Production Of Long-chain Polyunsaturated Fatty Acids Research Articles

Capability of DHA biosynthesis in a marine teleost, Pacific saury Cololabis saira: functional characterization of two paralogous Fads2 desaturases and Elovl5 elongase

The Pacific saury Cololabis saira is an important fishery resource in the Far East, where it is appreciated for its high lipid content, which includes long-chain polyunsaturated fatty acids (LC-PUFAs) such as docosahexaenoic acid (DHA). Although it has been hypothesized that these fatty acids are derived from their prey items, this study focused on evaluating the capacity of saury for endogenous production of LC-PUFAs. To describe its LC-PUFA biosynthetic pathway, we characterized fatty acid desaturase 2 (Fads2) and elongation of very long-chain fatty acid protein 5 (Elovl5) in Pacific saury. Two paralogous fads2 genes, namely fads2a and fads2b, were isolated from the saury and their recombinant protein showed ∆6∆8 and ∆4∆5 desaturase activities, respectively. Meanwhile, saury Elovl5 had elongase activity toward C18 and C20 PUFA. These three enzyme genes were expressed in the brain and liver, although fads2a was absent in the latter. Our results suggest that the saury has sufficient enzymatic functions, particularly in its brain, for DHA biosynthesis through the ∆4 pathway even from α-linolenic acid. This information provides novel insights into the origin of LC-PUFAs in Pacific saury and the future perspective for its potential as a source of such vital fatty acids for human consumption.

Novel Interactions of Myristic Acid and FADS3 Variants Predict Atopic Dermatitis among Indonesian Infants.

Fatty acids exert a range of different biological activities that could be relevant in the development of atopic dermatitis (AD). This study investigated the association of glycerophospholipid fatty acids (GPL-FA) with AD, and their interactions with single nucleotide polymorphisms (SNP) of the FADS1-3 gene cluster. Among 390 infants of the Indonesian ISADI study, GPL-FA were measured in umbilical plasma (P-0y) and in buccal cells at birth (B-0y), and again in buccal cells at AD onset or one year (B-1y). Prospective and cross-sectional associations with AD were assessed by logistic regression. Interactions of GPL-FA with 14 SNP were tested assuming an additive model. AD was diagnosed in 15.4% of participants. In B-1y, C18:2n-6 was inversely associated with AD; and positive associations were observed for C18:1n-9, C20:4n-6, C22:6n-3 and C20:4n-6/C18:2n-6. There were no prospective associations with AD, however, a significant interaction between the SNP rs174449 and B-0y C14:0 (myristic acid) was observed. This study indicates that Indonesian infants with AD have increased rates of endogenous long-chain polyunsaturated fatty acid production, as well as higher C18:1n-9 levels. GPL-FA measured at birth do not predict later AD incidence; however, genotype interactions reveal novel effects of myristic acid, which are modified by a FADS3 variant.

Ligand diversity contributes to the full activation of the jasmonate pathway in Marchantia polymorpha

In plants, jasmonate signaling regulates a wide range of processes from growth and development to defense responses and thermotolerance. Jasmonates, such as jasmonic acid (JA), (+)-7-iso-jasmonoyl-l-isoleucine (JA-Ile), 12-oxo-10,15(Z)-phytodienoic acid (OPDA), and dinor-12-oxo-10,15(Z)-phytodienoic acid (dn-OPDA), are derived from C18 (18 Carbon atoms) and C16 polyunsaturated fatty acids (PUFAs), which are found ubiquitously in the plant kingdom. Bryophytes are also rich in C20 and C22 long-chain polyunsaturated fatty acids (LCPUFAs), which are found only at low levels in some vascular plants but are abundant in organisms of other kingdoms, including animals. The existence of bioactive jasmonates derived from LCPUFAs is currently unknown. Here, we describe the identification of an OPDA-like molecule derived from a C20 fatty acid (FA) in the liverwort Marchantia polymorpha (Mp), which we term (5Z,8Z)-10-(4-oxo-5-((Z)-pent-2-en-1-yl)cyclopent-2-en-1-yl)deca-5,8-dienoic acid (C20-OPDA). This molecule accumulates upon wounding and, when applied exogenously, can activate known Coronatine Insensitive 1 (COI1) -dependent and -independent jasmonate responses. Furthermore, we identify a dn-OPDA-like molecule (Δ4-dn-OPDA) deriving from C20-OPDA and demonstrate it to be a ligand of the jasmonate coreceptor (MpCOI1-Mp Jasmonate-Zinc finger inflorescence meristem domain [MpJAZ]) in Marchantia. By analyzing mutants impaired in the production of LCPUFAs, we elucidate the major biosynthetic pathway of C20-OPDA and Δ4-dn-OPDA. Moreover, using a double mutant compromised in the production of both Δ4-dn-OPDA and dn-OPDA, we demonstrate the additive nature of these molecules in the activation of jasmonate responses. Taken together, our data identify a ligand of MpCOI1 and demonstrate LCPUFAs as a source of bioactive jasmonates that are essential to the immune response of M. polymorpha.

Inhibition of Δ-6 desaturase reduces fatty acid re-esterification in 3T3-L1 adipocytes independent of changes in n3-PUFA cellular content

Δ-6 desaturase (D6D) is a key enzyme in the synthesis of long-chain polyunsaturated fatty acids (LC-PUFA). Evidence suggests that reduced D6D activity not only disrupts LC-PUFA production, but also impacts whole body lipid handling and body weight; however, the mechanisms remain largely unexplored. Therefore, we investigated the effect of D6D inhibition on the regulation of lipid accumulation in 3T3-L1 adipocytes with and without changes in n-3 PUFA content. 3T3-L1 cells were treated with a D6D inhibitor (SC-26196) in the presence or absence of α-linolenic acid (ALA) throughout differentiation. We found that D6D inhibition blocked the conversion of ALA to eicosapentaenoic acid (EPA) and docosapentaenoic acid (DPAn-3) when ALA was supplemented, while no changes in n-3 PUFA content were observed in cells treated with the D6D inhibitor alone. D6D inhibited cells had reduced triacylglycerol (TAG) accumulation despite an EPA/DPA deficiency. In addition, analyses of cellular protein markers, as well as non-esterified fatty acids and glycerol release in medium, suggested an increase in lipolysis and a decrease in fatty acid re-esterification in D6D-inhibited cells, independent of n-3 PUFA changes. To provide further evidence, we treated cells with the D6D inhibitor in the presence or absence of EPA and compared them with ALA-treated cells. Although EPA further reduced TAG content, the reduced markers of fatty acid re-esterification were not affected by ALA or EPA. Collectively, this study provides new insight showing that D6D inhibition reduces TAG accumulation and fatty acid re-esterification in adipocytes independent of changes in n-3 PUFA cellular content.

Diatoms and fatty acid production in Arctic and estuarine ecosystems—a reassessment of marine food webs, with a focus on the timing of shorebird migration

This review highlights diatoms and their production of fatty acids at Arctic ice edges and temperate estuaries as strategic in initiating high spring productivity. As important as their energy production is their synthesis de novo of omega-3 long-chain polyunsaturated fatty acids (LCPUFAs), which are bioactive molecules fundamental to maintaining ecosystem processes, and are necessary for reproduction and growth for the entire marine food web. Without these fatty acids, consumers can suffer poor reproduction and growth even with sufficient energy intake, and shorebirds may need LCPUFAs for nonstop long-hop migration. Timing of high energy fatty acid and LCPUFA production by diatoms coincides with growth and reproduction of consumer populations, with the arrival of seabirds in the Arctic and staging of shorebirds in estuaries. The switch from diatom production of carbohydrates to fatty acids is triggered by changes in chemical and physical environmental factors, which can be muted by factors such as climate change or habitat modification. Higher ocean temperatures and lower pH from climate change alters the ability of diatoms to make LCPUFAs, and habitat degradation reduces the numbers of diatoms in the area, subsequently reducing the amounts of LCPUFA produced. Changes in LCPUFA output could impact functionality of ice-edge and estuarine ecosystems, impacting shorebird migration and consumer productivity. Research is scarce on marine consumer reproduction and growth comparing quantity vs. quality of ingested saturated, monounsaturated, and omega-3 LCPUFAs, and nonexistent for seabirds and shorebirds. Future studies of marine food webs should include the role of diatoms and their contribution of not just energy, but also LCPUFAs to the food web.

Food and Feed Safety of NS-B5ØØ27-4 Omega-3 Canola (Brassica napus): A New Source of Long-Chain Omega-3 Fatty Acids.

DHA canola, a genetically engineered Brassica napus (OECD Unique Identifier NS-B5ØØ27-4), has been developed as one of the first land-based production systems for omega-3 long-chain polyunsaturated fatty acids (LCPUFA), whose health benefits are well-established. Yet, the marine sources of these nutrients are under high pressures due to over-fishing and increasing demand. DHA canola is a plant-based source for these essential fatty acids that produces a high level of docosahexaenoic acid (DHA). This terrestrial system allows for sustainable, scalable and stable production of omega-3 LCPUFA that addresses not only the increasing market demand, but also the complex interplay of agriculture, aquaculture, and human nutrition. The vector used to produce the desired oil profile in DHA canola contains the expression cassettes of seven genes in the DHA biosynthesis pathway and was specifically designed to convert oleic acid to DHA in canola seed. The characterization and safety evaluation of food and feed produced from DHA canola are described and supported by a detailed nutritional analysis of the seed, meal, and oil. Aside from the intended changes of the fatty acid profile, none of the other compositional analytes showed biologically meaningful differences when compared to conventional canola varieties. In addition, the meal from DHA canola is compositionally equivalent to conventional canola meal. Further evidence of nutritional value and safety of DHA canola oil have been confirmed in fish feeding studies. Given that most human populations lack sufficient daily intakes of omega-3 LCPUFA, a dietary exposure assessment is also included. In conclusion, the results from these studies demonstrate it is safe to use products derived from DHA canola in human foods, nutraceuticals, or animal feeds.

Fatty acid profiles and omega-3 long-chain polyunsaturated fatty acids (LC-PUFA) biosynthesis capacity of three dual purpose chicken breeds

Westernized societies ingest an unhealthy high dietary omega-6/omega-3 fatty acid ratio of 20:1 or even higher. Seafood is the primary source of omega-3 long chain polyunsaturated fatty acids (LC-PUFA) for humans, mainly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which are involved in a plethora of physiological and health-related processes. However, the production of marine organisms with aquafeed formulations based on marine ingredients leads to serious environmental impacts on global pelagic fish resources, resulting in an unsustainable activity. The present study aims to gain further insight into the metabolism of fatty acids in chicken as a potential supply for omega-3 LC-PUFA. To this purpose, lipid classes and fatty acid profiles of thighs and hepatocytes, and the modification of [1-14C]18:3n-3 by hepatocytes isolated from three dual-purpose chicken breeds adapted to free-range culture systems were determined. Arachidonic acid highly accumulated in thighs meat (7.16–8.79 %) despite being barely supplied in the diet, with DHA (1.22–1.71 %) and n-3 docosapentaenoic acid (DPA, 22:5n-3; 1.02–1.14 %) being also relevant. Our experimental design with radiolabeled fatty acids was validated for the first time in terrestrial vertebrates. Chicken hepatocytes incubated with [1-14C]18:3n-3 produced a wide variety of C18-C24 intermediates demonstrating that the set of fatty acyl desaturases and elongases enzymes necessary to metabolize dietary C18 precursors are active for the production of LC-PUFA, including EPA, n-3 DPA and DHA.

Influence of dietary n‐3 polyunsaturated fatty acids (PUFAs) on growth, PUFA composition and the expression of FAD6 and ELOV5 in pacific white shrimp ( Litopenaeus vannamei )

Long-chain polyunsaturated fatty acids (LC-PUFAs) play crucial roles in growth, development and reproduction of Litopenaeus vannamei, one of the most important shrimp species cultivated in China in recent years. To investigate the effect of dietary n-3 PUFA composition on growth and the expression of LvFAD6 and LvELOV5 genes and fatty acid composition of this species, ten diets, which contain different levels of α-linolenic acid (C18:3 n-3, ALA), eicosapentaenoic acid (C20:5 n-3, EPA) and docosahexaenoic acid (C22:6 n-3, DHA), respectively, were prepared for the culture of this species lasting 42 days. The results showed that the identified cDNA sequence of fatty acyl desaturases 6 from L. vannamei (LvFAD6) was 739 bp, encoding a protein of 210 amino acids. The LvFAD6 peptide sequence possessed two histidine-rich motifs (QHHA and QIEHH), shared 96.6% identity with Apostichopus japonicus FAD6 and strongly clustered with FAD6 from A. japonicus and Octopus bimaculoides. The expression of LvFAD6 and LvELOV5 in muscle decreased with increase in dietary EPA, but increased with increase in dietary DHA. Dietary ALA has the potential to further increase the production of these health-beneficial LC-PUFAs in L. vannamei fed a diet with low levels of EPA and DHA, implying L. vannamei had the capacity to convert dietary ALA into EPA and DHA. These results suggest that the supplementation of ALA, EPA and DHA in diets for better growth of L. vannamei in ranges of 2.5% to 3.89%, 1.7% to 3.19% and 2.7% to 5.84% of total fatty acids respectively.

Nutritional regulation of genes responsible for long-chain (C20-24) and very long-chain (>C24) polyunsaturated fatty acid biosynthesis in post-larvae of gilthead seabream (Sparus aurata) and Senegalese sole (Solea senegalensis)

The fatty acyl elongases Elovl4 are pivotal components in the biosynthesis of very long-chain polyunsaturated fatty acids (VLC-PUFA) from long-chain polyunsaturated fatty acids (LC-PUFA). Thus, nutritional regulation of Elovl4, as well as other elongase and desaturase genes involved in LC-PUFA biosynthesis (e.g., Elovl5, Fads2) has been proposed as a strategy to enhance endogenous production of LC-PUFA and VLC-PUFA under intensive farming conditions. This study aimed at investigating the nutritional regulation of genes involved in the biosynthesis of VLC-PUFA (elovl4 isoforms a and b) and LC-PUFA (fads2, elovl5) in Sparus aurata and Solea senegalensis post-larvae fed three inert micro-diets with graded concentrations of dietary LC-PUFA by using different combinations of fish oil and soya oil. The effect of dietary LC-PUFA on survival, growth and fatty acid composition was also examined. The results denoted that, while no effects were observed in survival, fish fed the diet with the highest LC-PUFA content during 30 d showed a higher total length and wet weight. Gene expression results showed that fads2, elovl5, elovl4a and elovl4b can be regulated by dietary LC-PUFA content. Remarkably, our results denoted a differential elovl4 nutritional regulation associated to each species. The head is the body part studied with the highest elovl4 transcripts in both fish species.

Assessment of Fatty Acid Desaturase (Fads2) Structure-Function Properties in Fish in the Context of Environmental Adaptations and as a Target for Genetic Engineering.

Fatty acid desaturase 2 (Fads2) is the key enzyme of long-chain polyunsaturated fatty acid (LC-PUFA) biosynthesis. Endogenous production of these biomolecules in vertebrates, if present, is insufficient to meet demand. Hence, LC-PUFA are considered as conditionally essential. At present, however, LC-PUFA are globally limited nutrients due to anthropogenic factors. Research attention has therefore been paid to finding ways to maximize endogenous LC-PUFA production, especially in production species, whereby deeper knowledge on molecular mechanisms of enzymatic steps involved is being generated. This review first briefly informs about the milestones in the history of LC-PUFA essentiality exploration before it focuses on the main aim—to highlight the fascinating Fads2 potential to play roles fundamental to adaptation to novel environmental conditions. Investigations are summarized to elucidate on the evolutionary history of fish Fads2, providing an explanation for the remarkable plasticity of this enzyme in fish. Furthermore, structural implications of Fads2 substrate specificity are discussed and some relevant studies performed on organisms other than fish are mentioned in cases when such studies have to date not been conducted on fish models. The importance of Fads2 in the context of growing aquaculture demand and dwindling LC-PUFA supply is depicted and a few remedies in the form of genetic engineering to improve endogenous production of these biomolecules are outlined.

Polyunsaturated fatty acid production by Yarrowia lipolytica employing designed myxobacterial PUFA synthases

Long-chain polyunsaturated fatty acids (LC-PUFAs), particularly the omega-3 LC-PUFAs eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), and docosahexaenoic acid (DHA), have been associated with beneficial health effects. Consequently, sustainable sources have to be developed to meet the increasing demand for these PUFAs. Here, we demonstrate the design and construction of artificial PUFA biosynthetic gene clusters (BGCs) encoding polyketide synthase-like PUFA synthases from myxobacteria adapted for the oleaginous yeast Yarrowia lipolytica. Genomic integration and heterologous expression of unmodified or hybrid PUFA BGCs yielded different yeast strains with specific LC-PUFA production profiles at promising yield and thus valuable for the biotechnological production of distinct PUFAs. Nutrient screening revealed a strong enhancement of PUFA production, when cells were phosphate limited. This represents, to the best of our knowledge, highest concentration of DHA (16.8 %) in total fatty acids among all published PUFA-producing Y. lipolytica strains.

Growth performance and immune status in common carp Cyprinus carpio as affected by plant oil-based diets complemented with β-glucan

Omnivorous fish species such as the common carp (Cyprinus carpio) are able to biosynthesise long chain polyunsaturated fatty acids (LC-PUFAs) from plant oil PUFA precursors, but the influence of the amount and quality of the LC-PUFAs biosynthesised from these oils on the immunocompetence status of the fish has received little attention. This study aims to evaluate whether the conversion of PUFA by carp induces a sufficient biosynthesis of LC-PUFA to maintain a good immunocompetence status in this species. Six iso-nitrogenous (crude protein = 39.1%) and iso-lipidic (crude lipids = 10%) diets containing three different lipid sources (cod liver oil (CLO) as fish oil; linseed oil (LO) and sunflower oil (SFO) as plant oils) were formulated with or without β-glucan supplementation at 0.25 g/kg diet. Juvenile carp (16.3 ± 0.6 g initial body weight) were fed a daily ration of 4% body weight for 9 weeks and then infected at day 64 with the bacteria Aeromonas hydrophyla. No significant differences in survival rate, final body weight, specific growth rate and feed conversion rate were observed between diets. After bacterial infection, mortality rate did not differ between fish fed CLO and plant oil-based diets, indicating that the latter oils did not affect the overall immunocompetence status of common carp. Plant oil-based diets did not alter lysozyme activity in healthy and infected fish. No negative effects of plant oils on complement activity (ACH50) were observed in healthy fish, even if both plant oil-based diets induced a decrease in stimulated fish two days after infection. Furthermore, the levels of various immune genes (nk, lys, il-8, pla, pge, alox) were not affected by plant oil-based diets. The expression of pla and pge genes were higher in SFO-fed fish than in CLO ones, indicating that this plant oil rich in linoleic acid (LA) better stimulated the eicosanoid metabolism process than fish oil. In response to β-glucan supplementation, some innate immune functions seemed differentially affected by plant oil-based diets. LO and SFO induced substantial LC-PUFA production, even if fish fed CLO displayed the highest EPA and DHA levels in tissues. SFO rich in LA induced the highest ARA levels in fish muscle while LO rich in α-linolenic acid (ALA) sustained higher EPA production than SFO. A significantly higher fads−6a expression level was observed in SFO fish than in LO ones, but this was not observed for elovl5 expression. In conclusion, the results show that common carp fed plant oil-based diets are able to produce substantial amounts of LC-PUFA for sustaining growth rate, immune status and disease resistance similar to fish fed a fish oil-based diet. The differences in the production capacity of LC-PUFAs by the two plant oil-based diets were associated to a differential activation of some immune pathways, explaining how the use of these oils did not affect the overall immunocompetence of fish challenged with bacterial infection. Moreover, plant oil-based diets did not induce substantial negative effects on the immunomodulatory action of β-glucans, confirming that these oils are suitable for sustaining a good immunocompetence status in common carp.

Metabolic Engineering Strategies for the Enhanced Microalgal Production of Long-Chain Polyunsaturated Fatty Acids (LC-PUFAs).

Long-chain polyunsaturated fatty acids (LC-PUFAs), largely obtained from fish oil, serve as valuable dietary supplements with many health benefits, especially arachidonic acid, eicosapentaenoic acid, and docosahexaenoic acid. In recent years, interest in the sustainable production of LC-PUFAs using heterologous production hosts has drastically increased because overfishing and polluted oceans have led to a gradual decline in the supply of LC-PUFAs from fish oil in the last few decades. Some species of microalgae have been considered to be an ideal producer of LC-PUFAs as they inherently accumulate large amounts of LC-PUFAs and utilize CO2 using light energy. Also, a growing number of genetic toolboxes have become available, and the microalgal cultivation process is amenable to a scale-up. In fact,tremendous progress has been achieved in the development of high-performance strains of microalgae through metabolic engineering that has led to the efficient production of fatty acids and various derivatives in the last decade. This review discusses metabolic engineering strategies that have contributed to the enhanced production of LC-PUFAs using microalgae, including optimizing fatty acid biosynthetic pathway and intracellular supply of precursors and cofactors, as well as engineering transcription factors.

Optimizing long chain-polyunsaturated fatty acid synthesis in salmonids by balancing dietary inputs.

The increasing use of terrestrial plant lipids to replace of fish oil in commercial aquafeeds requires understanding synthesis and storage of long chain-polyunsaturated fatty acids (LC-PUFA) in farmed fish. Manipulation of dietary fatty acids may maximize tissue storage of LC-PUFA, through increased production and selective utilization. A data synthesis study was conducted to estimate optimal levels of fatty acids that may maximize the production and storage of LC-PUFA in the edible portion of salmonids. Data were compiled from four studies with Atlantic salmon, rainbow trout, and steelhead trout (total n = 180) which were fed diets containing different terrestrial-based oils to replace fish oil. LC-PUFA (%) were linearly correlated between diet and muscle tissue (p < 0.001; r2 > 44%), indicating proportional storage after consumption. The slope, or retention rate, was highest for docosahexaenoic acid (DHA) at 1.23, indicating that an additional 23% of DHA was stored in the muscle. Dietary saturated fatty acids were positively related to DHA stored in the muscle (p < 0.001; r2 = 22%), which may involve membrane structural requirements, as well as selective catabolism. DHA was found to be optimally stored with a dietary n-3: n-6 ratio of 1.03: 1. These new results provide a baseline of optimal dietary ratios that can be tested experimentally to determine the efficacy of balancing dietary fatty acids for maximum LC-PUFA storage.

Impact of temperature on fatty acid composition and nutritional value in eight species of microalgae.

Microalgae are considered a sustainable source of high-value products with health benefits. Marine algae-derived omega-3 long-chain polyunsaturated fatty acids (LC-PUFA), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), are considered dietary elements with effects on mental health, cognition enhancement, and cardiovascular protection. This study investigated the temperature effect on omega-3 LC-PUFA production in eight species of microalgae from various taxonomic groups, with a focus on achieving an optimal balance between omega-3 accumulation and efficient growth performance. Samples were batch-cultivated at four different temperatures, with constant light, and fatty acid methyl esters (FAME) were analyzed by gas chromatography. Several nutritional indices were calculated to assess the potential value of biomass produced for human consumption. Two promising candidates were identified suitable for batch cultivation and large-scale production: Nannochloropsis oculata for EPA and Isochrysis galbana for DHA production, with optimum productivities obtained between 14 and 20°C, and nutritional indices falling within the range required for nutritional benefit.

Molecular characterization, tissue distribution and differential nutritional regulation of putative Elovl5 elongase in silver barb (Puntius gonionotus)

Fatty acid desaturase (Fads) and elongase (Elovl) are two important enzymes involved in the biosynthesis of long-chain polyunsaturated fatty acids (LC-PUFAs) in fishes. Silver barb (Puntius gonionotus) is considered a promising medium carp species for freshwater aquaculture in Asia. Earlier, Δ6 fads cDNA was characterized to understand the molecular mechanism of LC-PUFA biosynthesis in this species. In the present study elovl5-like elongase full-length cDNA was cloned and characterized by RACE (rapid amplification of cDNA ends), and analyzed its basal expression in various organs/tissues by quantitative real time PCR (qRT-PCR) assay. Differential expression of elovl5 cDNA in various tissues following replacement of fish oil (FO) with linseed oil (LO) in the diet also studied. The full length cDNA sequence of silver barb elovl5-like elongase consisted of 1853bp, including an open reading frame (ORF) of 876bp encoding a precursor protein of 291 amino acids possessing the entire characteristic features of Elovl protein. The elovl5 transcript was constitutively expressed in all the tissues examined: muscle, liver, brain, kidney, skin, intestine, gill and eye. The expression of elovl5 gene was upregulated, whereas the serum biochemical constituents were either remain unchanged or lowered following replacement of FO with LO in the diets. We report for the first time, an in silico 3D structure model of elongase protein from silver barb. All these findings may contribute to a better understanding the potential regulatory mechanisms involved and may be useful for enhancing endogenous LC-PUFA production in silver barb.

Enrichment of Long-Chain Polyunsaturated Fatty Acids by Coordinated Expression of Multiple Metabolic Nodes in the Oleaginous Microalga Phaeodactylum tricornutum.

Microalgal long-chain polyunsaturated fatty acids (LC-PUFAs) have emerged as promising alternatives to depleting fish oils. However, the overproduction of LC-PUFAs in microalgae has remained challenging. Here, we report a sequential metabolic engineering strategy that systematically overcomes the metabolic bottlenecks and overproduces LC-PUFAs. Malonyl CoA-acyl carrier protein transacylase, catalyzing the first committed step in type II fatty acid synthesis, and desaturase 5b, involved in fatty acid desaturation, were coordinately expressed in Phaeodactylum tricornutum. Engineered microalgae hyper-accumulated LC-PUFAs, with arachidonic acid (ARA) and docosahexaenoic acid (DHA) contents of up to 18.98 μg/mg and 9.15 μg/mg (dry weight), respectively. Importantly, eicosapentaenoic acid (EPA) was accumulated up to a highest record of 85.35 μg/mg by metabolic engineering. ARA and EPA were accumulated mainly in triacylglycerides, whereas DHA was found exclusively in phospholipids. Combinatorial expression of these critical enzymes led to the optimal increment of LC-PUFAs without unbalanced metabolic flux and demonstrated the practical feasibility of generating sustainable LC-PUFA production.

Association of maternal weight with FADS and ELOVL genetic variants and fatty acid levels- The PREOBE follow-up.

Single nucleotide polymorphisms (SNPs) in the genes encoding the fatty acid desaturase (FADS) and elongase (ELOVL) enzymes affect long-chain polyunsaturated fatty acid (LC-PUFA) production. We aimed to determine if these SNPs are associated with body mass index (BMI) or affect fatty acids (FAs) in pregnant women. Participants (n = 180) from the PREOBE cohort were grouped according to pre-pregnancy BMI: normal-weight (BMI = 18.5–24.9, n = 88) and overweight/obese (BMI≥25, n = 92). Plasma samples were analyzed at 24 weeks of gestation to measure FA levels in the phospholipid fraction. Selected SNPs were genotyped (7 in FADS1, 5 in FADS2, 3 in ELOVL2 and 2 in ELOVL5). Minor allele carriers of rs174545, rs174546, rs174548 and rs174553 (FADS1), and rs1535 and rs174583 (FADS2) were nominally associated with an increased risk of having a BMI≥25. Only for the normal-weight group, minor allele carriers of rs174537, rs174545, rs174546, and rs174553 (FADS1) were negatively associated with AA:DGLA index. Normal-weight women who were minor allele carriers of FADS SNPs had lower levels of AA, AA:DGLA and AA:LA indexes, and higher levels of DGLA, compared to major homozygotes. Among minor allele carriers of FADS2 and ELOVL2 SNPs, overweight/obese women showed higher DHA:EPA index than the normal-weight group; however, they did not present higher DHA concentrations than the normal-weight women. In conclusion, minor allele carriers of FADS SNPs have an increased risk of obesity. Maternal weight changes the effect of genotype on FA levels. Only in the normal-weight group, minor allele carriers of FADS SNPs displayed reduced enzymatic activity and FA levels. This suggests that women with a BMI≥25 are less affected by FADS genetic variants in this regard. In the presence of FADS2 and ELOVL2 SNPs, overweight/obese women showed higher n-3 LC-PUFA production indexes than women with normal weight, but this was not enough to obtain a higher n-3 LC-PUFA concentration.

Effects of different dietary ratios of linolenic to linoleic acids or docosahexaenoic to eicosapentaenoic acids on the growth and immune indices in grouper, Epinephelus coioides

In order to investigate whether juvenile grouper (Epinephelus coioides) has biosynthetic ability of long-chain polyunsaturated fatty acids (LC-PUFA) and the proper ratio of docosahexaenoic (DHA) to eicosapentaenoic (EPA) acids in formula diets, a feeding trial was conducted with seven isonitrogenous (55.2%) and isolipidic (10.8%) diets (D0–D6) in sea floating net cages. D0 (control diet) used fish oil (FO) as lipid resource, D1–D3 contained blends of perilla and corn oils at linolenic/linoleic acids (ALA/LA) ratios of 1.0, 2.0 and 3.0, while D4–D6 contained blends of corn oil and purified EPA- or DHA-enriched oil at DHA/EPA ratios of 1.0, 2.0 and 3.0. After 9weeks of the feeding trial, the weight gain (WG) and specific growth rate (SGR) of D1–D3 groups were significantly lower than those of D4–D5 groups (P<0.05), but showed no significant differences compared with that of D0 or D6 group (P>0.05). D4 group showed the highest WG and SGR as well as the lowest estimated feed conversion rate (FCR), followed by the D5 group. The levels of EPA, DHA and docosapentaenoic acid (DPA) in liver of D1–D3 groups were significantly lower than those of D0 or D4–D6 groups (P<0.05), while those of EPA, DHA and total n-3 LC-PUFA in the muscle of D1–D3 groups were markedly lower than that of D4 group (P<0.05). Both the albumin and IgM concentrations in the serum of D1–D3 groups were significantly lower than those of D4 and D5 groups, respectively (P<0.05). Whereas the serum lysozyme activity of D3 group and alkaline phosphatase (AKP) activity of D1 group were respectively significantly lower than that of D0 and D4 groups (P<0.05). These results indicated that juvenile E. coioides might have a low biosynthetic capacity for production of LC-PUFA from LA and ALA, and good growth performance and health as well as high muscle LC-PUFA contents were obtained in fish fed D4 or D5 diets where the DHA/EPA ratio was about 1:1 or 2:1 and total n-3 LC-PUFA level was approximate to 1.83% in current study.

Biosynthesis of Polyunsaturated Fatty Acids in Sea Urchins: Molecular and Functional Characterisation of Three Fatty Acyl Desaturases from Paracentrotus lividus (Lamark 1816).

Sea urchins are broadly recognised as a delicacy and their quality as food for humans is highly influenced by their diet. Lipids in general and the long-chain polyunsaturated fatty acids (LC-PUFA) in particular, are essential nutrients that determine not only the nutritional value of sea urchins but also guarantee normal growth and reproduction in captivity. The contribution of endogenous production (biosynthesis) of LC-PUFA in sea urchins remained unknown. Using Paracentrotus lividus as our model species, we aimed to characterise both molecularly and functionally the repertoire of fatty acyl desaturases (Fads), key enzymes in the biosynthesis of LC-PUFA, in sea urchins. Three Fads, namely FadsA, FadsC1 and FadsC2, were characterised. The phylogenetic analyses suggested that the repertoire of Fads within the Echinodermata phylum varies among classes. On one hand, orthologues of the P. lividus FadsA were found in other echinoderm classes including starfishes, brittle stars and sea cucumbers, thus suggesting that this desaturase is virtually present in all echinoderms. Contrarily, the FadsC appears to be sea urchin-specific desaturase. Finally, a further desaturase termed as FadsB exists in starfishes, brittle stars and sea cucumbers, but appears to be missing in sea urchins. The functional characterisation of the P. lividus Fads confirmed that the FadsA was a Δ5 desaturase with activity towards saturated and polyunsaturated fatty acids (FA). Moreover, our experiments confirmed that FadsA plays a role in the biosynthesis of non-methylene interrupted FA, a group of compounds typically found in marine invertebrates. On the other hand, both FadsC desaturases from P. lividus showed Δ8 activity. The present results demonstrate that P. lividus possesses desaturases that account for all the desaturation reactions required to biosynthesis the physiological essential eicosapentaenoic and arachidonic acids through the so-called “Δ8 pathway”.