Tissue distribution of lipid and fatty acid metabolism and transcription factors genes in adult Atlantic bluefin tuna (Thunnus thynnus L.)

To determine expression of the major lipid pathways in tissues of adult bluefin tuna 8 individuals were used for collecting samples for tissue expression of key lipid metabolism genes. Triplicate sets of samples of brain, gills, heart, kidney, spleen, liver, intestine, white muscle, red muscle, adipose tissue ovary and testis were collected. Expression of genes was determined by qPCR. Tissue expression profiles showed that PUFA biosynthetic pathway genes were expressed in all tissues examined, highest expression in brain, liver and testis. Elongase elovl5 showed higher expression than desaturase fads2d6 in all other tissues, with low expression of in red muscle and ovaries. Transcription factors, pparα and pparγ showed parallel expression, with adipose tissue with the highest relative copy number, followed by intestine>testis>liver. The expression of lxr was low in liver, with highest expression in testis, brain and kidney. Similarly, rxr was poorly expressed in liver with higher expression in muscle, spleen and brain. The rank order of expression of srebp1 was brain, testis, ovary, intestines, kidney, gill, liver, white muscle, spleen, heart and red muscle. For srebp2 the highest expression was shown in brain, testis and adipose with lowest expression in heart and white muscle. Expression of fabp2 was highest in intestine, brain and heart with lower levels in liver, red muscle, adipose and kidney. fabp4 showed highest expression in ovaries with liver showing the lowest. fabp7 showed highest expression levels in brain and testis with lowest values in liver. cptI expression was highest in brain and lowest in liver. Similarly, the relative copy number for fas was highest in brain followed by gonads, gill and liver, with white muscle showing lowest expression. The expression of aco was highest in adipose and intestine, followed by liver, kidney and brain. The expression of hmgcl was highest in ovary followed by adipose, brain and testis, with lowest expression in liver. Expression of lpl was highest in testis and lowest in ovary, with liver and white muscle, adipose, heart, gills and red muscle, kidney, intestine showing intermediate levels of expression. This research was supported by projects from the Junta de Andalucía, Proyecto de Excelencia de Promoción General del Conocimiento Ref. RNM 0733, and Programa Estatal de Investigación del Ministerio de Economía y Competitividad Ref. AGL2014-52003-C2-1-R. AQUACULTURE, ENDOCRINOLOGY AND TOXICOLOGY 218. VI IBERIAN CONGRESS OF ICHTHYOLOGY MURCIA, SPAIN 21st · 24th June P-04

Molecular aspects of lipid metabolism, digestibility and antioxidant status of Atlantic bluefin tuna (T. thynnus L.) larvae during first feeding

Atlantic bluefin tuna (Thunnus thynnus L.; ABT) larvae were fed on enriched rotifers Brachionus rotundiformis and copepod nauplii Acartia tonsa from first feeding to 15 days post hatching. Rotifers were enriched with five different commercial products: OG, MG, AG and RA plus selenium and vitamin E. Copepods (COP) were cultured with the algae Rhodomonas baltica. Metabolic processes were studied by determining the expression of 30 genes related to lipid metabolism (transcription factors, fatty acid metabolism and lipid homeostasis), antioxidant enzymes, myogenesis and digestive enzymes. Growth and development parameters and high expression of myogenesis genes myhc2 and tropo indicated that COP were better than enriched rotifers as live prey for first feeding ABT. COP and AG-fed larvae showed the lowest values for the transcription factors pparã and srebp2. The expression of fas showed differences among treatments, with highest relative expression in COP-fed larvae and those fed with RA rotifers. In relation to fatty acid catabolism, larvae fed RA had the highest aco expression levels, with the lowest observed in those fed COP. The expression profiles of lipid homeostasis genes showed that larvae fed COP had higher fabp2 and 4 expressions. Larvae fed AG showed the lowest lpl expression levels, with highest values observed in larvae fed OG. Regarding antioxidant enzyme gene expression, sod showed highest values in larvae fed COP and RA, with larvae fed MG rotifers showing lowest expression levels. A similar pattern was observed for the expression of cat and gpx1 and 4. The expression of genes for digestive enzymes showed that tryp expression levels were highest in COP-fed larvae but, in contrast, COP-fed larvae showed the lowest anpep and alp levels. ABT larvae fed AG displayed the lowest expression level of pla2. bal1 and bal2 presented similar expression patterns, with highest values in COP-fed ABT and lowest expression in larvae fed AG rotifers. Copepods were a superior live prey for first feeding ABT larvae compared to enriched rotifers, as indicated by the higher growth and flexion index achieved by COP-fed larvae, possibly reflecting the higher protein content of the copepods.This work was supported by the Consejería de Innovación, Ciencia y Empresa de la Junta de Andalucía, Proyecto de Excelencia de Promoción General del Conocimiento [Ref. RNM 733, 2012], and Programa Estatal de Investigación del Ministerio de Economía y Competitividad [Ref. AGL2014-52003-C2-1-R, 2014].Versión del edito

Development of feeds for juvenile Atlantic bluefin tuna (Thunnus thynnus, L): effect of lipid level and source

In conclusion, the present study suggests that ABT juveniles can be grown on inert extruded dry feeds that result in good fish growth and accumulation of the health-promoting fatty acid DHA. Furthermore, a blend of VO and KO could be used as the dietary lipid source up to a dietary lipid level of 15 % without affecting fish performance. The expression of lipid metabolism genes in ABT liver showed a different response to dietary lipid level/fatty acid profile, consistent with previous data indicating limited n-3 LC-PUFA biosynthetic capability in ABT. However, gene expression showed some differences between the two trials, which highlight how the genetic background of different batches of ABT juveniles could affect the regulation of metabolic gene expression and thus be a factor in weaning success. The expression of antioxidant enzymes was also altered by diet, related to dietary contents of antioxidant nutrients. Thus, further studies are required in order to fully elucidate the lipid and fatty acid requirements of this iconic species in relation to dietary sources and production cost

Taurine metabolism and effects of inclusion levels in rotifer (Brachionus rotundiformis, Tschugunoff, 1921) on Atlantic bluefin tuna (Thunnus thynnus, L.) larvae

Taurine appears to be a crucial nutrient for teleosts, especially top predator species such as Atlantic bluefin tuna (Thunnus thynnus, L.; ABT). While dietary taurine supplementation has been highly recommended, there is a lack of studies on taurine assimilation and biosynthesis for this iconic species. The present study aims to provide insight into the molecular mechanisms involved in taurine biosynthesis and transport in ABT by studying tissue distribution and ontogenetic development of expression of cysteine dioxygenase (cdo), cysteine sulfinic acid decarboxylase (csad), 2-aminoethanethiol dioxygenase (ado) and taurine transporter (tauT) in response to graded levels of dietary taurine supplementation. The full open reading frame (ORF) for cdo and partial sequences for csad, ado and tauT were obtained, with the translated polypeptides being 202, 176, 166 and 324 amino acids, respectively. All three showed characteristics such as cupin motifs in Cdo and predicted N-glycosylation sites in Taut that are common to these genes in other species. Phylogenetic analysis showed that the ABT sequences clustered with sequences of other teleosts, and separately from mammals and molluscs. Tissue distribution varied, with adipose tissue, kidney, white muscle and testis/brain showing highest expression of cdo, csad, ado and tauT, respectively. Whole larvae expression of csad peaked at 15 dah, whereas the other genes generally increased throughout development to show highest expression at 25 dah. The nutritional trial was carried out by feeding ABT larvae from mouth opening to 14 days after hatching (dah) with rotifers (Brachionus rotundiformis) enriched with 4 different levels of taurine: 0.0 (tau0), 0.5 (tau0.5), 1.0 (tau1), and 2.0 g taurine per 106 rotifers (tau2). Rotifers effectively accumulated taurine with ABT larvae fed on treatment tau2 attaining the highest concentration of taurine. However, ABT larvae fed tau1 displayed higher growth and survival, and flexion index at 14 dah, than larvae fed the other taurine levels. Larvae fed tau1 also showed generally higher expression of tauT and cdo and digestive and antioxidant enzyme genes. While this study showed that larval ABT express taurine metabolism genes, suggesting possible synthesis that could contribute to the taurine pool in the fish, larval performance was enhanced by a level of dietary taurine (3.7 mg taurine g−1 rotifer) supplied by enrichment of rotifers at 1 g taurine per 106 rotifers.En prensa2,04

COPEPODS OR ROTIFERS? EVALUATING THE USE OF DIFFERENT FEEDING PROTOCOLS FOR LARVAE OF ATLANTIC BLUEFIN TUNA (Thunnus thynnus. L)

There are still many issues that require to be solved in larval rearing of Atlantic bluefin tuna (Thunnus thynnus; ABT) to prevent “mass-mortality” during this developmental stage. Initial data related to the feeding sequence of ABT larvae suggested that mortality observed during the first stages of life could be due partly to nutritional deficiencies. Previous studies demonstrated that copepods appeared to be a superior live prey compared to rotifers during the first two weeks of life. Our overarching aim was to evaluate different feeding strategies during first feeding of ABT larvae from a performance, compositional and molecular perspective. In order to do so, two groups of ABT larvae were fed with either copepod (Acartia tonsa; C) nauplii or rotifers (Brachionus rotundiformis; R) enriched with Algamac 3050® from mouth opening to 13 days after hatching (dah). After this, the group C-larvae was fed either Artemia enriched with Algamac 3050® (CA), Acartia nauplii and copepodites (CC) or sea bream (Sparus aurata) yolk-sac larvae (CY), while the R group passed on to being fed on Artermia enriched with Algamac 3050® (RA) up to 18 dah. After 13 dah, larvae fed C grew more than those fed R although there were no differences in survival. ABT larvae fed R accumulated highest eicosapentaenoate (EPA) but lowest docosahexaenoate (DHA) and total n-3 long-chain polyunsaturated fatty acids (LC-PUFA) than C-fed larvae, reflecting dietary contents. Indeed, there was no activation in the expression of the enzymes involved in LC-PUFA biosynthesis. However, the different live prey elicited regulation of transcription factor, digestive enzyme, lipid metabolism and oxidative stress genes. At 18 dah larvae fed CY and CA were the largest size with larvae fed RA displaying the lowest growth with no differences in survival among the dietary treatments. The highest DHA contents were found in ABT larvae fed CC and CY, whereas the lowest contents were found in RA-fed larvae. Indeed, RA-fed larvae showed the highest level of the intermediate product n-3 docosapentaenoate, which could be reflecting up-regulation in the biosynthetic pathway although this was not supported by gene expression data

Performance, feed utilization, and hepatic metabolic response of weaned juvenile Atlantic bluefin tuna (Thunnus thynnus L.): effects of dietary lipid level and source

The development of formulated diets and feeds is essential to increase production of farmed tuna species. There is limited knowledge of this topic, mainly on Pacific Bluefin tuna (Thunnus orientalis) in Japan, whereas no major attempts have been made with Atlantic Bluefin tuna (Thunnus thynnus; ABT). In the present study, two trials were performed using inert formulated diets as on-growing feeds for weaned ABT juvenile in order to establish adequate dietary levels of both lipid and omega-3 long-chain polyunsaturated fatty acids (LC-PUFA). In a first trial, ABT (initial weight = 2.9±0.9g) were fed for 10 days with either a commercial (Magokoro®, MGK) or two experimental feeds with two different lipid levels (15 or 20%) using krill oil (KO) as the single lipid source in order to estimate the suitable lipid content. Fish fed MGK displayed the highest growth, followed by 15KO, with no differences in fish survival. Thus, a lipid content of 15% was considered better than 20% for ABT juveniles. In the second trial, fish (initial weight = 3.3 ± 0.6g) were fed either MGK, 15KO or a feed containing 15% lipid with a combination (1:1, v/v) KO and rapeseed oil (RO) (15KORO). Fish fed 15KO and 15KORO showed the highest growth in terms of weight and fork length (including weight gain and SGR). Increasing dietary lipid level or adding RO to the feeds did not increase liver lipid content. The liver fatty acid profile largely reflected dietary intake confirming very limited LC-PUFA biosynthetic activity for this teleost species. In this respect, liver of fish fed 15KO and 20KO displayed the highest contents of docosahexaenoic acid (DHA). The hepatic expression of genes of lipid and fatty acid metabolism, transcription factors, and antioxidant enzymes was investigated with many of the genes showing regulation by both dietary lipid and LC-PUFA contents. The present study showed promising results that suggested ABT juveniles can be on grown on inert dry feeds that supported good fish growth and the accumulation of the health-promoting fatty acid DHA. Further studies are required in order to fully elucidate lipid and fatty acid requirements of this iconic species regarding dietary sources and production costs.En prensa1,52

Performance, feed utilization and hepatic molecular metabolic response of weaned juvenile Atlantic bluefin tuna (Thunnus thynnus, L): effect of lipid level and source

The development of formulated diets and feeds is essential to increase production of farmed tuna species. There is limited knowledge of this topic, mainly on Pacific Bluefin tuna (Thunnus orientalis) in Japan, whereas no major attempts have been made with Atlantic Bluefin tuna (Thunnus thynnus; ABT). In the present study, two trials were performed using inert formulated diets as on-growing feeds for weaned ABT juvenile in order to establish adequate dietary levels of both lipid and omega-3 long-chain polyunsaturated fatty acids (LC-PUFA). In a first trial, ABT (initial weight = 2.9±0.9g) were fed for 10 days with either a commercial (Magokoro®, MGK) or two experimental feeds with two different lipid levels (15 or 20%) using krill oil (KO) as the single lipid source in order to estimate the suitable lipid content. Fish fed MGK displayed the highest growth, followed by 15KO, with no differences in fish survival. Thus, a lipid content of 15% was considered better than 20% for ABT juveniles. In the second trial, fish (initial weight = 3.3 ± 0.6g) were fed either MGK, 15KO or a feed containing 15% lipid with a combination (1:1, v/v) KO and rapeseed oil (RO) (15KORO). Fish fed 15KO and 15KORO showed the highest growth in terms of weight and fork length (including weight gain and SGR). Increasing dietary lipid level or adding RO to the feeds did not increase liver lipid content. The liver fatty acid profile largely reflected dietary intake confirming very limited LC-PUFA biosynthetic activity for this teleost species. In this respect, liver of fish fed 15KO and 20KO displayed the highest contents of docosahexaenoic acid (DHA). The hepatic expression of genes of lipid and fatty acid metabolism, transcription factors, and antioxidant enzymes was investigated with many of the genes showing regulation by both dietary lipid and LC-PUFA contents. The present study showed promising results that suggested ABT juveniles can be on grown on inert dry feeds that supported good fish growth and the accumulation of the health-promoting fatty acid DHA. Further studies are required in order to fully elucidate lipid and fatty acid requirements of this iconic species regarding dietary sources and production costs

High Speed Photometry of SDSS J013701.06-091234.9

We present high speed photometry of the Sloan Digital Sky Survey cataclysmic variable SDSS J013701.06-091234.9 in quiescence and during its 2003 December superoutburst. The orbital modulation at 79.71\pm0.01 min is double humped; the superhump period is 81.702\pm0.007 min. Towards the end of the outburst late superhumps with a period of 81.29\pm0.01 min were observed. We argue that this is a system of very low mass transfer rate, and that it probably has a long outburst interval.Comment: 5 pages, 8 figures. Accepted for publication in MNRA

ATLANTIC BLUEFIN TUNA (Thunnus thynnus, L.) LARVAE ANTIOXIDANT MOLECULAR FUNCTIONS INDUCED BY DIETARY SELENIUM IN ROTIFER Brachionus rotundiformis

including iodothyronine deiodinases 1, 2 and 3 (dio1, dio2, dio3) was unaffected. Feeding Se enriched rotifers effectively increased Se in ABT larvae tissue. The improved growth observed in Se supplemented treatments might be related to an accelerated development as the flexion index was significantly higher in all Se enriched treatments compared to the non-supplemented control. A similar effect by Se supplementation has been previously described in Senegalese sole (Solea senegalensis) in relation to an enhanced thyroid hormone activity by Se supplementation (Ribeiro et al., 2012). The Se level of 0.10 µg g-1 dw measured in non-supplemented rotifers is below known requirements in fish (Antony Jesu Prabhu et al. 2016). In contrast, rotifers supplemented with the lowest Se level (Se3) contained 4.42 µg Se g-1, which might be sufficient to cover requirements for this mineral as selenoproteins displayed maximum expression in ABT larvae fed this treatment. The increased seleno-enzyme production might have contributed towards an improved antioxidant status in ABT larvae, indicated by a transcriptional downregulation of redox sensitive antioxidant enzymes cat and sod. In conclusion, rotifers without Se enrichment are suboptimal for ABT larvae at first feeding. A dietary Se level of 4.42 µg g-1 dw is recommended as it boosted growth performance and improved the antioxidant status in ABT larvae