The Dietary Inclusion of Ensiled Olive Cake Increases Unsaturated Lipids in Milk and Alters the Expression of Lipogenic Genes in Mammary and Adipose Tissue in Goats.

This study aimed to evaluate the effect of the dietary inclusion of ensiled OC on milk yield, composition, fatty acid (FA) profile, and the expression of selected genes involved in lipid metabolism in the udder and adipose tissue of goats. Seventy-two Damascus dairy goats in mid-lactation were assigned randomly to three iso-nitrogenous and iso-energetic diets containing 0, 10, and 20% of ensiled OC as a replacement of forage (OC0, OC10, and OC20, respectively) for 42 days. During weeks 5 and 6 of the trial, dry matter intake, milk yield, milk composition, and FA profiles were recorded, while mammary and perirenal adipose tissue samples were also collected from six animals per treatment from the OC0 and OC20 groups for gene expression analysis. No significant differences were observed among groups concerning milk yield, 4% fat-corrected milk, fat, or protein yield (kg/d). In contrast, the milk fat percentage was gradually increased with increasing OC inclusion rates in the diets, while milk protein percentages were elevated in both OC groups but significantly only in the milk of the OC20 group. The content of FA between C4:0 to C16:0 was reduced, while mono-unsaturated FA (MUFA) concentration was enhanced in the goat milk of OC groups. The OC feeding treatment was associated with the increased mammary expression of SLC2A1 (p < 0.05), VLDLR (p < 0.01), FABP3 (p < 0.01), and elevated SLC2A1 (p < 0.05) and FASN (p < 0.01) gene expression in the adipose tissue of goats fed the OC20 diet. Overall, OC can be used in goats' diets as a forage replacement, at least in the inclusion rate of 20% DM, since this could increase the milk protein and fat percentage and enrich its content with beneficial for human health lipids without adversely affecting milk production traits.

Effect of Farming System (Organic vs. Conventional) and Season on Composition and Fatty Acid Profile of Bovine, Caprine and Ovine Milk and Retail Halloumi Cheese Produced in Cyprus.

The present work aimed to evaluate the effect of farming practices and season on the fat and protein content and fatty acid (FA) profile of milk and Halloumi cheese produced in Cyprus. Over a year, raw bulk-tank milk samples from cow, goat, and sheep farms were collected seasonally from all organic (11) and representative conventional (44) dairy farms, whereas Fresh Halloumi cheese samples were collected monthly from retail outlets (48 organic and 48 conventional samples in total). The different farming practices did not affect the milk fat content of ruminants, while protein levels were decreased in organic bovine and caprine milk. Under organic farming practices, milk and cheese contained increased values of total mono-unsaturated FA (MUFA) and poly-unsaturated FA (PUFA), and specific FA, such as oleic, conjugated linoleic, linoleic, and α-linolenic acids. Total saturated FA (SFA) levels were particularly decreased in organic samples and, consequently, the atherogenic indices of milk and cheese were decreased. Season influenced milk and Halloumi cheese FA profile; spring samples had lower SFA and higher PUFA and MUFA concentrations. Overall, the organic farm practices improved the lipid profile of milk and Halloumi cheese, which is more likely attributed to the different feeding strategies applied in organic dairy farms.

Short-term forage substitution with ensiled olive cake increases beneficial milk fatty acids in lactating cows.

This study aimed to evaluate the effect of short-term forage substitution with ensiled olive cake (OC), on yield, composition and fatty acid (FA) profile of cows' milk. Mid-lactating Holstein-Friesian cows were randomly assigned for 21 days to two isoenergetic and isoproteic feeding treatments (12 animals per treatment), containing 0 and 10% DM of ensiled OC (C and OC groups, respectively). Milk yield was recorded daily, and milk samples were collected at 14 and 21 days of the trial for analyzing the fat, protein, and FA profile of milk. No significant differences were observed in milk yield, protein, and fat nor in protein and fat percentage of milk between groups. However, dietary supplementation with ensiled OC modified the FA profile of cow milk. Feeding cows with ensiled OC resulted in a significant decline of medium-chain FA, while long-chain and mono-unsaturated FA were risen in milk (P < 0.05). Among individual saturated FA, palmitic was particularly reduced, while among individual mono-unsaturated FA, increments of C18:1 cis-9 were demonstrated with the OC treatment (P < 0.05). Although total poly-unsaturated FA were decreased, the concentration of CLA cis-9, trans-11 tended to be elevated with OC feeding (P = 0.06). Overall, short-term forage substitution with ensiled OC improved, beneficially for human health, the lipid profile of milk without adversely affecting milk yield or milk composition of lactating cows.

ACAA2 and FASN polymorphisms affect the fatty acid profile of Chios sheep milk.

The objective of the research reported in this research communication was the identification and association of single nucleotide polymorphisms (SNP) in the ovine DGAT1, FASN, SCD1 and ACAA2 genes with milk fat percentage and fatty acid (FA) content. Three consecutive monthly milk samplings were obtained from a total of 429 purebred Chios ewes during mid-lactation. Genotypic data were jointly analyzed with 1184 fat content and 37 718 FA percentage records using mixed models. The 3' untranslated region (UTR) of the DGAT1 gene and the 5' and 3'UTRs of the SCD1 gene appeared to be monomorphic. The FASN g.14777C>T SNP on exon 31 was associated with C13:0 and the ACAA2 g.2982T>C SNP on the 3'UTR was associated with C9:0, C11:0, C12:1 cis-9, C13:0 and the ω6/ω3 index, while fat percentage was not affected by the identified SNPs. The results could be useful for breeding programs aiming to improve the quality and nutritional value of ovine milk.

NMR-Based Μetabolomics of the Lipid Fraction of Organic and Conventional Bovine Milk.

Origin and quality identification in dairy products is an important issue and also an extremely challenging and complex experimental procedure. The objective of the present work was to compare the metabolite profile of the lipid fraction of organic and conventional bovine milk using NMR metabolomics analysis. ¹H-NMR and 1D TOCSY NMR methods of analysis were performed on extracted lipid fraction of lyophilized milk. For this purpose, 14 organic and 16 conventional retail milk samples were collected monthly, and 64 bulk-tank (58 conventional and 6 organics) milk samples were collected over a 14-month longitudinal study in Cyprus. Data were treated with multivariate methods (PCA, PLS-DA). Minor components were identified and quantified, and modification of the currently used equations is proposed. A significantly increased % content of conjugated (9-cis, 11-trans)18:2 linoleic acid (CLA), α-linolenic acid, linoleic acid, allylic protons and total unsaturated fatty acids (UFA) and decreased % content for caproleic acid were observed in the organic samples compared to the conventional ones. The present work confirms that lipid profile is affected by contrasting management system (organic vs. conventional), and supports the potential of NMR-based metabolomics for the rapid analysis and authentication of the milk from its lipid profile.

Selective One-Dimensional Total Correlation Spectroscopy Nuclear Magnetic Resonance Experiments for a Rapid Identification of Minor Components in the Lipid Fraction of Milk and Dairy Products: Toward Spin Chromatography?

We report a rapid, direct, and unequivocal spin-chromatographic separation and identification of minor components in the lipid fraction of milk and common dairy products with the use of selective one-dimensional (1D) total correlation spectroscopy (TOCSY) nuclear magnetic resonance (NMR) experiments. The method allows for the complete backbone spin-coupling network to be elucidated even in strongly overlapped regions and in the presence of major components from 4 × 10(2) to 3 × 10(3) stronger NMR signal intensities. The proposed spin-chromatography method does not require any derivatization steps for the lipid fraction, is selective with excellent resolution, is sensitive with quantitation capability, and compares favorably to two-dimensional (2D) TOCSY and gas chromatography-mass spectrometry (GC-MS) methods of analysis. The results of the present study demonstrated that the 1D TOCSY NMR spin-chromatography method can become a procedure of primary interest in food analysis and generally in complex mixture analysis.

Direct nuclear magnetic resonance identification and quantification of geometric isomers of conjugated linoleic acid in milk lipid fraction without derivatization steps: overcoming sensitivity and resolution barriers.

We report the first successful direct and unequivocal identification and quantification of four minor geometric (9-cis, 11-trans) 18:2, (9-trans, 11-cis) 18:2, (9-cis, 11-cis) 18:2 and (9-trans, 11-trans) 18:2 conjugated linoleic acid (CLA) isomers in lipid fractions of lyophilized milk samples with the combined use of 1D (1)H-NMR, 2D (1)H-(1)H TOCSY and 2D (1)H-(13)C HSQC NMR. The significant sensitivity barrier has been successfully overcome under selective suppression of the major resonances, with over 10(4) greater equilibrium magnetization of the -(CH2)n-(1)H spins compared to that of the (1)H spins of the conjugated bonds of the CLA isomers. The resolution barrier has been significantly increased using reduced (13)C spectral width in the 2D (1)H-(13)C HSQC experiment. The assignment was confirmed with spiking experiments with CLA standard compounds and the method does not require any derivatization steps for the lipid fraction. The proposed method is selective, sensitive and compares favorably with the GS-MS method of analysis.

Genetic and proteomic analysis of the MHC class I repertoire from four ovine haplotypes.

Immunity to livestock diseases can be studied directly in the target animal, but its elucidation is often constrained by the lack of major histocompatibility complex (MHC)-defined animals. To address this issue, we have established an MHC-defined sheep resource flock generated around four diverse MHC haplotypes. Initial characterisation of the repertoire of transcribed MHC class I genes identified three class I transcripts associated with each haplotype. Nucleotide sequence, transcript abundance and phylogenetic analysis indicated that they represent alleles at up to four polymorphic loci that vary in number between the different haplotypes. The functional significance of each of these genes is evaluated here using complementary molecular genetic and proteomic approaches. We determine which genes give rise to proteins that localise to the surface of transfected cells. In addition, we provide data to support the generation of expressed products, based on immunoprecipitation of class I products from animals homozygous for each of the four MHC haplotypes followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This provides a clearer picture of the number of MHC class I loci in sheep and allows more rational prediction of their classical (class Ia) or non-classical (class Ib) nature. On the basis of the cellular localisation, phylogenetic and transcriptional analyses, we propose that the ovine MHC comprises a minimum of eight class I loci, with considerable variation between haplotypes.

Haplotype characterization of transcribed ovine major histocompatibility complex (MHC) class I genes.

The ovine major histocompatibility complex (MHC) remains poorly characterized compared with those of other livestock species. Molecular genetic analysis of the bovine MHC has revealed considerable haplotype and allelic diversity that earlier serological analysis had not detected. To develop cellular and molecular tools to support development of vaccines against intracellular pathogens of sheep, we have undertaken a molecular genetic analysis of four distinct ovine MHC haplotypes carried by two heterozygous Blackface rams. We have identified 12 novel class I transcripts and used a class I sequence-specific genotyping system to assign each of these transcripts to individual haplotypes. Using a combination of phylogenetic analysis, haplotype and transcript expression data, we identified at least four distinct polymorphic class I MHC loci, three of which appear together in a number of combinations in individual haplotypes. The haplotypes were further characterized at the highly polymorphic Ovar-DRB1 locus, allowing selection of the progeny of the two founder rams for the establishment of an MHC-defined resource population.