Effect of dietary conjugated linoleic acid (CLA) on lipid composition, metabolism and gene expression in Atlantic salmon (Salmo salar) tissues

Dietary conjugated linoleic acid (CLA) affects fat deposition and lipid metabolism in mammals, including livestock. To determine CLA effects in Atlantic salmon (Salmo salar), a major farmed fish species, fish were fed for 12 weeks on diets containing fish oil or fish oil with 2% and 4% CLA supplementation. Fatty acid composition of the tissues showed deposition of CLA with accumulation being 2 to 3 fold higher in muscle than in liver. CLA had no effect on feed conversion efficiency or growth of the fish but there was a decreased lipid content and increased protein content after 4% CLA feeding. Thus, the protein:lipid ratio in whole fish was increased in fish fed 4% CLA and triacylglycerol in liver was decreased. Liver beta-oxidation was increased whilst both red muscle beta-oxidation capacity and CPT1 activity was decreased by dietary CLA. Liver highly unsaturated fatty acid (HUFA) biosynthetic capacity was increased and the relative proportion of liver HUFA was marginally increased in salmon fed CLA. CLA had no effect on fatty acid Delta6 desaturase mRNA expression, but fatty acid elongase mRNA was increased in liver and intestine. In addition, the relative compositions of unsaturated and monounsaturated fatty acids changed after CLA feeding. CLA had no effect on PPARalpha or PPARgamma expression in liver or intestine, although PPARbeta2A expression was reduced in liver at 4% CLA feeding. CLA did not affect hepatic malic enzyme activity. Thus, overall, the effect of dietary CLA was to increase beta-oxidation in liver, to reduce levels of total body lipid and liver triacylglycerol, and to affect liver fatty acid composition, with increased elongase expression and HUFA biosynthetic capacity.     

Diversification of substrate specificities in teleostei Fads2: characterization of Δ4 and Δ6Δ5 desaturases of Chirostoma estor

Currently existing data show that the capability for long-chain PUFA (LC-PUFA) biosynthesis in teleost fish is more diverse than in other vertebrates. Such diversity has been primarily linked to the subfunctionalization that teleostei fatty acyl desaturase (Fads)2 desaturases have undergone during evolution. We previously showed that Chirostoma estor, one of the few representatives of freshwater atherinopsids, had the ability for LC-PUFA biosynthesis from C₁₈ PUFA precursors, in agreement with this species having unusually high contents of DHA. The particular ancestry and pattern of LC-PUFA biosynthesis activity of C. estor make this species an excellent model for study to gain further insight into LC-PUFA biosynthetic abilities among teleosts. The present study aimed to characterize cDNA sequences encoding fatty acyl elongases and desaturases, key genes involved in the LC-PUFA biosynthesis. Results show that C. estor expresses an elongase of very long-chain FA (Elovl)5 elongase and two Fads2 desaturases displaying Δ4 and Δ6/Δ5 specificities, thus allowing us to conclude that these three genes cover all the enzymatic abilities required for LC-PUFA biosynthesis from C₁₈ PUFA. In addition, the specificities of the C. estor Fads2 enabled us to propose potential evolutionary patterns and mechanisms for subfunctionalization of Fads2 among fish lineages.     

Ruthenium hydride and vinyl complexes supported by nitrogen-oxygen mixed-donor ligands

The Schiff base, 2-chlorophenylsalicylaldimine (HL₁), is formed readily from salicylaldehyde and 2-chloroaniline. After deprotonation, this ligand is found to react as a bidentate mixed-donor chelate with the complexes [RuRCl(CO)(BTD)(PPh₃)₂] (R = H, CHCHC₆H₅, CHCHC₆H₄Me-4, CHCH^tBu, CCCPhCHPh; BTD = 2,1,3-benzothiadiazole) to form the compounds [RuR(L₁)(CO)(PPh₃)₂] through displacement of the chloride and BTD ligands. An analogous reaction occurs with the osmium complex [OsHCl(CO)(BTD)(PPh₃)₂] to provide [OsH(L₁)(CO)(PPh₃)₂]. The compound [Ru(CHCHC₆H₄Me-4)(L₂)(CO)(PPh₃)₂] is formed through reaction of salicylaldehyde (HL₂) with [Ru(CHCHC₆H₄Me-4)Cl(CO)(BTD)(PPh₃)₂] in the presence of base. Two further ligands were investigated to extend the study to encompass 5- and 4-membered chelates; 8-hydroxyquinoline (HL₃) and 2-hydroxy-4-methylquinoline (HL₄) react with [Ru(CHCHPh)Cl(CO)(BTD)(PPh₃)₂] and [Ru(CHCHC₆H₄Me-4)Cl(CO)(BTD)(PPh₃)₂] in the presence of base to yield the complexes [Ru(CHCHPh)(L₃)(CO)(PPh₃)₂] and [Ru(CHCHC₆H₄Me-4)(L₄)(CO)(PPh₃)₂], respectively. The crystal structure of [Ru(CHCHC₆H₄Me-4)(L₁)(CO)(PPh₃)₂] is reported.     

Environmental and dietary influences on highly unsaturated fatty acid biosynthesis and expression of fatty acyl desaturase and elongase genes in liver of Atlantic salmon (Salmo salar)

Highly unsaturated fatty acid (HUFA) synthesis in Atlantic salmon (Salmo salar) was known to be influenced by both nutritional and environmental factors. Here we aimed to test the hypothesis that both these effectors involved similar molecular mechanisms. Thus, HUFA biosynthetic activity and the expression of fatty acyl desaturase and elongase genes were determined at various points during an entire 2 year production cycle in salmon fed diets containing either 100% fish oil or diets in which a high proportion (75% and 100%) of fish oil was replaced by C18 polyunsaturated fatty acid-rich vegetable oil. The results showed that HUFA biosynthesis in Atlantic salmon varied during the growth cycle with peak activity around seawater transfer and subsequent low activities in seawater. Consistent with this, the gene expression of Delta6 desaturase, the rate-limiting step in the HUFA biosynthetic pathway, was highest around the point of seawater transfer and lowest during the seawater phase. In addition, the expression of both Delta6 and Delta5 desaturase genes was generally higher in fish fed the vegetable oil-substituted diets compared to fish fed fish oil, particularly in the seawater phase. Again, generally consistent with this, the activity of the HUFA biosynthetic pathway was invariably higher in fish fed diets in which fish oil was substituted by vegetable oil compared to fish fed only fish oil. In conclusion, these studies showed that both nutritional and environmental modulation of HUFA biosynthesis in Atlantic salmon involved the regulation of fatty acid desaturase gene expression.     

Fatty acid utilisation and metabolism in caecal enterocytes of rainbow trout (Oncorhynchus mykiss) fed dietary fish or copepod oil

A combined fatty acid metabolism assay was employed to determine fatty acid uptake and relative utilisation in enterocytes isolated from the pyloric caeca of rainbow trout. In addition, the effect of a diet high in long-chain monoenoic fatty alcohols present as wax esters in oil derived from Calanus finmarchicus, compared to a standard fish oil diet, on caecal enterocyte fatty acid metabolism was investigated. The diets were fed for 8 weeks before caecal enterocytes from each dietary group were isolated and incubated with [1-14C]fatty acids: 16:0, 18:1n-9, 18:2n-6, 18:3n-3, 20:1n-9, 20:4n-6, 20:5n-3, and 22:6n-3. Uptake was measured over 2 h with relative utilisation of different [1-14C]fatty acids calculated as a percentage of uptake. Differences in uptake were observed, with 18:1n-9 and 18:2n-6 showing the highest rates. Esterification into cellular lipids was highest with 16:0 and C18 fatty acids, accounting for over one-third of total uptake, through predominant incorporation in triacylglycerol (TAG). The overall utilisation of fatty acids in phospholipid synthesis was low, but highest with 16:0, the most prevalent fatty acid recovered in intracellular phosphatidylcholine (PC) and phosphatidylinositol (PI), although exported PC exhibited higher proportions of C20/C22 polyunsaturated fatty acids (PUFA). Other than 16:0, incorporation into PC and PI was highest with C20/C22 PUFA and 20:4n-6 respectively. Recovery of labelled 18:1n-9 in exported TAG was 3-fold greater than any other fatty acid which could be due to multiple esterification on the glycerol 'backbone' and/or increased export. Approximately 20-40% of fatty acids taken up were beta-oxidised, and was highest with 20:4n-6. Oxidation of 20:5n-3 and 22:6n-3 was also surprisingly high, although 22:6n-3 oxidation was mainly attributed to retroconversion to 20:5n-3. Metabolic modification of fatty acids by elongation-desaturation was generally low at <10% of [1-14C]fatty acid uptake. Dietary copepod oil had generally little effect on fatty acid metabolism in enterocytes, although it stimulated the elongation and desaturation of 16:0 and elongation of 18:1n-9, with radioactivity recovered in longer n-9 monoenes. The monoenoic fatty acid, 20:1n-9, abundant in copepod oil as the homologous alcohol, was poorly utilised with 80% of uptake remaining unesterified in the enterocyte. However, the fatty acid composition of pyloric caeca was not influenced by dietary copepod oil.     

Can assignment tests measure dispersal?

Individual-based assignment tests are now standard tools in molecular ecology and have several applications, including the study of dispersal. The measurement of natal dispersal is vital to understanding the ecology of many species, yet the accuracy of assignment tests in situations where natal dispersal is common remains untested in the field. We studied a metapopulation of the grand skink, Oligosoma grande, a large territorial lizard from southern New Zealand. Skink populations occur on isolated, regularly spaced rock outcrops and are characterized by frequent interpopulation dispersal. We examined the accuracy of assignment tests at four replicate sites by comparing long-term mark-and-recapture records of natal dispersal with the results of assignment tests based on microsatellite DNA data. Assignment tests correctly identified the natal population of most individuals (65-100%, depending on the method of assignment), even when interpopulation dispersal was common (5-20% dispersers). They also provided similar estimates of the proportions of skinks dispersing to those estimated by the long-term mark-and-recapture data. Fully and partially Bayesian assignment methods were equally accurate but their accuracy depended on the stringency applied, the degree of genetic differentiation between populations, and the number of loci used. In addition, when assignments required high confidence, the method of assignment (fully or partially Bayesian) had a large bearing on the number of individuals that could be assigned. Because assignment tests require significantly less fieldwork than traditional mark-and-recapture approaches (in this study < 3 months vs. > 7 years), they will provide useful dispersal data in many applied and theoretical situations.     

Utilization of porcine pancreatic phospholipase A2 for the preparation of a marine fish oil enriched in (n - 3) polyunsaturated fatty acids

A simple and rapid method is described for the preparation of a marine oil fraction highly enriched in (n - 3) polyunsaturated fatty acids. Cod roe, containing lipid up to 15% of its dry weight, approximately 70% of which is phospholipid, was the starting material. Incubation of a concentrated aqueous extract of the roe with porcine pancreatic phospholipase A2 (EC 3.1.1.4) was the key step in the procedure. Extraction of the freeze-dried reaction product with diethyl ether containing formic acid produced an oil in a yield of 1.0 g/100 g wet wt of starting roe. The oil contained over 95% as free fatty acids, with 20:5 (n - 3) and 22:6 (n - 3) accounting for up to 24 and 40%, respectively, of the total free fatty acids. The therapeutic use of the oil is mentioned.