Profiling Phospholipids within Atlantic Salmon Salmo salar with Regards to a Novel Terrestrial Omega-3 Oil Source

The development and inclusion of novel oils derived from genetically modified (GM) oilseeds into aquafeeds, to supplement and supplant current terrestrial oilseeds, as well as fish oils, warrants a more thorough investigation into lipid biochemical alterations within finfish species, such as Atlantic salmon. Five tissues were examined across two harvesting timepoints to establish whether lipid isomeric alterations could be detected between a standard commercial diet versus a diet that incorporated the long-chain polyunsaturated fatty acids (LC-PUFA), EPA (eicosapentaenoic acid), and DHA (docosahexaenoic acid), derived from the GM oilseed Camelina sativa. Tissue-dependent trends were detected, indicating that certain organs, such as the brain, have a basal limit to LC-PUFA incorporation, though enrichment of these fatty acids is possible. Lipid acyl alterations, as well as putative stereospecific numbering (sn) isomer alterations, were also detected, providing evidence that GM oils may modify lipid structure, with lipids of interest providing a set of targeted markers by which lipid alterations can be monitored across various novel diets

A detailed NMR study of the solution stereodynamics in tricarbonylrhenium(I) halide complexes of the non-racemic chiral ligand 2,6-bis[(4R,5R)-4,5-dimethyl-1,3-dioxolan-2-yl]pyridine (L¹) and the molecular structure of fac-[ReBr(CO)₃(L¹)]

1 Tricarbonylrhenium(I) halide complexes of the non-racemic chiral ligand 2,6-bis[(4R, 5R)-dimethyl-1,3-dioxan-2-yl]pyridine (L¹), namely fac-[ReX(CO)₃(L¹)] (X = Cl, Br or I), have been prepared. In these complexes the ligand is bound in a bidentate fashion, with the N atom of the pyridine ring and an O atom of one of the acetal rings co-ordinated to the octahedral metal centre. The bidentate mode is confirmed by the X-ray structure of fac-[ReBr(CO)₃(L¹)]. There are four possible diastereoisomers, depending on the configuration at the metal centre and at the acetal-carbon atom of the co-ordinated ring; the X-ray structure of fac-[ReBr(CO)₃(L¹)] shows that the SR diastereoisomer is present in the solid state. In solution, three of the four possible diastereoisomers are observed, namely SR, RR and RS; their relative populations are in the order SR > RR > SS. Above ambient temperature the complexes are stereochemically non-rigid. The fluxional kinetics have been measured by a combination of standard band shape analysis and selective inversion experiments. Two distinct processes are present: an acetal ring flip and exchange of the pendant and co-ordinated acetal rings. The latter process occurs via two independent mechanisms, namely tick-tock and rotation pathways. The activation energies for the stereodynamics are in the ranges 72 – 73 kJ mol⁻¹ (tick-tock), 77 – 78 kJ mol⁻¹ (acetal ring flip) and 83 – 90 kJ mol⁻¹ (rotation) at 298 K

Effect of salinity on the biosynthesis of n-3 long-chain polyunsaturated fatty acids in silverside Chirostoma estor

The genus Chirostoma (silversides) belongs to the family Atherinopsidae, which contains around 150 species, most of which are marine. However, Mexican silverside (Chirostoma estor) is one of the few representatives of freshwater atherinopsids and is only found in some lakes of the Mexican Central Plateau. However, studies have shown that C. estor has improved survival, growth and development when cultured in water conditions with increased salinity. In addition, C. estor displays an unusual fatty acid composition for a freshwater fish with high docosahexaenoic acid (DHA) : eicosapentaenoic acid (EPA) ratios. Freshwater and marine fish species display very different essential fatty acid metabolism and requirements and so the present study investigated long-chain polyunsaturated fatty acid (LC-PUFA) biosynthesis to determine the capacity of C. estor for endogenous production of EPA and DHA, and the effect that salinity has on these pathways. Briefly, C. estor were maintained at three salinities (0, 5 and 15 ppt) and the metabolism of 14C-labelled 18:3n-3 determined in isolated hepatocyte and enterocyte cells. The results showed that C. estor has the capacity for endogenous biosynthesis of LC-PUFA from 18-carbon fatty acid precursors, but that the pathway was essentially only active in saline conditions with virtually no activity in cells isolated from fish grown in freshwater. The activity of the LCPUFA biosynthesis pathway was also higher in cells isolated from fish at 15 ppt compared to fish at 5 ppt, The pathway was around 5-fold higher in hepatocytes compared to enterocytes, although the majority of 18:3n-3 was converted to 18:4n-3 and 20:4n-3 in hepatocytes whereas the proportions of 18:3n-3 converted to EPA and DHA were higher in enterocytes. The data were consistent with the hypothesis that conversion of EPA to DHA could contribute, at least in part, to the generally high DHA:EPA ratios observed in the tissue lipids of C. estor

Investigation of highly unsaturated fatty acid metabolism in the Asian sea bass, Lates calcarifer

Lates calcarifer, commonly known as the Asian sea bass or barramundi, is an interesting species that has great aquaculture potential in Asia including Malaysia and also Australia. We have investigated essential fatty acid metabolism in this species, focusing on the endogenous highly unsaturated fatty acid (HUFA) synthesis pathway using both biochemical and molecular biological approaches. Fatty acyl desaturase (Fad) and elongase (Elovl) cDNAs were cloned and functional characterization identified them as ∆6 Fad and Elovl5 elongase enzymes, respectively. The ∆6 Fad was equally active towards 18:3n-3 and 18:2n-6, and Elovl5 exhibited elongation activity for C18-20 and C20-22 elongation and a trace of C22-24 activity. The tissue profile of gene expression for ∆6 fad and elovl5 genes, showed brain to have the highest expression of both genes compared to all other tissues. The results of tissue fatty acid analysis showed that the brain contained more docosahexaenoic acid (DHA, 22:6n-3) than flesh, liver and intestine. The HUFA synthesis activity in isolated hepatocytes and enterocytes using [1-14C]18:3n-3 as substrate was very low with the only desaturated product detected being 18:4n-3. These findings indicate that L. calcarifer display an essential fatty acid pattern similar to other marine fish in that they appear unable to synthesize HUFA from C18 substrates. High expression of ∆6 fad and elovl5 genes in brain may indicate a role for these enzymes in maintaining high DHA levels in neural tissues through conversion of 20:5n-3

Egg quality determinants in cod (Gadus morhua L.): egg performance and lipids in eggs from farmed and wild broodstock

Lipids and essential fatty acids, particularly the highly unsaturated fatty acids, 20:5n-3 (eicosapentaenoic acid; EPA), 22:6n-3 (docosahexaenoic acid; DHA) and 20:4n-6 (arachidonic acid, AA) have been shown to be crucial determinants of marine fish reproduction directly affecting fecundity, egg quality, hatching success, larval malformation and pigmentation. In Atlantic cod (Gadus morhua L.) culture, eggs from farmed broodstock can have much lower fertilisation and hatching rates than eggs from wild broodstock. The present study aimed to test the hypothesis that potential quality and performance differences between eggs from different cod broodstock would be reflected in differences in lipid and fatty acid composition. Thus eggs were obtained from three broodstock, farmed, wild/fed and wild/unfed, and lipid content, lipid class composition, fatty acid composition and pigment content were determined and related to performance parameters including fertilisation rate, symmetry of cell division and survival to hatching. Eggs from farmed broodstock showed significantly lower fertilisation rates, cell symmetry and survival to hatching rates than eggs from wild broodstock. There were no differences in total lipid content or the proportions of the major lipid classes between eggs from the different broodstock. However, eggs from farmed broodstock were characterised by having significantly lower levels of some quantitatively minor phospholipid classes, particularly phosphatidylinositol. There were no differences between eggs from farmed and wild broodstock in the proportions of saturated, monounsaturated and total polyunsaturated fatty acids. The DHA content was also similar. However, eggs from farmed broodstock had significantly lower levels of AA, and consequently significantly higher EPA/AA ratios than eggs from wild broodstock. Total pigment and astaxanthin levels were significantly higher in eggs from wild broodstock. Therefore, the levels of AA and phosphatidylinositol, the predominant AA-containing lipid class, and egg pigment content were positively related to egg quality or performance parameters such as fertilisation and hatching success rates, and cell symmetry

Access to interpreting services in England: secondary analysis of national data

Background: Overcoming language barriers to health care is a global challenge. There is great linguistic diversity in the major cities in the UK with more than 300 languages, excluding dialects, spoken by children in London alone. However, there is dearth of data on the number of non-English speakers for planning effective interpreting services. The aim was to estimate the number of people requiring language support amongst the minority ethnic communities in England. Methods: Secondary analysis of national representative sample of subjects recruited to the Health Surveys for England 1999 and 2004. Results: 298,432 individuals from the four main minority ethnic communities (Indian, Pakistani, Bangladeshi and Chinese) who may be unable to communicate effectively with a health professional. This represents 2,520,885 general practice consultations per year where interpreting services might be required. Conclusion: Effective interpreting services are required to improve access and health outcomes of non-English speakers and thereby facilitate a reduction in health inequalities

The efficiency of indicator groups for the conservation of amphibians in the Brazilian Atlantic Forest

The adequate selection of indicator groups of biodiversity is an important aspect of the systematic conservation planning. However, these assessments differ in the spatial scale, in the methods used and in the groups considered to accomplish this task, which generally produces contradictory results. The quantification of the spatial congruence between species richness and complementarity among different taxonomic groups is a fundamental step to identify potential indicator groups. Using a constructive approach, the main purposes of this study were to evaluate the performance and efficiency of eight potential indicator groups representing amphibian diversity in the Brazilian Atlantic Forest. Data on the geographic range of amphibian species that occur in the Brazilian Atlantic Forest was overlapped to the full geographic extent of the biome, which was divided into a regular equal-area grid. Optimization routines based on the concept of complementarily were applied to verify the performance of each indicator group selected in relation to the representativeness of the amphibians in the Brazilian Atlantic Forest as a whole, which were solved by the algorithm"simulated annealing", through the use of the software MARXAN. Some indicator groups were substantially more effective than others in regards to the representation of the taxonomic groups assessed, which was confirmed by the high significance of data (F = 312.76; p < 0.01). Leiuperidae was considered as the best indicator group among the families analyzed, as it showed a good performance, representing 71% of amphibian species in the Brazilian Atlantic Forest (i.e. 290 species), which may be associated with the diffuse geographic distribution of its species. This study promotes understanding of how the diversity standards of amphibians can be informative for systematic conservation planning on a regional scale

Highly unsaturated fatty acid synthesis in marine fish: Cloning, functional characterization, and nutritional regulation of fatty acyl delta6 desaturase of Atlantic cod (Gadus morhua L.)

Fish contain high levels of the n-3 highly unsaturated fatty acids (HUFA), eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids that are crucial to the health of higher vertebrates. Biosynthesis of HUFA requires enzyme-mediated desaturation of fatty acids. Here we report cloning and functional characterisation of a ∆6 fatty acyl desaturase of Atlantic cod (Gadus morhua), and describe its tissue expression and nutritional regulation. PCR primers were designed based on the sequences of conserved motifs in available fish desaturases and used to isolate a cDNA fragment from liver of cod. The full-length cDNA was obtained by Rapid Amplification of cDNA Ends (RACE). The cDNA for the putative fatty acyl desaturase was shown to comprise 1980bp which included a 5’-UTR of 261bp and a 3’-UTR of 375bp. Sequencing revealed that the cDNA included an ORF of 1344 bp that specified a protein of 447 amino acids. The protein sequence included three histidine boxes, two transmembrane regions, and an N-terminal cytochrome b5 domain containing the haem-binding motif HPGG, all of which are characteristic of microsomal fatty acid desaturases. The cDNA displayed Δ6 desaturase activity in a heterologous yeast expression system. Quantitative real time PCR assay of gene expression in cod showed that the ∆6 desaturase gene, was highly expressed in brain, relatively highly expressed in liver, kidney, intestine, red muscle and gill, and expressed at much lower levels in white muscle, spleen and heart. In contrast, the abundance of a cod fatty acyl elongase transcript was high in brain and gill, with intermediate levels in kidney, spleen, intestine and heart, and relatively low expression in liver. The expression of the Δ6 desaturase gene and the PUFA elongase gene may be under a degree of nutritional regulation, with levels being marginally increased in livers and intestine of fish fed a vegetable oil blend by comparison with levels in fish fed fish oil. However, this was not reflected in increased Δ6 desaturase activity in hepatocytes or enterocytes, which showed very little highly unsaturated fatty acid biosynthesis activity irrespective of diet. The study described has demonstrated that Atlantic cod express a fatty acid desaturase gene with functional Δ6 activity in a yeast expression system. This is consistent with an established hypothesis that the poor ability of marine fish to synthesise HUFA is not due to lack of a Δ6 desaturase, but rather to deficiencies in other parts of the biosynthetic pathway. However, further studies are required to determine why the Δ6 desaturase appears to be barely functional in cod under the conditions tested

Functional Desaturase Fads1 (Δ5) and Fads2 (Δ6) Orthologues Evolved before the Origin of Jawed Vertebrates

Long-chain polyunsaturated fatty acids (LC-PUFAs) such as arachidonic (ARA), eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids are essential components of biomembranes, particularly in neural tissues. Endogenous synthesis of ARA, EPA and DHA occurs from precursor dietary essential fatty acids such as linoleic and α-linolenic acid through elongation and Δ5 and Δ6 desaturations. With respect to desaturation activities some noteworthy differences have been noted in vertebrate classes. In mammals, the Δ5 activity is allocated to the Fads1 gene, while Fads2 is a Δ6 desaturase. In contrast, teleosts show distinct combinations of desaturase activities (e.g. bifunctional or separate Δ5 and Δ6 desaturases) apparently allocated to Fads2-type genes. To determine the timing of Fads1-Δ5 and Fads2-Δ6 evolution in vertebrates we used a combination of comparative and functional genomics with the analysis of key phylogenetic species. Our data show that Fads1 and Fads2 genes with Δ5 and Δ6 activities respectively, evolved before gnathostome radiation, since the catshark Scyliorhinus canicula has functional orthologues of both gene families. Consequently, the loss of Fads1 in teleosts is a secondary episode, while the existence of Δ5 activities in the same group most likely occurred through independent mutations into Fads2 type genes. Unexpectedly, we also establish that events of Fads1 gene expansion have taken place in birds and reptiles. Finally, a fourth Fads gene (Fads4) was found with an exclusive occurrence in mammalian genomes. Our findings enlighten the history of a crucially important gene family in vertebrate fatty acid metabolism and physiology and provide an explanation of how observed lineage-specific gene duplications, losses and diversifications might be linked to habitat-specific food web structures in different environments and over geological timescales

Interactions between dietary docosahexaenoic acid and other long-chain polyunsaturated fatty acids on performance and fatty acid retention in post-smolt Atlantic salmon (Salmo salar)

A study with varying dietary inclusion levels (1, 5, 10, 15 and 20 g kg-1) of docosahexaenoic acid (DHA; 22:6n-3) was conducted with post-smolt (111 &plusmn; 2.6 g; mean &plusmn; S.) Atlantic salmon (Salmo salar) over a 9-week period. In addition to the series of DHA inclusion levels, the study included further diets that had DHA at 10 g kg-1 in combination with either eicosapentaenoic acid (EPA; 20:5n-3) or arachidonic acid (ARA; 20:4n-6), both also included at 10 g kg-1. An additional treatment with both EPA and DHA included at 5 g kg-1 (total of 10 g kg-1 long-chain polyunsaturated fatty acids, LC-PUFA) was also included. After a 9-week feeding period, fish were weighed, and carcass, blood and tissue samples collected. A minor improvement in growth was seen with increasing inclusion of DHA. However, the addition of EPA further improved growth response while addition of ARA had no effect on growth. As with most lipid studies, the fatty acid composition of the whole body lipids generally reflected that of the diets. However, there were notable exceptions to this, and these implicate some interactions among the different LC-PUFA in terms of the fatty acid bio- chemistry in this species. At very low inclusion levels, DHA retention was substantially higher (*250 %) than that at all other inclusion levels (31&ndash;58 %). The inclusion of EPA in the diet also had a positive effect on the retention efficiency of DHA. However, EPA retention was highly variable and at low DHA inclusion levels there was a net loss of EPA as this fatty acid was most likely elongated to produce DHA, consistent with increased DHA retention with addi- tional EPA in the diet. Retention of DPA (22:5n-3) was high at low levels of DHA, but diminished with increasing DHA inclusion, similar to that seen with DHA retention. The addition of EPA to the diet resulted in a substantial increase in the efficiency of DPA retention; the inclusion of ARA had the opposite effect. Retention of ARA was unaffected by DHA inclusion, but the addition of either EPA or ARA to the diet resulted in a substantial reduction in the efficiency of ARA retention. No effects of dietary treatment were noted on the retention of either linolenic (18:3n-3) or linoleic (18:2n-6) acids. When the total n-3 LC-PUFA content of the diet was the same but consisted of either DHA alone or as a combination of EPA plus DHA, the performance effects were similar