of 30 juvenile crabs (approximately initial weight 17.63 ± 0.08 g). The results indicated that the highest percent weight gain (PWG) and specific growth rate (SGR) was presented in crabs fed the diet with 1.24% histidine (P < 0.05). Crabs fed the diet with 1.45% histidine showed remarkably higher activities of alanine transaminase (ALT) and aspartate transaminase (AST) in hemolymph than those fed the other diets (P < 0.05). Meanwhile, the concentrations of malondialdehyde (MDA) in hemolymph and hepatopancreas remarkably decreased with dietary histidine levels increasing from 0.43% to 1.79%, and activities of total superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC) and hydroxyl radical scavenging activity in hemolymph and hepatopancreas exhibited the significant trend of first increasing to 1.17% histidine and then decreasing (P < 0.05). The concentrations of carnosine significantly increased with the increase of dietary histidine levels (P < 0.05). Moreover, dietary histidine levels could significantly influence the mRNA levels of genes related to TOR pathway such as tor, akt, s6k1, s6, eif4e1a, eif4e2and eif4e3, and antioxidant such as cat, cMnsod, gpx, trxand prx(P < 0.05). Two slope broken-line model analysis of PWG against the dietary histidine levels indicated that the optimal dietary histidine requirement was estimated to be 1.17% of the dry matter (2.60% of the dietary protein) for juvenile swimming crab. In conclusion, dietary 1.24% histidine can significantly increase the carnosine contents in muscle, hepatopancreas and hemolymph, improve antioxidant performance and activate the TOR signaling pathway.

An 8-week feeding trial was conducted to evaluate the effects of replacing fish meal with spray-dried egg meal (SEM) on growth performance, anti-oxidative capacity, mTOR pathway, and cholesterol metabolism in juvenile Portunus trituberculatus. The results showed that the highest percent weight gain was observed in crabs fed diet with 10% replacement of fish meal with SEM. Essential amino acids in the muscle and hepatopancreas significantly decreased with the increase of dietary replacement levels of fish meal with SEM. The lowest content of MDA in hepatopancreas was occurred at those fed diet with 40% replacement of fish meal with SEM, the activities of T-AOC, GSH-PX and content of GSH in hepatopancreas significantly increased with dietary SEM replacement fish meal increasing from 10% to 50%. The expression of akt, eif4e1a, eif4e2, eif4e3, and 4ebp1significantly up-regulated, however, the expression of torwas showed significantly linear down-regulated, and the highest expression of akt, eif4e2and 4ebp1were found in crabs fed diet with 20% replacement of fish meal with SEM, the highest expression of eif4e1awas observed in those fed diet with 10% replacement of fish meal with SEM, the highest expression of eif4e3was occurred in crabs fed diet with 30% replacement of fish meal with SEM. In cholesterol metabolism, the expression of abcg1and lrp2significantly down-regulated and the expression of npc1significantly up-regulated with increase of replacement level of fish meal with SEM. The content of T-CHO in hemolymph and hepatopancreas significantly linear increased. In conclusion, based on quadratic broken-line regression analysis of PWG against the dietary replacement of fish meal with SEM, the optimal replacement level of fish meal with SEM was estimated to be 15.94% for juvenile swimming crab.

The present study aimed to evaluate the effects of dietary inositol level on the growth performance and barriers function of hemolymph and intestine of Litopenaeus vannamei. Six isonitrogenous and isolipidic diets were formulated with analyzed inositol levels being 0.86 (basal diet), 1.34, 1.40, 2.30, 3.88, and 7.50 g kg−1, respectively. The results indicated that appropriate dietary inositol could improve the growth performance of Litopenaeus vannamei. The optimal dietary inositol requirement was 1.44 g kg−1according to broken-line regression analysis with the WGR as the evaluation index. The activities of non-specific immune enzymes (SOD, ACP, ALP and NOS) in hemolymph and the expression levels of genes involved in the Toll signaling pathway (toll, pelleand traf6) in hemocyte of shrimp fed diet containing 1.40 g kg−1inositol significantly higher than those in shrimp fed diet containing 0.86 g kg−1inositol, and the activities of PO and SAHR in hemolymph and the expression levels of genes involved in immune enzymes (Mn-sod, Cu/Zn sod, acp, akpand po) in hemocyte of shrimp fed diet containing 1.40, 2.30, 3.88 and 7.50 g kg−1inositol significantly higher than those in shrimp fed diet containing 0.86 g kg−1inositol, suggesting that appropriate dietary inositol can improve hemolymph immunity by increasing the activities of antioxidant and immune enzymes in the hemolymph. In addition, the intestinal VH and the expression levels of genes involved in intestinal health (zo-1, cyclC, tgfβand il2) in shrimp fed diet containing 1.40 g kg−1inositol were the highest. Meanwhile, the inositol content, VW, and the expression levels of genes involved in the IMD pathway (imd) and anti-lipopolysaccharide factor (alf) in the intestine were significantly correlated with dietary inositol level. These results suggested that appropriate inositol can improve intestinal health by improving intestinal morphology and the expression of health-related genes. Although dietary inositol had no significant effect on the microbial composition spectrum diversity in the hemolymph and intestine of L. vannamei, it contributed to increasing the abundance of beneficial bacteria and decreasing the abundance of opportunistic pathogens. KEGG analysis showed that dietary inositol level had significantly positive effects on multiple immune and metabolic pathways. In conclusion, dietary appropriate inositol enhanced hemolymph homeostasis and host immune defense by improving the barrier function of immune and microbial in hemolymph and improved the intestine health status by enhancing the barrier function of physical, immune and microbial in intestine.

Three isonitrogenous diets (41% crude protein) with 4.87%, 10.75% and 16.10% lipid were designed to evaluate the effects of dietary lipid levels growth, antioxidant capacity, expression levels of genes related to lipid and glucose metabolism of juvenile Scylla paramamosain. Each diet was fed to four replicates of 32 crabs with initial weight 5.10 ± 0.05 g for 6 weeks. The results indicated that crabs fed diet with 10.75% lipid exhibited significantly higher percent weight gain (PWG) and specific growth rate (SGR) than those fed diet with 4.87% lipid (P < 0.05), and crabs fed diets with 10.75% and 16.10% lipid had significantly higher protein efficiency ratio (PER) and feed efficiency (FE) than those fed diet with 4.87% lipid (P < 0.05). Crabs fed diet with 4.87% lipid had the lowest concentrations of lipid and glycogen in muscle, lipid content in hepatopancreas and hemolymph glucose among all treatments (P < 0.05). The concentrations of free fatty acid (FFA), triglyceride (TG), total cholesterol (TCHO) and pyruvate in hemolymph and hepatopancreas increased significantly with the dietary lipid levels increasing from 4.87% to 16.10% (P < 0.05), while the concentration of crustacean insulin-like (INS) in hemolymph and crustacean hyperglycemic hormone (CHH) in hepatopancreas significantly decreased with the increase of dietary lipid levels (P < 0.05). In the hepatopancreas, The Oil Red-positive area significantly increased with increase of dietary lipid levels (P < 0.05). Crabs fed diet with 16.10% lipid showed the lowest concentration of glutathione peroxidase (GSH-Px) (P < 0.05). On the contrary, the highest concentration of malondialdehyde (MDA) was observed at crabs fed diet with 16.10% lipid (P < 0.05). Crabs fed diets with 10.75% and 16.10% lipid down-regulated expression of genes related to long-chain-PUFA biosynthesis and lipid synthesis in hepatopancreas than those fed diet with 4.87% lipid (P < 0.05). The mRNA level of cpt1significantly up-regulated with the increase of dietary lipid levels, while the mRNA expression level cpt2, acox1and fabp4in hepatopancreas significantly down-regulated in crabs fed diets with 10.75% and 16.10% lipid (P < 0.05). The expression of gkand pfkin crabs fed diet with 16.10% lipid significantly down-regulated than those in other groups (P < 0.05). The expressions level of genes related to gluconeogenesis significantly down-regulated in crabs fed diets with 10.75% and 16.10% lipid (P < 0.05). The lowest mRNA expression of glucose transporter 4 (glut4) was occurred in crabs fed diet with 16.10% lipid (P < 0.05). Overall, the results of present study demonstrated that the dietary 10.75% lipid can improve the growth and feed utilization for juvenile mud crab, in addition, dietary lipid can affect the antioxidant capacity, lipid metabolism, glucose metabolism and insulin signaling pathway, dietary excessive lipid resulted in lipid deposition of hepatopancreas for juvenile mud crab.

Vegetable oil (VO) is increasingly replacing fish oil (FO) in the commercial diet of crustaceans, partly due to a global shortage of FO. But little is known about how different amounts of substitution affect the health and metabolism of crustaceans. A six-week feeding trial was conducted to evaluated the effects of different replacement levels of FO with linseed oil (LO) on growth performance, antioxidant and digestive capacity and fatty acid metabolism in swimming crab, Portunus trituberculatus. Five isonitrogenous and isolipidic diets (480 g kg−1protein and 120 g kg−1lipid) were formulated to contain graded levels of LO, and FO was replaced with LO at 0%, 25%, 50%, 75% and 100%, respectively. The results showed that 50% of FO could be replaced by LO without affecting the growth performance and feed utilization for juvenile swimming crab. In hemolymph, 100% dietary replacement of FO with LO, the activities of alanine aminotransferase (ALT) and aspartate transaminase (AST) exhibited significantly higher than those fed the other diets, the concentrations of triacylglycerol (TAG), total cholesterol (T-CHO) and high-density lipoprotein cholesterol (HDL-C) significantly increased with increase of dietary replacement of FO with LO, which may lead to hyperlipidemia in swimming crabs. The number of R cells and their lipid droplets in hepatopancreas gradually increased with increase of replacement of FO with LO, the results confirmed the occurrence of lipid accumulation in the hepatopancreas. The activities of glutathione (GSH), and glutathione peroxidase (GPX) in hepatopancreas significantly decreased with increase of replacement of FO with LO, and crabs fed diet with 50% replacement of FO with LO had higher activities of trypsin and lipase (LPS) than those fed the other diets. Long-chain polyunsaturated fatty acids (LC-PUFA), n-3/n-6 PUFA in hepatopancreas and muscle significantly decreased with the increase of substitution level of FO with LO. Moreover, the expression of genes related to LC-PUFA synthesis as well as fatty acid β-oxidation, such as elongase 4 (elovl4), hepatocyte nuclear factor 4-α (hnf4α), retinoid X receptor (rxr), carnitine palmitoyltransferase 2 (cpt2) and acyl-CoA oxidase (acox2), were down-regulated with increase of dietary replacement of FO with LO. In conclusion, it is suggested that the optimal substitution ratio of LO for FO should be less than 50% in the diet of Portunus trituberculatus.

A 22-week feeding experiment was executed to assess the impact of dietary ARA supplementation on the growth, tissue proximate composition, hemolymph biochemical, tissue fatty acids profile, ARA metabolite and sex steroid hormone, and the mRNA expression of ovarian development related gene of female mud crab in the pre-devolopmental stage. Crabs were fed with six diets containing different ARA levels (0.04%, 0.25%, 0.46%, 0.89%, 1.25%, and 1.57%, dry matter). The results showed that the increasing dietary ARA levels had significant effects on survival, percent weight gain (PWG), specific growth rate (SGR), and feed efficiency (FE), which showed an increasing trend initially and then showed a decreasing trend. According to the two-slope broken-line model and quadratic curve model for the relationship between PWG and dietary ARA levels, optimal dietary ARA levels were estimated to be 0.46% and 0.70%. The lipid contents in muscle and hepatopancreas decreased significantly with the increasing dietary ARA levels. Similarly, total cholesterol (T-CHO) and triglyceride (TG) concentrations were significantly decreased with the increasing dietary ARA levels. In the hepatopancreas and muscle, ARA contents significantly increased with the increasing dietary ARA levels. In hemolymph and hepatopancreas, progesterone (Prog), pregnenolone (Preg), estradiol (E2) and vitellogen (Vtg) concentrations presented earlier increase and later decrease trend. In the hepatopancreas, prostaglandin E2(PGE2) and 5-hydroxyeicosatetraenoic acid (5-HETE) concentrations showed an upward trend. In the muscle, PGE2concentration presented earlier increasing and later decreasing trend. In the ovary, the increasing dietary ARA levels produced similar effects on the expressions of genes, including steroidogenic acute regulatory protein (star), 3β-hydroxysteroid dehydrogenase (3βhsd), vitellogenin (vtg) and vitellogenin receptor (vtgr). Thus, moderate dietary ARA levels promoted growth, improved ARA deposition and metabolism, increased sex steroid hormone and Vtg levels, and enhanced ovarian development of female mud crab in the pre-developmental stage.

Hexavalent chromium (Cr6+) is a metallic contaminant that in organisms and poses a great threat to biological health and ecology due to its toxicity and biomagnification. However, the potential health risks to mud crabs and humans from dietary excessive Cr6+were currently unknown. Therefore, a 9-week feeding trial was conducted to evaluate the toxic effects of dietary Cr6+levels (1.4, 77.5 and 312.2 mg/kg) for mud crab (Scylla paramamosain). The results indicated that dietary Cr6+levels had no significant effect on growth performance for mud crab. Cr concentrations in hepatopancreas and muscle significantly increased with dietary Cr6+levels increasing from 1.4 to 312.2 mg/kg. The antioxidant capacity of mud crab was enhanced by increased the activities of superoxide dismutase (SOD) and scavenging capability for hydroxyl free radical (SCHR) to resist the Cr6+toxicity. However, there was no statistical difference in the rate of apoptosis in hemocytes among all treatments. Expression levels of caspase 2gene related to apoptosis in hemocytes significantly up-regulated with dietary Cr6+level increasing from 1.4 to 312.2 mg/kg. For food safety, crabs fed diets with 77.5 and 312.2 mg/kg Cr6+had target hazard quotient (THQ) values greater than 1 for the hepatopancreas, indicating a serious risk for human consumption. According to the target cancer risk (TCR) analysis, people who consumed muscle in mud crab fed diet with 312.2 mg/kg Cr6+were at unacceptable cancer risk. Overall, the results of the present study indicated mud crab had a strong resistance to Cr6+toxicity, and dietary excessive Cr6+did not decline growth performance and survival, oxidative stress and apoptosis in mud crab. Excess Cr6+were accumulated in tissues such as hepatopancreas and muscle, and consumption of crabs tissues fed diets with 77.5 and 312.2 mg/kg Cr6+will provoke health and cancer risk in humans.

Cholesterol, which acts as a substrate for the synthesis of ovarian sex hormones, could promote ovarian development. However, crustaceans cannot synthesize cholesterol and must obtain it from diet for growth, molt and gonadal development. Three isonitrogenous and isolipidic diets were formulated to contain 0.12%, 1.00% and 2.50% cholesterol, respectively. The results showed that dietary 1.00% and 2.50% cholesterol significantly increased percent weight gain (PWG) and specific growth rate (SGR), moreover, crabs fed diet with 1.00% cholesterol had the highest gonadosomatic index (GSI) among all treatments. Cholesterol (CHO) and triglyceride (TG) concentrations in tissues significantly increased with dietary cholesterol levels increasing from 0.12% to and 2.50%. Targeted metabolomics analyses indicated that crabs fed 1.00% cholesterol diet exhibited higher steroid hormones, such as progesterone, pregnenolone, estrone, estradiol and estriol than those fed the other diets. In addition, crabs fed the 1.00% cholesterol diet significantly up-regulated the expression of genes related to cholesterol transport, steroid hormone synthesis and ovarian development in hepatopancreas and ovary. Transcriptomics analyses revealed that DEGs (differentially expressed genes) in crabs fed 1.00% cholesterol diet were associated with “metabolism of xenobiotics by cytochrome P450″ and “progesterone-mediated oocyte maturation”. In conclusion, the results of present study demonstrated that dietary 1.00% cholesterol promoted growth and ovarian development by improving the cholesterol transport and steroid hormone synthesis for female swimming crabs.

An 8-week feeding trial was conducted to determine the optimal dietary phenylalanine requirement of juvenile swimming crab (Portunus trituberculatus). Six experimental diets (45.0% crude protein and 8.0% crude lipid) were formulated to contain 0.89%, 1.15%, 1.41%, 1.64%, 1.90% and 2.18% phenylalanine, respectively. Each diet was randomly divided into triplicate groups with 30 juvenile swimming crabs (initial weight 22.87 ± 0.03 g). The highest percent weight gain (PWG) and feed efficiency (FE) were presented in crabs fed with 1.64% phenylalanine diet, and the lowest PWG and FE were observed in crabs fed diet with 0.89% phenylalanine (P < 0.05), while no significant difference was presented in survival among all treatments (P > 0.05). Crabs fed the diet containing 0.89% phenylalanine had the lowest content of crude lipid in hepatopancreas among all treatments (P < 0.05). Hematological parameters related to glucose and lipids metabolism and enzyme activities involved in glycolysis and gluconeogenesis were significantly affected by dietary phenylalanine levels (P < 0.05). The contents of saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs), n-6 polyunsaturated fatty acids (n-6 PUFAs) and n-3 polyunsaturated fatty acids (n-3 PUFAs) in the hepatopancreas were notably affected by the dietary phenylalanine levels (P < 0.05). The enzyme activities related to glycolysis, gluconeogenesis and glycogen content in hepatopancreas were significantly influenced by dietary phenylalanine levels (P < 0.05). The mRNA levels of genes related to glycolysis and gluconeogenesis in the hepatopancreas were significantly affected by dietary phenylalanine levels (P < 0.05). Moreover, the insulin and mammalian target of rapamycin (mTOR) signaling pathway were notably activated by dietary phenylalanine levels (P < 0.05). Based on two slope broken-line regression analysis of PWG against the dietary phenylalanine levels, the optimal dietary phenylalanine requirement was estimated to be 1.60% dry matter (3.55% dietary protein) for juvenile swimming crab.

Chromium (Cr) is an essential micronutrient for all animals, which is also an important component of glucose tolerance factor. The present study was conducted to evaluate the effect of dietary Cr levels on growth, Cr accumulation in tissues, glucose and lipid metabolism and insulin signal pathway of juvenile mud crab. Six isonitrogenous and isolipidic diets were formulated to contain 1.4 (basal diet), 1.7, 2.0, 2.2, 2.6 and 3.7mgkg-1Cr, respectively. The results indicated that crabs fed with 2.2 and 2.6mgkg-1Cr diets had the highest percent weight gain (PWG) and specific growth rate (SGR) among all treatments. Cr concentration in hepatopancreas significantly increased with increase of dietary Cr levels. Crabs fed with 3.7mgkg-1Cr diet exhibited lower antioxidant enzyme activities and higher malondialdehyde (MDA) in hepatopancreas and hemolymph than those fed the other diets. Crabs fed the diets with 2.2 and 2.6mgkg-1Cr showed higher activities of hexokinase (HK), phosphofructokinase (PFK) and pyruvate kinase (PK). The expression levels of genes related to insulin signaling pathway, glycolysis and fat synthesis, glycolipid metabolism (such as phosphatidylinositol 3-kinase (pi3k), hexokinase (hk), and fatty acid synthase (fas)) in hepatopancreas were significantly up-regulated. The expression levels of genes related to gluconeogenesis and lipolysis such as fructose-1,6-bisphosphatase (fbp) and carnitine palmitoyltransferase 1 (cpt1) were significantly down-regulated in crabs fed with optimal Cr diets. In conclusion, the optimal dietary Cr requirement was estimated to be 2.34mgkg-1for juvenile mud crab based on two slope broken-line regression analysis of PWG against dietary Cr levels, the results of present study indicated that optimal dietary Cr promoted the growth and Cr accumulation in tissues, and modulated insulin signaling pathway to trigger glycolysis and glycogenesis to maintain glucose homeostasis.

An eight-week feeding trial was conducted to investigate the effects of dietary lipid sources on growth performance, fatty acids composition in tissue and expression of genes related to lipid metabolism in juvenile largemouth bass (Micropterus salmoides). Four experimental diets were formulated to contain 6% coconut oil (CO), rapeseed oil (RO), linseed oil (LO) and fish oil (FO), respectively. Each diet was fed to triplicate groups of juvenile for 8 weeks. The results indicated that final body weight (FBW), percent weight gain (PWG), specific growth rate (SGR), feed efficiency (FE), survival, hepatopancreas index (HSI), viscerosomatic index (VSI) and intraperitoneal fat ratio (IPF) were not significantly influenced by dietary different lipid sources (P＞0.05). Moreover, proximate composition in whole body, dorsal muscle and liver were also not significantly affected by dietary different lipid sources (P＞0.05). Fish fed with FO diet was significantly higher MDA concentration in hepatic and serum than those fed with CO, RO and LO diets (P<0.05). However, T-AOC, SOD and GSH activities in hepatic and serum were not affected by different lipid sources (P＞0.05). Fish fed with RO diet had significantly higher relative expression of fasand accin liver than those fed the other diets, the highest expression of lplwas occurred at fish fed with CO diet (p<0.05). Meanwhile, fish fed with CO and FO diets had significantly lower expression of elovl5and Δ6fadin liver than those fed with RO diet (p<0.05). In conclusion, the results of present study indicated that vegetable oils such as CO, RO and LO can potentially be used as dietary FO substitutes for juvenile largemouth bass.

Cholesterol, as an indispensable nutrient, regulates molting and growth in crustacean. As crustaceans are unable to biosynthesize cholesterol de novo, it is central to understand how dietary cholesterol affects molting in crustaceans. An 8-week feeding trial was conducted to evaluate the effects of dietary cholesterol level (0.12%, 0.43%, 0.79%, 1.00%, 1.30% and 2.50%) on growth, cholesterol metabolism and expression of genes related to lipid and ecdysone metabolism in female swimming crabs (Portunus trituberculatus). A total of 192 crabs (1.41 ± 0.05 g) were randomly distributed into 192 aquaria. Each treatment had 4 replicates with each replicate containing 8 crabs. Crabs fed the 1.00% cholesterol diet showed best growth performance, and thus based on percent weight gain, the optimal dietary cholesterol requirement was calculated at 1.01%. Tissue cholesterol concentrations were positively correlated with dietary cholesterol level. The contents of functional fatty acids in hepatopancreas significantly increased as dietary cholesterol increased from 0.12% to 2.50% (P< 0.05). The expression levels of genes related to lipogenesis pathway, lipid catabolism and fatty acid oxidation were significantly down-regulated with increased dietary cholesterol level (P< 0.05). The highest expression levels of cholesterol transport genes, low-density lipoprotein receptor (ldlr) and low-density lipoprotein receptor-related protein 2 (lrp2) occurred in crabs fed the 1.30% cholesterol diet. Moreover, hormones related to molting such as crustacean hyperglycemic hormone (CHH), methyl farnesoate (MF), molt-inhibiting hormone (MIH), and ecdysone in hemolymph were significantly influenced by dietary cholesterol level (P< 0.05). The highest expression levels of ecdysone receptor (ecr) and chitinase 1 (chi1) in eyestalk and hepatopancreas were found in crabs fed the diet containing 1.00% cholesterol (P< 0.05). In conclusion, the optimal dietary level was beneficial to functional fatty acid accumulation, regulated lipid metabolism, promoted the ecdysone signalling pathway by improving the cholesterol transport, and improved the molting rate and growth of swimming crabs.

Abstract: This paper addresses the non-preemptive scheduling problem of scheduling jobs on identical parallel machines to minimize the maximum completion time or makespan. The problem has been proved to be NP-hard in the strong sense. The NP-hardness of the problem motivates us to develop a new methodology to obtain near-optimal solutions. We formulate the problem as an integer programming and then propose a new iterated local search (ILS) algorithm based on a variable number of cyclic exchanges to solve it. The properties of the solutions are derived and the results are used to improve the computational efficiency of our algorithm. Computational experiments show that the cyclic exchange neighborhood embedded in an iterated local search framework is effective for solving the scheduling problems with up to 1000 jobs and 40 machines within a reasonable amount of computation time.

•The optimal dietary P requirement was estimated to be 15.91–16.83 g kg−1for juvenile swimming crab.•There is a positive correlation between P bioaccumulation in tissue and dietary P levels.•Dietary optimal P level increase mitochondrial respiration and strengthen related enzyme activities in hepatopancreas.

An eight-week feeding trial was conducted to investigate the effects of dietary carbohydrate sources on growth performance, glycogen content in tissue, insulin signaling pathway, hepatic glucose transport and metabolism in juvenile swimming crab (Portunus trituberculatus). Four isonitrogenous (approximately 45.4% crude protein) and isolipidic (approximately 9.0% crude lipid) experimental diets were formulated to contain corn starch, wheat starch, dextrin and potato starch. Each diet was randomly assigned to triplicate groups of 30 juvenile swimming crabs (approximately 5.6 ± 0.1g) that were stocked in rectangle plastic baskets. The results indicated that crabs fed with corn starch, wheat starch and dextrin diets had significantly higher final body weight (FBW), weight gain (WG), specific growth rate (SGR) and feed efficiency (FE) than those fed with potato starch diet. However, crabs fed with potato starch diet had the highest feed intake (FI) among all treatments. Crabs fed with corn starch, wheat starch and dextrin diets exhibited significantly higher glycogen in tissue and ATP content in hepatopancreatic than those fed with potato starch diet. The lowest concentration of insulin-like and hyperglycemic hormone were observed in crabs fed with potato starch diet. The lowest relative expression levels of PK, FBpase, PEPCK, GLUT1, GLUT2, GK, G6PDH, 6PGDH, ACC, IGF1R, PI3K and FOXO were occurred at crabs fed with dextrin diet. In conclusion, swimming crab could make better use of corn starch, wheat starch and dextrin, however, potato starch is not suitable as carbohydrate source. The results demonstrated that the ratio of amylose/amylopectin in diets was positively correlated with expression levels of genes related to glycolipid metabolism, glucose transport and insulin pathway.

•Carps fed faba bean had poor growth performance, higher intramuscular collagen content, smaller fibre diameter and wider intermyofibrillar space. However, the fibre density was not influenced.•The faba bean components showed lower crude protein and amino acid imbalance, especially, the deficiency in the sulfuric amino acid.•The raw- fillet and cooked-fillet texture of FBG were both significantly greater than that of CFG.