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The solute carrier family 25 member 1 (Slc25a1)-dependent mitochondrial citrate shuttle is responsible for exporting citrate from the mitochondria to the cytoplasm for supporting lipid biosynthesis and protein acetylation. Previous studies on Slc25a1 concentrated on pathological models. However, the importance of Slc25a1 in maintaining metabolic homeostasis under normal nutritional conditions remains poorly understood. Here, we investigated the mechanism of mitochondrial citrate shuttle in maintaining lipid metabolism homeostasis in male Nile tilapia (Oreochromis niloticus). To achieve the objective, we blocked the mitochondrial citrate shuttle by inhibiting Slc25a1 under normal nutritional conditions. Slc25a1 inhibition was established by feeding Nile tilapia with 250 mg/kg 1,2,3-benzenetricarboxylic acid hydrate for 6 weeks or intraperitoneal injecting them with dsRNA to knockdown slc25a1b for 7 days. The Nile tilapia with Slc25a1 inhibition exhibited an obesity-like phenotype accompanied by fat deposition, liver damage and hyperglycemia. Moreover, Slc25a1 inhibition decreased hepatic citrate-derived acetyl-CoA, but increased hepatic triglyceride levels. Furthermore, Slc25a1 inhibition replenished cytoplasmic acetyl-CoA through enhanced acetate pathway, which led to hepatic triglycerides accumulation. However, acetate-derived acetyl-CoA caused by hepatic Slc25a1 inhibition did not activate de novo lipogenesis, but rather modified protein acetylation. In addition, hepatic Slc25a1 inhibition enhanced fatty acids esterification through acetate-derived acetyl-CoA, which increased Lipin1 acetylation and its protein stability. Collectively, our results illustrate that inhibiting mitochondrial citrate shuttle triggers lipid anabolic remodeling and results in lipid accumulation, indicating the importance of mitochondrial citrate shuttle in maintaining lipid metabolism homeostasis., Competing Interests: Declaration of competing interest None., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Ammonia is a common environmental stress factor that constrains aquaculture industry development. This study evaluated the effect of carbohydrate levels and ammonia stress in oriental river prawn (Macrobrachium nipponense). The experiment had six treatments containing two water ammonia levels (0 and 5 mg/L) and three dietary carbohydrate levels (low carbohydrate diet (LCD, 10%), medium carbohydrate diet [MCD, 20%], and high carbohydrate diet [HCD, 30%]), and lasted six weeks. The results showed that the prawns fed on MCD had higher weight gain than those fed on LCD and HCD during ammonia stress. Moreover, the prawns fed on MCD had significantly lower acid phosphatase and alkaline phosphatase activities during ammonia stress. Feeding the prawns on the MCD increased B cells in the hepatopancreas during ammonia stress. Interestingly, the prawns fed on MCD had significantly lower superoxide dismutase activity compared to LCD and HCD during ammonia stress. Moreover, the prawns fed on MCD had significantly lower pyruvate kinase activity and pyruvate and lactic acid contents, while those fed on LCD had significantly higher succinic dehydrogenase, 6-phosphogluconic dehydrogenase, and phosphoenol pyruvate carboxykinase activities during ammonia stress. The prawns fed on the MCD increased significantly glutaminase activity and decreased the ammonia content in the serum during ammonia exposure. In addition, feeding the prawns on MCD decreased significantly the expression of apoptosis and inflammation-related genes. Taken together, the MCD supplied energy required to counteract ammonia stress, which increased growth, improved antioxidant capacity, facilitated ammonia excretion, and alleviated inflammation and apoptosis of the oriental river prawn., (© 2023 Wiley Periodicals LLC.)

Cyhexatin (CYT), an organotin acaricide, is extensively utilized in developing countries to mitigate plant diseases caused by mites and minimize agricultural crop losses. However, the comprehensive mechanisms underlying the developmental stage of non-target organisms remain largely unexplored. In this study, zebrafish embryos were firstly exposed to CYT (0.06, 0.12, and 0.20 ng/mL, referred to as CYTL, CYTM, and CYTH, respectively) from 2 hpf (hours post fertilization) to 30 dpf (days post fertilization). No developmental toxicity was observed in the CYTL and CYTM groups, except for induced deformed phenotypes in the CYTM group at 120 hpf. However, exposure to CYTH resulted in significant reductions in spontaneous movement (24 hpf), heart rate (48 hpf), hatching rate (48 and 72 hpf), body weight (30 dpf), whole body length (30 dpf), and locomotion (30 dpf). Additionally, CYTH exposure induced morphological malformations, including spinal curvature, pericardial edema, and tail curvature in zebrafish larvae. Moreover, CYTH treatment induced apoptosis, increased reactive oxygen species (ROS) production, and resulted in significant reductions in free T3, cholesterol, estradiol, and testosterone levels in zebrafish larvae, while free T4 levels were increased. RNA-Seq analysis indicated that CYTH exposure led to significant alterations in the genome-wide gene expression profiles of zebrafish, particularly in the thyroid hormone and steroid biosynthesis signaling pathways, indicating endocrine disruption. Furthermore, CYTH exposure induced global DNA hypomethylation, reduced S-adenosylmethionine (SAM) levels and the SAM/S-adenosylhomocysteine (SAH) ratio, elevated SAH levels, and suppressed the mRNA expression of DNA methyltransferases (DNMTs) while also downregulating DNMT1 at both the gene and protein levels in zebrafish larvae. Overall, this study partially elucidated the developmental toxicity and endocrine disruption caused by CYT in zebrafish, providing evidence of the environmental hazards associated with this acaricide., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this study., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Hypoxia is a common environmental stress factor in aquatic organisms, which varies among fish species. However, the mechanisms underlying the ability of fish species to tolerate hypoxia are not well known. Here, we showed that hypoxia response in different fish species was affected by lipid catabolism and preference for lipid or carbohydrate energy sources. Activation of biochemical lipid catabolism through peroxisome proliferator-activated receptor alpha (Pparα) or increasing mitochondrial fat oxidation in tilapia decreased tolerance to acute hypoxia by increasing oxygen consumption and oxidative damage and reducing carbohydrate catabolism as an energy source. Conversely, lipid catabolism inhibition by suppressing entry of lipids into mitochondria in tilapia or individually knocking out three key genes of lipid catabolism in zebrafish increased tolerance to acute hypoxia by decreasing oxygen consumption and oxidative damage and promoting carbohydrate catabolism. However, anaerobic glycolysis suppression eliminated lipid catabolism inhibition-promoted hypoxia tolerance in adipose triglyceride lipase ( atgl ) mutant zebrafish. Using 14 fish species with different trophic levels and taxonomic status, the fish preferentially using lipids for energy were more intolerant to acute hypoxia than those preferentially using carbohydrates. Our study shows that hypoxia tolerance in fish depends on catabolic preference for lipids or carbohydrates, which can be modified by regulating lipid catabolism.

Fish feeding habit determines the digestive tract structure and intestinal microflora. However, the relationship between feeding habit, digestive intestinal morphology, and microbial diversity of omnivorous, herbivorous, plankton feeder, and carnivorous fish from the same environment has not been compared. This study compared the digestive enzyme activities, intestinal morphology, and intestinal microflora of omnivorous ( Carassius auratus ), herbivorous ( Ctenopharyngodon idellus ), carnivorous ( Siniperca chuatsi ), and plankton feeder ( Schizothorax grahami ) fishes and predicted the potential functions of specific microflora on different nutrients. Twelve intestine samples were collected from each of the four fishes from Dianchi Lake. The composition and diversity of microbial communities were determined by using high-throughput sequencing of 16S rDNA. The results showed that the carnivorous fish ( S. chuatsi ) had higher trypsin and pancrelipase activities in the hepatopancreas and enteropeptidase in the intestine, but lower amylase activities in the intestine. The carnivorous fish intestine had more microvilli branches and complex structures than other fish species in the order carnivorous > herbivorous > plankton feeder > omnivorous. The intestinal microflora diversity was higher in the omnivorous fish and followed the order omnivorous > herbivorous > plankton feeder > carnivorous. Acinetobacter species and Bacteroides species were the most dominant flora in the carnivorous and herbivorous fishes, respectively. Acinetobacter species and Pseudomonas species might help the host to digest protein, while Bacteroidetes species may help the host to digest cellulose. Taken together, feeding habit determines the digestive enzyme activities, intestinal tissue morphology, and differential colonization of fish intestinal flora. The knowledge obtained is useful in feed formulation and feeding practices for the studied fish species., Competing Interests: The authors declare that they have no conflicts of interest., (© 2023 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.)

Liver health is important to maintain survival and growth of fish. Currently, the role of dietary docosahexaenoic acid (DHA) in improving fish liver health is largely unknown. This study investigated the role of DHA supplementation in fat deposition and liver damage caused by D-galactosamine (D-GalN) and lipopolysaccharides (LPS) in Nile tilapia (Oreochromis niloticus). Four diets were formulated as control diet (Con), Con supplemented with 1 % DHA, 2 % DHA and 4 % DHA diets, respectively. The diets were fed to 25 Nile tilapia (2.0 ± 0.1 g, average initial weight) in triplicates for four weeks. After the four weeks, 20 fish in each treatment were randomly selected and injected with a mixture of 500 mg D-GalN and 10 μL LPS per mL to induce acute liver injury. The results showed that the Nile tilapia fed on DHA diets decreased visceral somatic index, liver lipid content and serum and liver triglyceride concentrations than those fed on the Con diet. Moreover, after D-GalN/LPS injection, the fish fed on DHA diets decreased alanine aminotransferase and aspartate transaminase activities in the serum. The results of liver qPCR and transcriptomics assays together showed that the DHA diets feeding improved liver health by downregulating the expression of the genes related to toll-like receptor 4 (TLR4) signaling pathway, inflammation and apoptosis. This study indicates that DHA supplementation in Nile tilapia alleviates the liver damage caused by D-GalN/LPS through increasing lipid catabolism, decreasing lipogenesis, TLR4 signaling pathway, inflammation, and apoptosis. Our study provides novel knowledge on the role of DHA in improving liver health in cultured aquatic animals for sustainable aquaculture., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Vitellogenins (Vtgs) are essential for female reproduction in oviparous animals, yet the exact roles and mechanisms remain unknown. In the present study, we knocked out vtg1, which is the most abundant Vtg in zebrafish, Danio rerio via the CRISPR/Cas 9 technology. We aimed to identify the roles of Vtg1 and related mechanisms in reproduction and development. We found that, the Vtg1-deficient female zebrafish reduced gonadosomatic index, egg production, yolk granules and mature follicles in ovary compared to the wide type (WT). Moreover, the Vtg1-deficient zebrafish diminished hatching rates, cumulative survival rate, swimming capacity and food intake, but increased malformation rate, and delayed swim bladder development during embryo and early-larval phases. The Vtg1-deficiency in female broodstock inhibited docosahexaenoic acid-enriched phosphatidylcholine (DHA-PC) transportation from liver to ovary, which lowered DHA-PC content in ovary and offspring during larval stage. However, the Vtg1-deficient zebrafish increased gradually the total DHA-PC content via exogeneous food intake, and the differences in swimming capacity and food intake returned to normal as they matured. Furthermore, supplementing Vtg1-deficient zebrafish with dietary PC and DHA partly ameliorated the impaired female reproductive capacity and larval development during early phases. This study indicates that, DHA and PC carried by Vtg1 are crucial for female fecundity, and affect embryo and larval development through maternal-nutrition effects. This is the first study elucidating the nutrient and physiological functions of Vtg1 and the underlying biochemical mechanisms in fish reproduction and development., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The regulation of cholesterol metabolism in fish is still unclear. Statins play important roles in promoting cholesterol metabolism development in mammals. However, studies on the role of statins in cholesterol metabolism in fish are currently limited. The present study evaluated the effects of statins on cholesterol metabolism in fish. Nile tilapia ( Oreochromis niloticus ) were fed on control diets supplemented with three atorvastatin levels (0, 12, and 24 mg/kg diet, ATV0, ATV12, and ATV24, respectively) for 4 wk. Intriguingly, the results showed that both atorvastatin treatments increased hepatic cholesterol and triglyceride contents mainly through inhibiting bile acid synthesis and efflux, and compensatorily enhancing cholesterol synthesis in fish liver ( P < 0.05). Moreover, atorvastatin treatment significantly inhibited hepatic very-low-density lipoprotein (VLDL) assembly and thus decreased serum VLDL content ( P < 0.05). However, fish treated with atorvastatin significantly reduced cholesterol and triglycerides contents in adipose tissue ( P < 0.05). Further molecular analysis showed that atorvastatin treatment promoted cholesterol synthesis and lipogenesis pathways, but inhibited lipid catabolism and low-density lipoprotein (LDL) uptake in the adipose tissue of fish ( P < 0.05). In general, atorvastatin induced the remodeling of lipid distribution between liver and adipose tissues through blocking VLDL efflux from the liver to adipose tissue of fish. Our results provide a novel regulatory pattern of cholesterol metabolism response caused by atorvastatin in fish, which is distinct from mammals: cholesterol inhibition by atorvastatin activates hepatic cholesterol synthesis and inhibits its efflux to maintain cholesterol homeostasis, consequently reduces cholesterol storage in fish adipose tissue.

Ammonia is one of the common stress factors in aquaculture. However, the effect of chronic ammonia exposure in juvenile oriental river prawn (Macrobrachium nipponense) is currently unexplored. This study explored the effects of chronic ammonia on juvenile healthy oriental river prawns. Fifty prawns (0.123 ± 0.003 g) were exposed to 0, 5, and 15 mg/L total ammonia nitrogen (TAN) in triplicates for 28 days. The effects of chronic ammonia challenge were evaluated on growth, antioxidant capacity, hepatopancreas and gill morphology, and glucose and ammonia metabolism. The results showed that, the chronic ammonia exposure reduced significantly survival rate and weight gain of prawns. The prawns exposed to 15 mg/L ammonia had induced oxidative stress. However, the prawn exposed to 15 mg/L ammonia had significantly lower aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and acid phosphatase activities in the serum. Furthermore, exposure of prawns to 15 mg/L ammonia increased the activities of hexokinase, pyruvate kinase, pyruvate and lactic acid content, and glutamine synthase activity. However, the prawns exposed to 15 mg/L ammonia, reduced succinic dehydrogenase, 6-phosphogluconic dehydrogenase, phosphoenolpyruvate carboxykinase, glutamate synthase, and glutamate dehydrogenase activities but increased ammonia content in serum. The exposure of ammonia deformed lumen, damaged basement membrane and decreased secretory cells in the hepatopancreas, disordered gill epithelial and pillar cells, and caused gill filament base vacuolation. Our study indicates that chronic ammonia stress impairs growth performance, tissue morphology, induces oxidative stress, and alters glucose and ammonia metabolism in juvenile oriental river prawns., (© 2022 Wiley Periodicals LLC.)

The roles of dietary cholesterol in fish physiology are currently contradictory. The issue reflects the limited studies on the metabolic consequences of cholesterol intake in fish. The present study investigated the metabolic responses to high cholesterol intake in Nile tilapia ( Oreochromis niloticus ), which were fed with four cholesterol-contained diets (0.8, 1.6, 2.4 and 3.2%) and a control diet for eight weeks. All fish-fed cholesterol diets showed increased body weight, but accumulated cholesterol (the peak level was in the 1.6% cholesterol group). Then, we selected 1.6% cholesterol and control diets for further analysis. The high cholesterol diet impaired liver function and reduced mitochondria number in fish. Furthermore, high cholesterol intake triggered protective adaptation via (1) inhibiting endogenous cholesterol synthesis, (2) elevating the expression of genes related to cholesterol esterification and efflux, and (3) promoting chenodeoxycholic acid synthesis and efflux. Accordingly, high cholesterol intake reshaped the fish gut microbiome by increasing the abundance of Lactobacillus spp. and Mycobacterium spp., both of which are involved in cholesterol and/or bile acids catabolism. Moreover, high cholesterol intake inhibited lipid catabolic activities through mitochondrial β-oxidation, and lysosome-mediated lipophagy, and depressed insulin signaling sensitivity. Protein catabolism was elevated as a compulsory response to maintain energy homeostasis. Therefore, although high cholesterol intake promoted growth, it led to metabolic disorders in fish. For the first time, this study provides evidence for the systemic metabolic response to high cholesterol intake in fish. This knowledge contributes to an understanding of the metabolic syndromes caused by high cholesterol intake or deposition in fish., Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-022-00158-7., Competing Interests: Conflict of interestAll authors declare no conflicts of interests., (© The Author(s) 2023.)

Hypoxia is one of the challenges in prawns aquaculture. However, the role of thiamine, which is a coenzyme in carbohydrate metabolism with antioxidant properties, in reducing hypoxia in prawns aquaculture is currently unknown. We investigated the effects of thiamine on antioxidant status, carbohydrate metabolism and acute hypoxia in oriental river prawn, Macrobrachium nipponense. One thousand eight hundred prawns (0.123 ± 0.003 g) were fed five diets (60 prawns each tank, six replicates per diet) supplemented with graded thiamine levels (5.69, 70.70, 133.67, 268.33 and 532.00 mg/kg dry mater) for eight weeks and then exposed to hypoxia stress for 12 h followed by reoxyegnation for 12 h. The results showed that, under normoxia, prawns fed the 133.67 or 268.33 mg/kg thiamine diet had significantly lower glucose 6-phosphatedehydrogenase, succinate dehydrogenase and phosphoenolpyruvate carboxykinase activities than those fed the other diets. Moreover, total antioxidant capacity (T-AOC) increased significantly when prawns were fed the 133.67 mg/kg thiamine diet. Superoxide dismutase (SOD) activity and malonaldehyde (MDA) content also increased significantly when prawns were fed the 268.33 or 532.00 mg/kg thiamine diet under hypoxia. And the significantly increased SOD activity and MDA level also observed in prawns fed 532.00 mg/kg thiamine under reoxygenation. Under normoxia, prawns fed the 70.70 or 133.67 mg/kg thiamine diet decreased the mRNA expressions of AMP-activated protein kinase-alpha (AMPK-α), pyruvate dehydrogenase-E1-α subunit (PDH-E1-α) and hypoxia-inducible factor-1s (HIF-1α, HIF-1β), but increased the mRNA expressions of phosphofructokinase (PFK) significantly. After 12 h of hypoxia, the energy metabolism related genes (AMPK-β, AMPK-γ, PFK, PDH-E1-α), hypoxia-inducible factor related genes (HIF-1α, HIF-1β) and thiamine transporter gene (SLC19A2) were up-regulated significantly in prawns fed the 133.67 or 268.33 mg/kg thiamine diets. After 12 h of reoxygenation, prawns fed the 133.67 or 268.33 mg/kg diet significantly decreased the SOD activity, MDA level and SLC19A2 mRNA expression compared with other diets. The optimum thiamine was 161.20 mg/kg for minimum MDA content and 143.17 mg/kg for maximum T-AOC activity based on cubic regression analysis. In summary, supplementing 143.17 to 161.20 mg/kg thiamine in the diets for M. nipponense improves the antioxidant capacity under normoxia and reduces the oxidative damage under hypoxia stress., (Copyright © 2022. Published by Elsevier Ltd.)

The wide use of high-fat diet (HFD) causes negative effects on flesh quality in farmed fish. l-carnitine, a lipid-lowering additive, enhances mitochondrial fatty acid β-oxidation. However its roles in alleviating the effects of HFD on flesh quality in fish are unknown. We fed Nile tilapia with medium-fat diet (MFD, 6% dietary lipid), high-fat diet (HFD, 12% dietary lipid) and HFCD supplemented with l-carnitine (HFCD + 400 mg/kg l-carnitine) for 10 weeks. The HFD-fed fish had higher fat deposition, pH value, myofiber density and flesh hardness than those fed on MFD. However, feeding the fish with the HFCD improved lipid catabolism, which increased significantly lactic acid content and myofiber diameter in muscle, thus reduced pH and hardness values. HFCD also reduced endoplasmic reticulum stress and myofiber apoptosis caused by HFD in the fish. Our study suggests that dietary l-carnitine supplementation alleviates the negative effects of HFD on flesh quality of farmed fish., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Pyruvate dehydrogenase kinases (PDKs)-pyruvate dehydrogenase E1α subunit (PDHE1α) axis plays an important role in regulating glucose metabolism in mammals. However, the regulatory function of PDKs-PDHE1α axis in the glucose metabolism of fish is not well known. This study determined whether PDKs inhibition could enhance PDHE1α activity, and improve glucose catabolism in fish. Nile tilapia fingerlings (1.90 ± 0.11 g) were randomly divided into 4 treatments in triplicate (30 fish each) and fed control diet without dichloroacetate (DCA) (38% protein, 7% lipid and 45% corn starch) and the control diet supplemented with DCA, which inhibits PDKs through binding the allosteric sites, at 3.75 (DCA3.75), 7.50 (DCA7.50) and 11.25 g/kg (DCA11.25), for 6 wk. The results showed that DCA3.75, DCA7.50 and DCA11.25 significantly increased weight gain, carcass ratio and protein efficiency ratio ( P  < 0.05) and reduced feed efficiency ( P  < 0.05) of Nile tilapia. To investigate the effects of DCA on growth performance of Nile tilapia, we selected the lowest dose DCA3.75 for subsequent analysis. Nile tilapia fed on DCA3.75 significantly reduced the mesenteric fat index, serum and liver triglyceride concentration and total lipid content in whole fish, and down-regulated the expressions of genes related to lipogenesis ( P  < 0.05) compared to the control. The DCA3.75 treatment significantly improved glucose oxidative catabolism and glycogen synthesis in the liver, but significantly reduced the conversion of glucose to lipid ( P  < 0.05). Furthermore, the DCA3.75 treatment significantly decreased the PDK2/4 gene and protein expressions ( P  < 0.05), accordingly stimulated PDHE1α activity by decreasing the phosphorylated PDHE1α protein level. In addition, DCA3.75 treatment significantly increased the phosphorylated levels of key proteins involved in insulin signaling pathway and glycogen synthase kinase 3β ( P  < 0.05). Taken together, the present study demonstrates that PDK2/4 inhibition by using DCA promotes glucose utilization in Nile tilapia by activating PDHE1α and improving insulin sensitivity. Our study helps to understand the regulatory mechanism of glucose metabolism for improving dietary carbohydrate utilization in farmed fish., Competing Interests: We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, and there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the content of this paper., (© 2022 Chinese Association of Animal Science and Veterinary Medicine. Publishing services by Elsevier B.V. on behalf of KeAi Communications Co. Ltd.)

The demand for collagen has been increasing over years due to its wide application in food, cosmetics and biomedicine industries. The synthesis of collagen protein in fish depends on instructions provided by collagen, type I, alpha 1 (COL1A1) gene. However, cloning, tissue distribution and mRNA expression of COL1A1 gene in a gel-producing Chu's croaker (Nibea coibor) is currently unknown. This study cloned the cDNA of COL1A1 gene (GenBank accession number: MK641512) from six N. coibor fish. The distribution and mRNA expression pattern of COL1A1 was analyzed in eight tissues of N. coibor. The COL1A1 cDNA had a full length of 6130 bp and contained a 4344 bp open reading frame (ORF) encoding a polypeptide of 1448 amino acids. The homology of N. coibor COL1A1 amino acid had 98% similarity with Larimichthys crocea, indicating conservatism with other members in same family (Sciaenidae). The deduced polypeptide contained the same signal peptides, C-propeptide and N-propeptide domains, and triple helix domains, which are the characteristics of type I collagen in vertebrates. The mRNA of COL1A1 gene was expressed significantly higher in the spine of N. coibor than in all other tissues (P < 0.05), followed by swim bladder, skin and scales. The swim bladder had higher collagen and hydroxyproline contents than other tissues, followed by spine >, scales > and > skin (P < 0.05). Our study successfully cloned the COL1A1 gene from N. coibor for the first time. The COL1A1 gene contained all the features of collagen pro-α1(I) chain proteins, and shared high homology with other marine teleost. COL1A1 gene in N. coibor is highly expressed in spine and swim bladder, consistent with collagen distribution. Our study contributes to better understanding on collagen biosynthesis in N. coibor tissues for various industrial uses., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Neutral lipases-mediated lipolysis and acid lipases-moderated lipophagy are two main processes for degradation of lipid droplets (LDs). However, the individual and interactive roles of these metabolic pathways are not well known across vertebrates. This study explored the roles of lipolysis and lipophagy from the aspect of neutral and acid lipases in zebrafish. We established zebrafish strains deficient in either adipose triglyceride lipase (atgl -/- ; AKO fish) or lysosomal acid lipase (lal -/- ; LKO fish) respectively, and then inhibited lipolysis in the LKO fish and lipophagy in the AKO fish by feeding diets supplemented with the corresponding inhibitors Atglistatin and 3-Methyladenine, respectively. Both the AKO and LKO fish showed reduced growth, swimming activity, and oxygen consumption. The AKO fish did not show phenotypes in adipose tissue, but mainly accumulated triacylglycerol (TAG) in liver, also, they had large LDs in the hepatocytes, and did not stimulate lipophagy as a compensation response but maintained basal lipophagy. The LKO fish reduced total lipid accumulation in the body but had high cholesterol content in liver; also, they accumulated small LDs in the hepatocytes, and showed increased lipolysis, especially Atgl expression, as a compensatory mechanism. Simultaneous inhibition of lipolysis and lipophagy in zebrafish resulted in severe liver damage, with the potential to trigger mitophagy. Overall, our study illustrates that lipolysis and lipophagy perform individual and interactive roles in maintaining homeostasis of TAG and cholesterol metabolism. Furthermore, the interactive roles of lipolysis and lipophagy may be essential in regulating the functions and form of mitochondria., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL)-mediated lipolysis play important roles in lipid catabolism. ATGL is considered the central rate-limiting enzyme in the mobilization of fatty acids in mammals. Currently, severe fat accumulation has been commonly detected in farmed fish globally. However, the ATGL-mediated lipolysis and the potential synergy among ATGL, HSL, and autophagy, which is another way for lipid breakdown, have not been intensively understood in fish. In the present study, we added Atglistatin as an ATGL-specific inhibitor into the zebrafish diet and fed to the fish for 5 weeks. The results showed that the Atglistatin-treated fish exhibited severe fat deposition, reduced oxygen consumption, and fatty acid β-oxidation, accompanied with increased oxidative stress and inflammation. Furthermore, the Atglistatin-treated fish elevated total and phosphorylation protein expressions of HSL. However, the free fatty acids and lipase activities in organs were still systemically reduced in the Atglistatin-treated fish, and the autophagy marker LC3 was also decreased in the liver. On the other hand, glycogenolysis was stimulated but blood glucose was higher in the Atglistatin-treated fish. The transcriptomic analysis also provided the hint that the protein turnover efficiency in Atglistatin-treated fish was likely to be accelerated, but the protein content in whole fish was not affected. Taken together, ATGL plays crucial roles in energy homeostasis such that its inhibition causes loss of lipid-sourced energy production, which cannot be compensated by activation of HSL, autophagy, and utilization of other nutrients.

Background: Fish cannot use carbohydrate efficiently and instead utilize protein for energy supply, thus limiting dietary protein storage. Protein deposition is dependent on protein turnover balance, which correlates tightly with cellular energy homeostasis. Mitochondrial fatty acid β-oxidation (FAO) plays a crucial role in energy metabolism. However, the effect of remodeled energy homeostasis caused by inhibited mitochondrial FAO on protein deposition in fish has not been intensively studied., Objectives: This study aimed to identify the regulatory role of mitochondrial FAO in energy homeostasis maintenance and protein deposition by studying lipid, glucose, and protein metabolism in fish., Methods: Carnitine-depleted male Nile tilapia (initial weight: 4.29 ± 0.12 g; 3 mo old) were established by feeding them with mildronate diets (1000 mg/kg/d) for 6 wk. Zebrafish deficient in the carnitine palmitoyltransferase 1b gene (cpt1b) were produced by using CRISPR/Cas9 gene-editing technology, and their males (154 ± 3.52 mg; 3 mo old) were used for experiments. Normal Nile tilapia and wildtype zebrafish were used as controls. We assessed nutrient metabolism and energy homeostasis-related biochemical and molecular parameters, and performed 14C-labeled nutrient tracking and transcriptomic analyses., Results: The mitochondrial FAO decreased by 33.1-88.9% (liver) and 55.6-68.8% (muscle) in carnitine-depleted Nile tilapia and cpt1b-deficient zebrafish compared with their controls (P < 0.05). Notably, glucose oxidation and muscle protein deposition increased by 20.5-24.4% and 6.40-8.54%, respectively, in the 2 fish models compared with their corresponding controls (P < 0.05). Accordingly, the adenosine 5'-monophosphate-activated protein kinase/protein kinase B-mechanistic target of rapamycin (AMPK/AKT-mTOR) signaling was significantly activated in the 2 fish models with inhibited mitochondrial FAO (P < 0.05)., Conclusions: These data show that inhibited mitochondrial FAO in fish induces energy homeostasis remodeling and enhances glucose utilization and protein deposition. Therefore, fish with inhibited mitochondrial FAO could have high potential to utilize carbohydrate. Our results demonstrate a potentially new approach for increasing protein deposition through energy homeostasis regulation in cultured animals., (Copyright © The Author(s) on behalf of the American Society for Nutrition 2020.)

Environmental estrogens, including bisphenol A (BPA) and 17β-estradiol (E2), which are widely used in industries and medicine, pose a severe ecological threat to fish due to feminization induction. However, the related metabolic basis for reproductive feminization in male fish has not been well addressed. We first found that female zebrafish exhibited higher lipid accumulation and lipogenesis activity than males. Next, we exposed male and female zebrafish to E2 (200 ng/L) or BPA (100 μg/L) for six weeks, and observed an early-phase reproductive feminization in males, accompanied with reduced spermatids, significant fat deposition and lipogenic gene expressions that mimicked female patterns. Cellular signaling assays revealed that, E2 or BPA modulated lipid metabolism in males mainly through lowering 5' AMP-activated protein kinase (AMPK) and upregulating the lipogenic mechanistic target of rapamycin (mTOR) pathways. For the first time, we show that environmental estrogens could alter lipid metabolism in male fish to a female pattern (metabolic feminization) prior to gonad feminization in male fish, to allows males to accumulate efficiently lipids to harmonize with the feminized gonads. This study suggests that negative effects of environmental estrogens, as hazardous materials, on vertebrate health are more complicated than originally thought., Competing Interests: Declaration of Competing Interest All authors declare that we do not have any commercial or associative interest that represents a conflict of interest in connection with the work submitted., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Antibiotics used in global aquaculture production cause various side effects, which impair fish health. However, the use of dietary composition such as carbohydrate, which is one of the dominant components in fish diets to attenuate the side effects induced by antibiotics, remains unclear. We determined the ability of high carbohydrate diet to protect Nile tilapia (Oreochromis niloticus) from oxytetracycline-induced side effects. Triplicate groups of thirty O. niloticus (9.50 ± 0.08 g) were fed on medium carbohydrate (MC; 335 g/kg) and high carbohydrate (HC; 455 g/kg) diets without and with 2.00 g/kg diet of oxytetracycline (80 mg/kg body weight/day) hereafter, MCO and HCO for 35 days. Thereafter, we assessed growth performance, hepatic nutrients composition and metabolism, microbiota abundance, immunity, oxidative and cellular stress, hepatotoxicity, lipid peroxidation and apoptosis. To understand the possible mechanism of carbohydrate protection on oxytetracycline, we assessed the binding effects and efficiencies of mixtures of medium and high starch with oxytetracycline as well as the MCO and HCO diets. The O. niloticus fed on the MCO and HCO diets had lower growth rate, nutrients utilization and survival rate than those fed on the MC and HC diets, respectively. Dietary HCO increased hepatosomatic index and hepatic protein content of O. niloticus than MCO diet. The O. niloticus fed on the HCO diet had lower mRNA expression of genes related to protein, glycogen and lipid metabolism compared to those fed on the MCO diet. Feeding O. niloticus on the HCO diet increased innate immunity and reduced pathogenic bacteria, pro-inflammation, hepatotoxicity, cellular stress and apoptosis than the MCO diet. The high starch with oxytetracycline and HCO diet had higher-oxytetracycline binding effects and efficiencies than the medium starch with oxytetracyline and MCO diet, respectively. Our study demonstrates that, high carbohydrate partially protects O. niloticus from oxytetracycline-induced side effects by binding the antibiotic. Incorporating high carbohydrate in diet formulation for omnivorous fish species alleviates some of the side effects caused by antibiotics., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Environmental estrogenic compounds are important pollutants, which are widely distributed in natural water bodies. They produce various adverse effects on fish, but their concentration-dependent toxicities in fish metabolism and health are not fully understood. This study investigated the effects of 17β-estradiol (E2) and bisphenol A (BPA) at low and high concentrations on lipid deposition, inflammation and antioxidant response in male zebrafish. We measured fish growth parameters, gonad development, lipid contents and the activities of inflammatory and antioxidant enzymes, as well as their mRNA expressions. All E2 and BPA concentrations used increased body weight, damaged gonad structure and induced feminization in male zebrafish. The exposure of zebrafish to E2 and BPA promoted lipid accumulation by increasing total fat, liver triglycerides and free fatty acid contents, and also upregulated lipogenic genes expression, although they decreased total cholesterol content. Notably, zebrafish exposed to low concentrations of E2 (200 ng/L) and BPA (100 μg/L) had higher lipid synthesis and deposition compared to high concentrations (2000 ng/L and 2000 μg/L, respectively). However, the high concentrations of E2 and BPA increased inflammation and antioxidant response. Furthermore, BPA caused greater damage to fish gonad development and more severe lipid peroxidation compared to E2. Overall, the results suggest that the toxic effects of E2 and BPA on zebrafish are concentration-dependent such that, the relative low concentrations used induced lipid deposition, whereas the high ones caused adverse effects on inflammation and antioxidant response., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

l-Carnitine is essential for mitochondrial β-oxidation and has been used as a lipid-lowering feed additive in humans and farmed animals. d-Carnitine is an optical isomer of l-carnitine and dl-carnitine has been widely used in animal feeds. However, the functional differences between l- and d-carnitine are difficult to study because of the endogenous l-carnitine background. In the present study, we developed a low-carnitine Nile tilapia model by treating fish with a carnitine synthesis inhibitor, and used this model to investigate the functional differences between l- and d-carnitine in nutrient metabolism in fish. l- or d-carnitine (0·4 g/kg diet) was fed to the low-carnitine tilapia for 6 weeks. l-Carnitine feeding increased the acyl-carnitine concentration from 3522 to 10 822 ng/g and alleviated the lipid deposition from 15·89 to 11·97 % in the liver of low-carnitine tilapia. However, as compared with l-carnitine group, d-carnitine feeding reduced the acyl-carnitine concentration from 10 822 to 5482 ng/g, and increased lipid deposition from 11·97 to 20·21 % and the mRNA expression of the genes involved in β-oxidation and detoxification in the liver. d-Carnitine feeding also induced hepatic inflammation, oxidative stress and apoptosis. A metabolomic investigation further showed that d-carnitine feeding increased glycolysis, protein metabolism and activity of the tricarboxylic acid cycle and oxidative phosphorylation. Thus, l-carnitine can be physiologically utilised in fish, whereas d-carnitine is metabolised as a xenobiotic and induces lipotoxicity. d-Carnitine-fed fish demonstrates increases in peroxisomal β-oxidation, glycolysis and amino acid degradation to maintain energy homeostasis. Therefore, d-carnitine is not recommended for use in farmed animals.

Colonizing germ-free (GF) zebrafish with specific bacterial species provides the possibility of understanding the influence on host biological processes including gene expression, development, immunity, and behavioral responses. It also enlightens our understanding on the host-microbe interactions within the physiological context of a living host. However, the responses of GF zebrafish to various colonization conditions such as bacterial concentrations, colonization time points, and exposure duration remain unclear. To address this issue, we explored the responses of GF zebrafish by using two bacterial species at varying concentrations, colonization time points and exposure duration. Therefore, we mono-associated GF zebrafish with Escherichia coli DH5α or Bacillus subtilis WB800N at concentrations ranging from 10 2 to 10 7 CFU/ml either at 3 day post fertilization (dpf) or 5 dpf for 24 or 48 h. We evaluated the responses of GF zebrafish by analyzing the survival rate, colonization efficiency, nutrients metabolism, intestinal cell proliferation, innate immunity, stress, and behavior responses by comparing it to conventionally raised zebrafish (CONR) and GF zebrafish. The results indicated that the final bacteria concentrations ranging from 10 2 to 10 4 CFU/ml did not cause any mortality when GF mono-associated larvae were exposed to either E. coli DH5α or B. subtilis WB800N at 3 or 5 dpf, while concentrations ranging from 10 6 to 10 7 CFU/ml increased the mortality, particularly for 5 dpf owing to the decrease in dissolved oxygen level. The E. coli DH5α mainly induced the expression of genes related to nutrients metabolism, cell proliferation and immunity, while B. subtilis WB800N mainly upregulated the expression of genes related to immunity and stress responses. Moreover, our data revealed that GF zebrafish showed higher levels of physical activity than CONR and the microbial colonization reduced the hyperactivity of GF zebrafish, suggesting colonization of bacteria affected behavior characteristics. This study provides useful information on bacterial colonization of GF zebrafish and the interaction between the host and microbiota., (Copyright © 2019 Tan, Limbu, Qian, Qiao, Du and Zhang.)

Antibiotics and high fat diets are commonly used independently in global aquaculture production for fish health management and to spare the use of costly protein as energy sources, respectively, causing relatively similar metabolic effects and stresses. However, it is unknown whether dietary high fat worsens or attenuates the adverse effects caused by antibiotics in fish. We determined the ability of high fat diet to influence the adverse effects of oxytetracycline on Nile tilapia, Oreochromis niloticus. Thirty Nile tilapia weighing 8.45 ± 0.15 g were fed on medium fat (MF; 70 g/kg) and high fat (HF; 120 g/kg) diets and the same fat levels supplemented with 2.00 g/kg diet of OTC (80 mg/kg body weight/day) hereafter, MFO and HFO for 65 days. The general growth performance, feed efficiency and intestinal health of fish were evaluated. The Nile tilapia fed on HFO diet had significantly lower growth rate, body protein content and feed efficiency compared to those fed on MFO diet. Dietary HFO affected the intestine histomorphology, which decreased dramatically the tight junction proteins of Nile tilapia and induced microbiota dysbiosis compared to MFO diet. The Nile tilapia fed on HFO diet had increased oxidative stress, which stimulated drug detoxification response, caused endoplasmic reticulum stress and apoptosis compared to those fed on MFO diet. The new findings from our study demonstrate that, the adverse effects of antibiotics in fish are different at medium and high fat contents. Feeding fish with high fat diets with antibiotics worsen the adverse effects. This enlightens our understanding on the risks of antibiotics misuse and also suggests that antibiotics should be more strictly limited in aquaculture, in which high fat diets are currently widely used in fish production worldwide., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Lipophagy degrades lipid droplets (LDs) through the lysosomal degradative pathway, thus plays important roles in regulating lipid metabolism in mammals. However, information on the existence and functions of lipophagy in fish lipid metabolism is still limited. In the present study, we confirmed the existence of lipophagy by observing the structures of LDs sequestered in autophagic vacuoles in the zebrafish liver cell line (ZFL) via electronic microscopy. Moreover, starved cells increased the mRNA expression of the microtubule-associated protein 1A/1B light chain 3 beta ( LC3 ), which is a marker protein for autophagy and protein conversion from LC3-I to LC3-II. Inhibiting autophagy with chloroquine increased significantly the LDs content and decreased fatty acid β-oxidation and esterification activities in the ZFL cells cultured in the fed state. Furthermore, inhibiting autophagy function downregulated the mRNA expression of the genes and their proteins related to lipid metabolism. Altogether, the present study verified the existence of lipophagy and its essential regulatory roles in lipid metabolism in fish cells.

In the present paper, we investigated the molecular cloning and functional characterization of elongase of very long chain fatty acid (elovl) and fatty acyl desaturase (fads) genes in a marine teleost, Nibea diacanthus. The elongase cDNA sequence encoded a polypeptide of 294 amino acids exhibiting Elovl5 activity, which effectively elongated both C18 (18:2n-6, 18:3n-3 and 18:3n-6) and C20 (20:4n-6 and 20:5n-3) polyunsaturated fatty acids. The desaturase cDNA sequence specified a polypeptide of 445 amino acids indicating Δ6 desaturation activity, which coul converted C18:2n-6 and C18:3n-3 to C18:3n-6 and C18:4n-3, respectively. Tissue distribution analysis by quantitative real-time PCR revealed that the elovl5 was primarily expressed in intestine and liver, while the fads2 was mainly expressed in liver and brain. These results increase our knowledge of the ability of endogenous highly unsaturated fatty acids (HUFA) biosynthesis in marine carnivorous fish., (Copyright © 2019 Elsevier B.V. All rights reserved.)

High fat diets are commonly used in aquaculture to reduce feed cost in Nile tilapia, but impair its lipid homeostasis. This study evaluated the role of forskolin on reducing fat accumulation in Nile tilapia (Oreochromis niloticus) by using in vitro and in vivo experiments. The use of 50 μM forskolin in vitro increased free fatty acid and glycerol release, but decreased triglyceride in adipocytes and hepatocytes. The adipose triglyceride lipase (ATGL), protein kinase cAMP-dependent type I regulatory subunit alpha (PKAR I) and other genes related to β-oxidation (peroxisome proliferator activated receptor alpha, PPARα and carnitine O-palmitoyltransferase 1, CPT1) were significantly up-regulated. After feeding a high-fat diet for six weeks, O. niloticus were fed with 0 (control), 0.5 and 1.5 mg/kg forskolin for two weeks to determine whether forskolin could reduce fat accumulation in vivo. Fish fed the two levels of forskolin decreased significantly the hepatosomatic and mesenteric fat indices. The total lipid in the whole fish and liver together with the serum glycerol content were lower in fish fed on forskolin than in the control. The fish fed on forskolin diets exhibited smaller areas of lipid droplets in adipose and liver tissues. Lipolysis related genes (ATGL, hormone-sensitive lipase, HSL; monoacylglycerol lipase, MGL; and protein kinase cAMP-activated catalytic subunit, PKAC) and β-oxidation genes (PPARα; fatty acid binding protein 1, FABP1; and CPT1) in the adipose were up-regulated. Similarly, in the liver lipolysis genes such as ATGL and PKAR I and β-oxidation genes (PPARα, FABP1, CPT1 and acyl-CoA oxidase, ACO) showed an increasing trend with the increase of forskolin doses. This study indicates that forskolin can reduce fat accumulation in the adipose and liver by stimulating lipolysis and β-oxidation in O. niloticus., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Dietary α-lipoic acid (LA), β-glucan (Gluc) and l-carnitine (L-Ca) are commonly used additives to promote fish growth and stress resistance in aquaculture production. However their mechanisms and efficiencies in helping fish to resist diseases have not been compared before. In this study, we fed Nile tilapia (Oreochromis niloticus) with diets containing appropriate doses of LA, Gluc and L-Ca for five weeks and further intraperitoneally injected the fish with Aeromonas hydrophila. After dietary treatment, none of the additives affected the fish growth, but dietary Gluc and L-Ca reduced protein and lipid body contents in fish, respectively. After A. hydrophila challenge, all fish treated with the three dietary additives showed higher survival rate, but those fed on dietary L-Ca had lower survival than those fed on LA and Gluc diets, indicating high protection efficiency of LA and Gluc. The protective mechanisms of the three feed additives were quite different under A. hydrophila infection. Dietary LA induced higher total antioxidant capacity and higher mRNA expression of anti-oxidative genes than other additives in liver and also activated partly the immune function in serum and spleen. Gluc largely increased the immune function by activating the immunity enzymes in serum, inducing inflammation in liver and increasing the expression of immune genes in spleen and head kidney. Gluc also increased partly the antioxidant capacity in serum and liver and lipid catabolism in liver. L-Ca largely increased lipid catabolism in liver while it increased partly the antioxidant capacities in serum and liver. Taken together, these results indicate that, dietary LA, Gluc and L-Ca have various protective mechanisms and differ in their efficiencies on resisting A. hydrophila infection in Nile tilapia., (Copyright © 2018. Published by Elsevier Ltd.)

Key Points: In a cold environment, mammals increase their food intake while fish decrease or stop feeding. However, the physiological value of fasting during cold resistance in fish is currently unknown. Fasting for more than 48 h enhanced acute cold resistance in zebrafish, which correlated with lipid catabolism and cell damage attenuation. Lipid catabolism and autophagy were necessary for cold resistance in fish and the inhibition of mitochondrial fatty acid β-oxidation or autophagy weakened the fasting-induced cold resistance. Repression of mechanistic target of rapamycin (mTOR) signalling pathway by rapamycin largely mimicked the beneficial effects of fasting in promoting cold resistance, suggesting mTOR signalling may be involved in the fasting-induced cold resistance in fish. Our study demonstrates that fasting may be a protective strategy for fish to survive under cold stress., Abstract: In cold environments, most homeothermic animals increase their food intake to supply more energy to maintain body temperature, whereas most poikilothermic animals such as fishes decrease or even stop feeding under cold stress. However, the physiological value of fasting during cold resistance in poikilotherms has not been explained. Here, we show that moderate fasting largely enhanced cold resistance in fish. By using pharmacological (fenofibrate, mildronate, chloroquine and rapamycin) and nutritional approaches (fatty acids diets and amino acids diets) in wild-type or specific gene knock-out zebrafish models (carnitine palmitoyltransferase-1b-deficient strain, CPT1b -/- , or autophagy-related protein 12-deficient strain, ATG12 -/- ), we verified that fasting-stimulated lipid catabolism and autophagy played essential roles in the improved cold resistance. Moreover, suppression of the mechanistic target of rapamycin (mTOR) pathway by using rapamycin mostly mimicked the beneficial effects of fasting in promoting cold resistance as either the physiological phenotype or transcriptomic pattern. However, these beneficial effects were largely reduced when the mTOR pathway was activated through high dietary leucine supplementation. We conclude that fasting helps fish to resist cold stress by modulating lipid catabolism and autophagy, which correlates with the mTOR signalling pathway. Therefore, fasting can act as a protective strategy of fish in resisting coldness., (© 2019 The Authors. The Journal of Physiology © 2019 The Physiological Society.)

Dietary fish oil used in aquafeed transfers marine pollutants to farmed fish. However, the entire transfer route of marine pollutants in dietary fish oil from ocean to table fish has not been tracked quantitatively. To track the entire transfer route of marine pollutants from wild fish to farmed fish through dietary fish oil and evaluate the related human health risks, we obtained crude and refined fish oils originating from the same batch of wild ocean anchovy and prepared fish oil-containing purified aquafeeds to feed omnivorous lean Nile tilapia and carnivorous fatty yellow catfish for eight weeks. The potential human health risk of consumption of these fish was evaluated. Marine persistent organic pollutants (POPs) were concentrated in fish oil, but were largely removed by the refining process, particularly dioxins and polychlorinated biphenyls (PCBs). The differences in the POP concentrations between crude and refined fish oils were retained in the fillets of the farmed fish. Fillets fat content and fish growth were positively and negatively correlated to the final POPs deposition in fillets, respectively. The retention rates of marine POPs in the final fillets through fish oil-contained aquafeeds were 1.3%-5.2%, and were correlated with the POPs concentrations in feeds and fillets, feed utilization and carcass ratios. The dietary crude fish oil-contained aquafeeds are a higher hazard ratio to consumers. Prohibiting the use of crude fish oil in aquafeed and improving growth and feed efficiency in farmed fish are promising strategies to reduce health risks originating from marine POPs., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The use of antibiotics for anti-infection and growth promotion has caused the overuse of antibiotics in aquaculture. However, the benefit or risk of the long-term use of antibiotics on fish growth or health has not been fully addressed. In the present study, zebrafish were fed with sulfamethoxazole (SMX) or oxytetracycline (OTC) at the therapeutic concentrations (100 and 80 mg/kg body weight per day, respectively) for 6 weeks to mimic the long-term use of antibiotics. The digestive enzyme activities were higher in both antibiotic treatments, and higher oxygen consumption rate was found in OTC treated group. As a result, SMX increased the weight gain of zebrafish, and OTC treatment did not show significant prompting effect on growth. The mortality was higher in SMX or OTC treated group on 2nd-4th day after exposure to Aeromonas hydrophila. Lower alkaline phosphatase (AKP) and acid phosphatase (ACP) activities were found in OTC treated group, while higher malondialdehyde (MDA) content was found in the intestine of both SMX and OTC treated zebrafish. Furthermore, feeding OTC decreased the intestinal microbial richness. This study revealed that long-term use of legal aquaculture concentrations of antibiotics caused systemic adverse effects on fish gut health; stringent policy for use of antibiotics in fish is urgent.

Background: Antibiotics used globally to treat human and animal diseases exist ubiquitously in the environment at low doses because of misuse, overdose and poor absorption after ingestion, coupled with their high-water solubility and degradation resistance. However, the systemic chronic effects of exposure to low environmental concentrations of antibiotics (LECAs) and legal aquaculture doses of antibiotics (LADAs) in fish and their human health risk are currently unknown., Objective: To investigate the in vivo chronic effects of exposure to LECAs and LADAs using oxytetracycline (OTC) and sulfamethoxazole (SMZ) in Nile tilapia (Oreochromis niloticus) and their human health risk., Methods: Twenty O. niloticus weighing 27.73 ± 0.81 g were exposed to water containing LECAs (OTC at 420 ng/L and SMZ at 260 ng/L) and diets supplemented with LADAs (OTC 80 mg/kg/day and SMZ 100 mg/kg/day) for twelve weeks. General physiological functions, metabolic activities, intestinal and hepatic health were systemically evaluated. The possible human health risks of the consumption of the experimental Nile tilapia fillets in adults and children were assessed by using risk quotient., Results: After exposure, we observed retarded growth performance accompanied by reduced nutrients digestibility, feed efficiency, organ indices, and lipid body composition in treated fish. Antibiotics distorted intestinal morphological features subsequently induced microbiota dysbiosis and suppressed intestinal tight junction proteins. Exposure of fish to LECAs and LADAs induced oxidative stress, suppressed innate immunity, stimulated inflammatory and detoxification responses, concomitantly inhibited antioxidant capacity and caused lipid peroxidation in intestine and liver organs. Both LECAs and LADAs enhanced gluconeogenesis, inhibited lipogenesis and fatty acid beta oxidation in intestine and liver organs. The exposure of fish to LECAs and LADAs induced anaerobic glycolytic pathway and affected intestinal fat catabolism in intestine while halted aerobic glycolysis, increased hepatic fat catabolism, and induced DNA damage in liver. The hazard risk quotient in children for fish treated with OTCD was >1 indicating human health risk., Conclusion: Overall, both LECAs and LADAs impair general physiological functions, nutritional metabolism, and compromise fish immune system. Consumption of fish fed with legal OTC provokes health risk in children. Global stringent prohibition policy for use of antibiotics in aquaculture production and strategies to limit their release into the environment are urgently required to protect human health., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Understanding how intestinal microbiota alters energy homeostasis and lipid metabolism is a critical process in energy balance and health. However, the exact role of intestinal microbiota in the regulation of lipid metabolism in fish remains unclear. Here, we used two zebrafish models (germ-free and antibiotics-treated zebrafish) to identify the role of intestinal microbiota in lipid metabolism. Conventional and germ-free zebrafish larvae were fed with egg yolk. Transmission electron microscopy was used to detect the presence of lipid droplets in the intestinal epithelium. The results showed that, microbiota increased lipid accumulation in the intestinal epithelium. The mRNA sequencing technology was used to assess genes expression level. We found majority of the differentially expressed genes were related to lipid metabolism. Due to the limitation of germ-free zebrafish larvae, antibiotics-treated zebrafish were also used to identify the relationship between the gut microbiota and the host lipid metabolism. Oil-red staining showed antibiotics-treated zebrafish had less intestinal lipid accumulation than control group. The mRNA expression of genes related to lipid metabolism in liver and intestine was also quantified by using real-time PCR. The results indicated that apoa4 , hsl , cox15 , slc2a1a , and lss were more related to intestinal bacteria in fish, while the influence of intestinal microbiota on the activity of fabp6 , acsl5 , cd36 , and gpat2 was different between the liver and intestine. This study identified several genes regulated by intestinal microbiota. Furthermore, the advantages and disadvantages of each model have been discussed. This study provides valuable information for exploring host-microbiota interactions in zebrafish in future.

Insulin-like growth factor-1 (IGF-1) plays a crucial role in regulating growth in vertebrates whereas suppressors of cytokine signaling (SOCS) act as feedback inhibitors of the GH/IGF-1 axis. Although SOCS-2 binds the IGF-1 receptor and inhibits IGF-1-induced STAT3 activation, presently there is no clear evidence as to whether IGF-1 could induce SOCS gene expression. The current study aimed to determine whether IGF-1 could induce the transcription of SOCS in juvenile Nile tilapia ( Oreochromis niloticus ). We show that there is a common positive relationship between the mRNA expression of IGF-I and SOCS-2 under different nutritional statuses and stimulants, but not the mRNA expression of SOCS-1 and SOCS-3 Furthermore, rhIGF-1 treatment and transcriptional activity assay confirmed the hypothesis that IGF-1 could induce SOCS-2 expression, whereas it had no effect or even decreased the expression of SOCS-1 and SOCS-3 Overall, we obtained evidence that the transcription of SOCS-2, but not SOCS-1 or SOCS-3, could be induced by IGF signaling, suggesting that SOCS-2 serves as a feedback suppressor of the IGF-1 axis in juvenile Nile tilapia., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)

Impaired mitochondrial fatty acid β-oxidation has been correlated with many metabolic syndromes, and the metabolic characteristics of the mammalian models of mitochondrial dysfunction have also been intensively studied. However, the effects of the impaired mitochondrial fatty acid β-oxidation on systemic metabolism in teleost have never been investigated. In the present study, we established a low-carnitine zebrafish model by feeding fish with mildronate as a specific carnitine synthesis inhibitor [0.05% body weight (BW)/d] for 7 weeks, and the systemically changed nutrient metabolism, including carnitine and triglyceride (TG) concentrations, fatty acid (FA) β-oxidation capability, and other molecular and biochemical assays of lipid, glucose, and protein metabolism, were measured. The results indicated that mildronate markedly decreased hepatic carnitine concentrations while it had no effect in muscle. Liver TG concentrations increased by more than 50% in mildronate-treated fish. Mildronate decreased the efficiency of liver mitochondrial β-oxidation, increased the hepatic mRNA expression of genes related to FA β-oxidation and lipolysis, and decreased the expression of lipogenesis genes. Mildronate decreased whole body glycogen content, increased glucose metabolism rate, and upregulated the expression of glucose uptake and glycolysis genes. Mildronate also increased whole body protein content and hepatic mRNA expression of mechanistic target of rapamycin ( mtor ), and decreased the expression of a protein catabolism-related gene. Liver, rather than muscle, was the primary organ targeted by mildronate. In short, mildronate-induced hepatic inhibited carnitine synthesis in zebrafish caused decreased mitochondrial FA β-oxidation efficiency, greater lipid accumulation, and altered glucose and protein metabolism. This reveals the key roles of mitochondrial fatty acid β-oxidation in nutrient metabolism in fish, and this low-carnitine zebrafish model could also be used as a novel fish model for future metabolism studies.

Antibiotics have been widely used in human and veterinary medicine to both treat and prevent disease. Due to their high water solubility and low bioavailability, many antibiotic residues have been found in aquatic environments. Fish are an indispensable link between the environmental pollution and human health. However, the chronic effects of environmental concentrations of antibiotics in fish have not been thoroughly investigated. Sulfamethoxazole (SMX) and oxytetracycline (OTC) are frequently detected in aquatic environments. In this study, zebrafish were exposed to SMX (260 ng/L) and OTC (420 ng/L) for a six-week period. Results indicated that exposure to antibiotics did not influence weight gain of fish but increased the metabolic rate and caused higher mortality when treated fish were challenged with Aeromonas hydrophila. Furthermore, exposure to antibiotics in water resulted in a significant decrease in intestinal goblet cell numbers, alkaline phosphatase (AKP), acid phosphatase (ACP) activities, and the anti-oxidant response while there was a significant increase in expression of inflammatory factors. Antibiotic exposure also disturbed the intestinal microbiota in the OTC-exposed group. Our results indicated that environmental antibiotic concentrations can impair the gut health of zebrafish. The potential health risk of antibiotic residues in water should be evaluated in the future., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Growing mixed-sex Nile tilapia, Oreochromis niloticus in earthen ponds to table size is a major challenge due to its early maturity and prolific breeding. This study determined the effects of two medicinal plants; Aspilia plant, Aspilia mossambicensis and Neem tree, Azadirachta indica on hatchlings production, growth performance, feed utilization, survival and haematology of O. niloticus . Experimental diets were prepared by adding 1.0, 2.0, 4.0 and 8.0 g of either A. mossambicensis or A. indica leaf powders into a kg of the control diet subsequently administered daily to twenty triplicates of O. niloticus for three months. Both A. mossambicensis and A. indica leaf powder at the used doses, reduced significantly hatchlings production of O. niloticus when compared to the control ( P  < .05) . The lowest value of hatchlings count was found in A. indica dose 8.0 g kg -1 ( P  < .05). The use of A. mossambicensis leaf powder at a dose of 4.0 g kg -1 improved significantly growth performance and feed utilization ( P  < .05). In contrast, survival rate was not affected significantly by the two plants ( P  > .05). Both plants differentially increased significantly haematological parameters such as Hb concentration, packed cell volume (PCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), white blood cells (WBC), monocyte and lymphocytes while reduced significantly neutrophils and eosinophils ( P  < .05). In conclusion, A. mossambicensis and A. indica leaf powders control prolific breeding of O. niloticus , modulate its growth performance and feed utilization. The two plants also modulate haematological parameters of O. niloticus indicating immunological response towards stress or intoxication, however, the values obtained were not beyond the recommended range for healthy fish.

Visceral adipose tissue (VAT) and subcutaneous adipose tissue (SCAT) have different structures and metabolic functions and play different roles in the regulation of the mammal endocrine system. However, little is known about morphology and physiological and metabolic functions between VAT and SCAT in fish. We compared the morphological, physiological, and biochemical characteristics of VAT and SCAT in Nile tilapia and measured their functions in energy intake flux, lipolytic ability, and gene expression patterns. SCAT contained more large adipocytes and nonadipocytes than VAT in Nile tilapia. VAT had higher lipid content and was the primary site for lipid deposition. Conversely, SCAT had higher hormone-induced lipolytic activity. Furthermore, SCAT had a higher percentage of monounsaturated and lower polyunsaturated fatty acids than VAT. SCAT had higher mitochondrial DNA, gene expression for fatty acid β-oxidation, adipogenesis, and brown adipose tissue characteristics, but it also had a lower gene expression for inflammation and adipocyte differentiation than VAT. SCAT and VAT have different morphological structures, as well as physiological and metabolic functions in fish. VAT is the preferable lipid deposition tissue, whereas SCAT exhibits higher lipid catabolic activity than VAT. The physiological functions of SCAT in fish are commonly overlooked. The present study indicates that SCAT has specific metabolic characteristics that differ from VAT. The differences between VAT and SCAT should be considered in future metabolism studies using fish as models, either in biomedical or aquaculture studies., (Copyright © 2017 the American Physiological Society.)

Energy metabolism plays important roles in stress resistance and immunity in mammals, however, such functions have not been established in fish. In the present study, Nile tilapia (Oreochromis niloticus) was fed with mildronate, an inhibitor of mitochondrial fatty acid (FA) β-oxidation, for six weeks subsequently challenged with Aeromonas hydrophila and ammonia nitrogen exposure. Mildronate treatment reduced significantly l-carnitine concentration and mitochondrial FA β-oxidation efficiency, while it increased lipid accumulation in liver. The fish with inhibited hepatic FA catabolism had lower survival rate when exposed to Aeromonas hydrophila and ammonia nitrogen. Moreover, fish fed mildronate supplemented diet had lower immune enzymes activities and anti-inflammatory cytokine genes expressions, but had higher pro-inflammatory cytokine genes expressions. However, the oxidative stress-related biochemical indexes were not significantly affected by mildronate treatment. Taken together, inhibited mitochondrial FA β-oxidation impaired stress resistance ability in Nile tilapia mainly through inhibiting immune functions and triggering inflammation. This is the first study showing the regulatory effects of lipid catabolism on stress resistance and immune functions in fish., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Suppressor of cytokine signaling (SOCS) proteins are inverse feedback regulators of cytokine and hormone signaling mediated by the Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathway that are involved in immunity, growth and development of organisms. In the present study, three SOCS genes, SOCS-1, SOCS-2 and SOCS-3, were identified in an economically important fish, Nile tilapia (Oreochromis niloticus) referred to as NtSOCS-1, NtSOCS-2 and NtSOCS-3. Multiple alignments showed that, the three SOCS molecules share highly conserved functional domains, including the SRC homology 2 (SH2) domain, the extended SH2 subdomain (ESS) and the SOCS box with others vertebrate counterparts. Phylogenetic analysis indicated that NtSOCS-1, 2 and 3 belong to the SOCS type II subfamily. Whereas NtSOCS-1 and 3 showed close evolutionary relationship with Perciformes, NtSOCS-2 was more related to Salmoniformes. Tissue specific expression results showed that, NtSOCS-1, 2 and 3 were constitutively expressed in all nine tissues examined. NtSOCS-1 and 3 were highly expressed in immune-related tissues, such as gills, foregut and head kidney. However, NtSOCS-2 was superlatively expressed in liver, brain and heart. In vivo, NtSOCS-1 and 3 mRNA levels were up-regulated after lipopolysaccharide (LPS) challenge while NtSOCS-2 was down-regulated. In vitro, LPS stimulation increased NtSOCS-3 mRNA expression, however it inhibited the transcription of NtSOCS-1 and 2. Collectively, our findings suggest that, the NtSOCS-1 and 3 might play significant role(s) in innate immune response, while NtSOCS-2 may be more involved in metabolic regulation., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Unaffordability of commercial feeds to semi-intensive Nile tilapia, Oreochromis niloticus farmers has led to reliance on supplemental feeding and fertilization for nutrition of their fish without a scientific basis. This study compared the growth, survival, condition factor and yield performance of O. niloticus fed on mixed ingredients (MI) and rice bran alone (RB) diets reared in fertilized earthen ponds in small scale farmers' ponds. The study also determined phytoplankton composition, biomass and abundance in the experimental ponds. Quadruplicate ponds were stocked with O. niloticus and African sharptooth catfish, Clarias gariepinus at a stocking ratio of 1:3. The initial mean weights of O. niloticus and C. gariepinus were 14.62 ± 0.61 and 20.34 ± 1.44 g respectively. The fish were fed on MI and RB diets with protein content of 184.74 and 126.06 g kg(-1) respectively for 270 days. The results showed that growth performance, survival rate and condition factor of O. niloticus were not affected by either feeding with MI or RB diet (p > 0.05). Moreover, there were no significant differences obtained in phytoplankton biomass and abundance in O. niloticus ponds fed on the two diets (p > 0.05). Furthermore, feeding O. niloticus either on MI or RB diet did not affect the net and gross fish yields (p > 0.05). The study revealed that the use of either MI or RB diet does not significantly affect the performance of O. niloticus cultured in semi-intensive earthen ponds where natural food organisms are an integral part of the culture system. Rice bran could be used for semi-intensive culture of O. niloticus in fertilized ponds to boost the production based on its availability and low cost.

Phytoplankton can indirectly indicate health status of coral reefs due to their sensitivity to changes in water quality parameters. This study explored the spatial and temporal variability in water quality and nutrients in relation to phytoplankton community composition and chlorophyll a concentration at Bawe, Mnemba, Chumbe and Pongwe coral reef sites in Unguja Island. In situ measurements of dissolved oxygen, temperature, salinity and pH were done every month for 1 year. Surface water samples were collected for determination of phytoplankton composition, nutrients and chlorophyll a concentration. Dissolved oxygen, temperature, salinity and pH did not differ significantly among the four sites (p > 0.05) but showed significant temporal variations among months (p < 0.05). Bawe had significantly higher phosphate concentration (1.45 ± 0.57 µg L(-1)) than Chumbe (0.74 ± 0.53 µg L(-1)), Mnemba (0.42 ± 0.30 µg L(-1)) and Pongwe (0.28 ± 0.10 µg L(-1); p < 0.05). Similarly, Bawe had significantly higher nitrate concentration (0.81 ± 0.43 µg L(-1)) than Mnemba (0.33 ± 0.14 µg L(-1)) and Pongwe (0.24 ± 0.13 µg L(-1); p < 0.05) but similar to Chumbe (0.90 ± 0.35 µg L(-1); p > 0.05). However, values obtained at all the studied sites were less than 3 and 14 mg L(-1) for phosphate and nitrate, respectively, for eutrophic oceans. Phytoplankton species were dominated by Bacillariophyceae (70.83 %) and some species identified such as Ceratium sp., Dinophysis sp., Protoperidinium sp., Prorocentrum sp., Oscillatoria sp. and Dictyocha fibula are known to produce toxins that affect fish species. Bawe had significantly higher chlorophyll a concentration (0.47 ± 0.07 mg L(-1)) than Mnemba (0.33 ± 0.04 mg L(-1)) and Chumbe (0.33 ± 0.04 mg L(-1); p < 0.05). Chlorophyll a concentration was spatially inversely related to distance from Unguja town (p < 0.05) while it was temporally significantly positively correlated with dissolved oxygen, nitrate and phosphate (p < 0.05). The study revealed that, the coral reef sites have low nutrient levels and are in good health. The existence of toxic phytoplankton species suggests careful consumption of fisheries resources at the four coral reef sites and frequent monitoring for Harmful Algal Blooms (HABs) is required. The higher nutrients and chlorophyll a concentrations at Bawe Island compared to other sites calls for mechanisms to limit the release of domestic sewage from households and hotels to safeguard the coral reefs.

This study was conducted to assess the reproductive biology and early breeding behaviour of female Oreochromis niloticus reared in monoculture and polyculture with Clarias gariepinus in earthen ponds for 8 months. Results revealed no significant difference in length at first maturity (L50) between females reared in monoculture and polyculture systems. Similarly, no significant differences were detected in absolute fecundity, relative fecundity, gonado-somatic index and condition factor between the two culture systems. The absolute fecundity was more strongly correlated with total length and body weight than with ovary weight. The study concluded that early breeding of O. niloticus in captivity is not affected by the culture systems used. Therefore, O. niloticus production in either system can be improved only through proper pond management techniques.

Fatty acids (FAs) particularly ω3 and ω6 polyunsaturated fatty acids (PUFAs) play important role in human health. This study aimed to investigate the composition and levels of selected ω3 PUFAs in four commercial fish species, Nile perch (Lates niloticus), Nile tilapia (Oreochromis niloticus), Tilapia zillii, and dagaa (Rastrineobola argentea) from Mwanza Gulf in Lake Victoria. The results indicated that 36 types of FAs with different saturation levels were detected. These FAs were dominated by docosahexaenoic (DHA), eicosapentaenoic (EPA), docosapentaenoic (DPA), and eicosatetraenoic acids. O. niloticus had the highest composition of FAs (34) compared to L. niloticus (27), T. zillii (26), and R. argentea (21). The levels of EPA differed significantly among the four commercial fish species (F = 6.19,  P = 0.001). The highest EPA levels were found in R. argentea followed by L. niloticus and O. niloticus and the lowest in T. zillii. The DPA levels showed no significant difference among the four fish species studied (F = 0.652,  P = 0.583). The study concluded that all four commercial species collected from Mwanza Gulf are good for human health, but R. argentea is the best for consumption because it contains higher levels of ω3 FAs, mainly EPA.