M. Tor

Age-related changes in intramuscular and subcutaneous fat content and fatty acid composition in growing pigs using longitudinal data

The evolution of intramuscular fat (IMF) and subcutaneous backfat (SB) content and fatty acid composition with age was investigated. A repeated sampling scheme (n=517) for longissimus dorsi (LM) and SB specimens was performed from 160 to 220 days on 216 purebred Duroc barrows. IMF content increased linearly with age at a rate of 0.05%/day, with major compositional changes occurring in C18:1 and C18.2, which increased and decreased by around 4%, respectively. Backfat thickness increase rate declined with age, showing similar compositional changes to IMF but halved in magnitude. Fat content itself also influenced fat composition. At a fixed age, both saturated (SFA) (C16:0) and monounsaturated (MUFA) (C18:1) increased with IMF but only SFA (C16:0 and C18:0) with SB. Aging mostly affected MUFA while fatness SFA. A delay in age at slaughter will enhance IMF and C18:1 in relation to overall fatness but at expense of decreasing body and loin growth rate.

A Functional Variant in the Stearoyl-CoA Desaturase Gene Promoter Enhances Fatty Acid Desaturation in Pork

There is growing public concern about reducing saturated fat intake. Stearoyl-CoA desaturase (SCD) is the lipogenic enzyme responsible for the biosynthesis of oleic acid (18∶1) by desaturating stearic acid (18∶0). Here we describe a total of 18 mutations in the promoter and 3′ non-coding region of the pig SCD gene and provide evidence that allele T at AY487830:g.2228T>C in the promoter region enhances fat desaturation (the ratio 18∶1/18∶0 in muscle increases from 3.78 to 4.43 in opposite homozygotes) without affecting fat content (18∶0+18∶1, intramuscular fat content, and backfat thickness). No mutations that could affect the functionality of the protein were found in the coding region. First, we proved in a purebred Duroc line that the C-T-A haplotype of the 3 single nucleotide polymorphisms (SNPs) (g.2108C>T; g.2228T>C; g.2281A>G) of the promoter region was additively associated to enhanced 18∶1/18∶0 both in muscle and subcutaneous fat, but not in liver. We show that this association was consistent over a 10-year period of overlapping generations and, in line with these results, that the C-T-A haplotype displayed greater SCD mRNA expression in muscle. The effect of this haplotype was validated both internally, by comparing opposite homozygote siblings, and externally, by using experimental Duroc-based crossbreds. Second, the g.2281A>G and the g.2108C>T SNPs were excluded as causative mutations using new and previously published data, restricting the causality to g.2228T>C SNP, the last source of genetic variation within the haplotype. This mutation is positioned in the core sequence of several putative transcription factor binding sites, so that there are several plausible mechanisms by which allele T enhances 18∶1/18∶0 and, consequently, the proportion of monounsaturated to saturated fat.

Estimating intramuscular fat content and fatty acid composition in live and post-mortem samples in pigs.

Repeated muscle sampling (n=732) on 216 pigs was performed to evaluate the effect of live sampling and sample size on intramuscular fat (IMF) content and fatty acid (FA) composition. The sampling scheme consisted of 1-3 biopsies of longissimus dorsi (LM), a small and a big post-mortem sample of LM, and a big post-mortem sample of gluteus medius (GM). IMF was determined by quantitative gas chromatography after direct transesterification. Data on LM were jointly analyzed using a mixed model on age with heterogeneous residual variances across sampling methods. Biopsies overestimated IMF and polyunsaturated FA content and underestimated monounsaturated FA content with decreasing sample size. Potential for bias and not sampling variance is the major limitation of using live samples for assessing changes in IMF with age. Small post-mortem samples of LM were as informative as big samples of GM for inferring IMF content but not IMF composition.

Genome-Wide Association Study Singles Out SCD and LEPR as the Two Main Loci Influencing Intramuscular Fat Content and Fatty Acid Composition in Duroc Pigs.

Intramuscular fat (IMF) content and fatty acid composition affect the organoleptic quality and nutritional value of pork. A genome-wide association study was performed on 138 Duroc pigs genotyped with a 60k SNP chip to detect biologically relevant genomic variants influencing fat content and composition. Despite the limited sample size, the genome-wide association study was powerful enough to detect the association between fatty acid composition and a known haplotypic variant in SCD (SSC14) and to reveal an association of IMF and fatty acid composition in the LEPR region (SSC6). The association of LEPR was later validated with an independent set of 853 pigs using a candidate quantitative trait nucleotide. The SCD gene is responsible for the biosynthesis of oleic acid (C18:1) from stearic acid. This locus affected the stearic to oleic desaturation index (C18:1/C18:0), C18:1, and saturated (SFA) and monounsaturated (MUFA) fatty acids content. These effects were consistently detected in gluteus medius, longissimus dorsi, and subcutaneous fat. The association of LEPR with fatty acid composition was detected only in muscle and was, at least in part, a consequence of its effect on IMF content, with increased IMF resulting in more SFA, less polyunsaturated fatty acids (PUFA), and greater SFA/PUFA ratio. Marker substitution effects estimated with a subset of 65 animals were used to predict the genomic estimated breeding values of 70 animals born 7 years later. Although predictions with the whole SNP chip information were in relatively high correlation with observed SFA, MUFA, and C18:1/C18:0 (0.48–0.60), IMF content and composition were in general better predicted by using only SNPs at the SCD and LEPR loci, in which case the correlation between predicted and observed values was in the range of 0.36 to 0.54 for all traits. Results indicate that markers in the SCD and LEPR genes can be useful to select for optimum fatty acid profiles of pork.

Genetic Marker Discovery in Complex Traits: A Field Example on Fat Content and Composition in Pigs

Among the large number of attributes that define pork quality, fat content and composition have attracted the attention of breeders in the recent years due to their interaction with human health and technological and sensorial properties of meat. In livestock species, fat accumulates in different depots following a temporal pattern that is also recognized in humans. Intramuscular fat deposition rate and fatty acid composition change with life. Despite indication that it might be possible to select for intramuscular fat without affecting other fat depots, to date only one depot-specific genetic marker (PCK1 c.2456C>A) has been reported. In contrast, identification of polymorphisms related to fat composition has been more successful. For instance, our group has described a variant in the stearoyl-coA desaturase (SCD) gene that improves the desaturation index of fat without affecting overall fatness or growth. Identification of mutations in candidate genes can be a tedious and costly process. Genome-wide association studies can help in narrowing down the number of candidate genes by highlighting those which contribute most to the genetic variation of the trait. Results from our group and others indicate that fat content and composition are highly polygenic and that very few genes explain more than 5% of the variance of the trait. Moreover, as the complexity of the genome emerges, the role of non-coding genes and regulatory elements cannot be disregarded. Prediction of breeding values from genomic data is discussed in comparison with conventional best linear predictors of breeding values. An example based on real data is given, and the implications in phenotype prediction are discussed in detail. The benefits and limitations of using large SNP sets versus a few very informative markers as predictors of genetic merit of breeding candidates are evaluated using field data as an example.

Response to selection for decreased backfat thickness at restrained intramuscular fat content in Duroc pigs

Intramuscular fat (IMF) content is a relevant trait for the pig industry and consumers. However, selection for IMF has the undesired correlated effect of decreasing lean growth. A selection experiment was performed to investigate the effects of selection against backfat thickness (BT) at restrained IMF. Barrows from a purebred Duroc line were allocated into a selected (n = 165) or a control (n = 185) group based on their litter predicted EBV. Litters in the selected group were selected against BT at 180 d at restrained IMF in gluteus medius (GM) whereas those in the control group were chosen randomly. Realized selection intensities and genetic responses for BT, IMF in GM, and BW were estimated using a 3-trait multivariate animal mixed model under a Bayesian setting. Correlated responses for other traits were estimated similarly but using a 4-trait model, where other traits were added to the previous 3-trait model 1 at a time. Selected pigs had less BT than control pigs [-1.22 mm, with highest posterior density interval at 95% (HPD95; -2.47, -0.75)] with restrained decrease in IMF, both in GM [-0.16%; HPD95 (-0.36, +0.05)] and in LM [-0.15%; HPD95 (-0.37, +0.09)]. However, the realized selection intensity for IMF in GM denotes that the restriction on IMF was incomplete [-0.18; HPD95 (-0.36, +0.02)]. Selection decreased BW [-1.64 kg; HPD95 (-2.47, -0.75)] but increased carcass lean weight [+0.66 kg; HPD95 (+0.14, +1.22)], indicating that the response in BT offsets the unfavorable correlated response in BW. Selected pigs were shorter [-0.50 cm; HPD95 (-0.81, -0.20)] but with similar ham weight and loin depth. These results provide evidence that lean weight can be improved restraining the genetic change in IMF. However, they also stress that a complete restriction on IMF is difficult to achieve unless selection is practiced on a big population where IMF is accurately predicted.

A polymorphism in the stearoyl-CoA desaturase gene promoter increases monounsaturated fatty acid content in dry-cured ham

Data on 125 dry-cured hams from purebred Duroc pigs were used to examine whether the favorable effect of the T allele in the promoter region of the stearoyl-CoA desaturase gene (AY487830:g.2228T>C) on monounsaturated fatty acid (MUFA) content in green ham is maintained after the curing process. It is shown that pigs carrying the T allele produced dry-cured hams with increased C16:1, C18:1n-9, C18:1n-7, and MUFA and decreased C18:0 and saturated fatty acid (SFA) content. The TT pigs had 1.39% more MUFA and 1.62% less SFA than the CC animals, while gilts had 0.74% more MUFA and 0.34% less SFA than barrows. The correlation between MUFA in green and dry-cured hams (n=53) was high (r=0.88), with TT pigs being more effective in retaining increased MUFA in green hams until the end of the curing period. It is concluded that increasing the presence of the T allele could have more impact than gender to produce hams with a high level of MUFA.

Hepatic lipogenic enzyme expression in pigs is affected by selection for decreased backfat thickness at constant intramuscular fat content

This study investigated (i) whether genetic selection for decreased backfat thickness at constant intramuscular fat (IMF) affects the expression of lipogenic enzymes in pig liver and (ii) whether expression of the hepatic lipogenic enzymes is related to subcutaneous fat and IMF fatty acid composition. The enzymes investigated were fatty acid synthase (FAS), stearoyl-CoA desaturase and Δ6-desaturase (Δ6d). Experiments were conducted on 30 barrows (15 controls and 15 selected). Selected pigs had lower backfat thickness, which was accompanied by a reduced expression of the hepatic FAS and Δ6d when compared to control pigs. There was a trend towards a positive relationship between FAS and Δ6d protein expression and saturated and polyunsaturated fatty acids content respectively, in subcutaneous fat but not in muscle. It was concluded that selection against backfat thickness is associated with changes in expression of the hepatic FAS and Δ6d proteins. The changes in protein expression did not influence significantly the tissue fatty acid composition under these conditions.

The effect of SCD and LEPR genetic polymorphisms on fat content and composition is maintained throughout fattening in Duroc pigs

The effects of the stearoyl-CoA desaturase (SCD; AY487830:g.2228T>C) and leptin receptor (LEPR; NM_001024587:g.1987C>T) polymorphisms on fat content and fatty acid (FA) composition were investigated throughout fattening. Samples of Longissimus thoracis (LT) and subcutaneous fat (SF) from 214 Duroc barrows were collected from 160days to slaughter age (220days) using a longitudinal design. Results indicated that the positive effect of the T allele at the SCD gene on monounsaturated FA and of the T allele at the LEPR gene on saturated FA are maintained throughout the growing-finishing period, both in LT and SF. In LEPR, however, compositional changes, particularly in SF, are a result of increased fatness. There is very limited evidence of genotype by age interaction, and thus it is concluded that the combined selection for the SCD T and LEPR C alleles is a good strategy to increase the MUFA/SFA ratio regardless of the age at slaughter.

Fast determination of oleic acid in pork by flow injection analysis/mass spectrometry

In some Mediterranean products such as olive oil or ham, oleic acid is the most abundant component of the total fat. Due to the large volume of trade in these products, it may be necessary to analyze oleic fatty acids in high numbers of samples in short periods of time. However, using classic lipid analysis techniques, it is not always possible to cope with these high demands. To solve this problem, a high-throughput analytical method for oleic fatty acid quantification in pork is presented. The purpose of the method is to avoid liquid chromatography processes using a flow injection analysis (FIA) system based on electrospray ionization mass spectrometry. The use of pentadecanoic fatty acid as an internal standard overcame matrix effects. The oleic FIA technique could be used as a suitable method for discriminating carcass samples for selection and labeling by oleic acid content when large numbers of pork samples must be processed in a short period of time.