Genome-wide association study identifies markers associated with carcass and meat quality traits in Italian Large White pigs.

A GWAS was performed using the genotypes obtained by PorcineSNP60 v2 BeadChip and 11 phenotypic traits (carcass lean meat percentage; backfat thickness; Longissimus thoracis muscle thickness; lightness; backfat thickness measured with caliper at the midline; meat pH measured at about 1 h post mortem and 24 h post mortem; CIE L*, a* and b* color parameters; and water-holding capacity). Three markers were associated with three of the phenotypic traits considered: M1GA0009592 (SSC7) with backfat thickness and lean meat content, DIAS0002910 (SSC6) and ALGA0109856 (SSC6) with water-holding capacity. The marker M1GA0009592, associated with backfat thickness, lies in a QTL region near the gene JARID2, which is a transcription factor also involved in the regulation of adipose-derived stem cell pluripotency. The results seem to indicate a possible role of these genomic regions in the regulation of pig carcass fatness (i.e. backfat at last rib) and water-holding capacity.

Genome-wide association study identifies markers associated with meat ultimate pH in Duroc pigs.

In this study we aimed to identify genomic regions associated with muscle pH, meat colour and water-holding capacity in a population of 280 Italian Duroc pigs genotyped by the Illumina PorcineSNP60 v2 Genotyping BeadChip. After quality control, the remaining 32 597 SNPs and 278 subjects were used to perform a genome-wide association study with the genabel package, using a kinship matrix in a model with the effects of sex, age and slaughter day. Bonferroni correction was applied, and the significant markers and regions were then further investigated to identify the nearest genes and the linkage disequilibrium (LD) between markers. Four markers (ASGA0082344, ASGA0095635, DBWU0000985 and CASI0005117) were significantly associated with ultimate pH (pHu ); no significant association was detected for the other traits. The four significant variants, located from 16.841 to 17.643 Mb on chromosome 3, were found within or close to the sequences of the sulfatase modifying factor 2 (SUMF2), lysine acetyltransferase 8 (KAT8), serine protease 8 (PRSS8) and phosphorylase kinase catalytic subunit gamma 2 (PHKG2) genes. The four associated markers lie in two LD blocks, suggesting that the observed effect is related to mutations located in two regions: the first one where SUMF2 is mapped and the second one where genes KAT8, PRSS8 and PHKG2 are located.

Identification of differentially expressed small RNAs and prediction of target genes in Italian Large White pigs with divergent backfat deposition.

The identification of the molecular mechanisms regulating pathways associated with the potential for fat deposition in pigs can lead to the detection of key genes and markers for the genetic improvement of fat traits. Interactions of microRNAs (miRNAs) with target RNAs regulate gene expression and modulate pathway activation in cells and tissues. In pigs, miRNA discovery is far from saturation, and the knowledge of miRNA expression in backfat tissue and particularly of the impact of miRNA variations is still fragmentary. Using RNA-seq, we characterized the small RNA (sRNA) expression profiles in Italian Large White pig backfat tissue. Comparing two groups of pigs divergent for backfat deposition, we detected 31 significant differentially expressed (DE) sRNAs: 14 up-regulated (including ssc-miR-132, ssc-miR-146b, ssc-miR-221-5p, ssc-miR-365-5p and the moRNA ssc-moR-21-5p) and 17 down-regulated (including ssc-miR-136, ssc-miR-195, ssc-miR-199a-5p and ssc-miR-335). To understand the biological impact of the observed miRNA expression variations, we used the expression correlation of DE miRNA target transcripts expressed in the same samples to define a regulatory network of 193 interactions between DE miRNAs and 40 DE target transcripts showing opposite expression profiles and being involved in specific pathways. Several miRNAs and mRNAs in the network were found to be expressed from backfat-related pig QTL. These results are informative for the complex mechanisms influencing fat traits, shed light on a new aspect of the genetic regulation of fat deposition in pigs and facilitate the prospective implementation of innovative strategies of pig genetic improvement based on genomic markers.

Transcriptional profiling of subcutaneous adipose tissue in Italian Large White pigs divergent for backfat thickness.

Fat deposition is a widely studied trait in pigs because of its implications with animal growth efficiency, technological and nutritional characteristics of meat products, but the global framework of the biological and molecular processes regulating fat deposition in pigs is still incomplete. This study describes the backfat tissue transcription profile in Italian Large White pigs and reports genes differentially expressed between fat and lean animals according to RNA-seq data. The backfat transcription profile was characterised by the expression of 23 483 genes, of which 54.1% were represented by known genes. Of 63 418 expressed transcripts, about 80% were non-previously annotated isoforms. By comparing the expression level of fat vs. lean pigs, we detected 86 robust differentially expressed transcripts, 72 more highly expressed (e.g. ACP5, BCL2A1, CCR1, CD163, CD1A, EGR2, ENPP1, GPNMB, INHBB, LYZ, MSR1, OLR1, PIK3AP1, PLIN2, SPP1, SLC11A1, STC1) and 14 lower expressed (e.g. ADSSL1, CDO1, DNAJB1, HSPA1A, HSPA1B, HSPA2, HSPB8, IGFBP5, OLFML3) in fat pigs. The main functional categories enriched in differentially expressed genes were immune system process, response to stimulus, cell activation and skeletal system development, for the overexpressed genes, and unfolded protein binding and stress response, for the underexpressed genes, which included five heat shock proteins. Adipose tissue alterations and impaired stress response are linked to inflammation and, in turn, to adipose tissue secretory activity, similar to what is observed in human obesity. Our results provide the opportunity to identify biomarkers of carcass fat traits to improve the pig production chain and to identify genetic factors that regulate the observed differential expression.

Genome-wide study on intramuscular fat in Italian Large White pig breed using the PorcineSNP60 BeadChip.

Genome-wide association study results are presented for intramuscular fat in Italian Large White pig breed. A total of 886 individuals were genotyped with PorcineSNP60 BeadChip. After quality control performed with plink software and in R environment, 49 208 markers remained for the association analysis. The genome-wide association studies was conducted using linear mixed model implemented in GenABEL. We detected seven new SNPs of genes till now not found associated to intramuscular fat (IMF). Three markers map in a wide intergenic region rich of QTL linked to fat traits, one map 388 kb upstream the gene SDK1, one map inside PPP3CA gene, one inside SCPEP1 gene and the last is not mapped in the porcine genome yet. Associations here presented indicate a moderate effect of these genes on IMF. In particular, PPP3CA, that is involved in the oxidative metabolism of skeletal muscle, could be considerated as an interesting candidate gene for IMF content in pigs. However, further studies are needed to clarify the role of these genes on the physiological processes involved in IMF regulation. These results may be useful to control this trait that is important in terms of nutritional, technological and organoleptic characteristics of fresh meat and processed products.

Relationship between perilipin genes polymorphisms and growth, carcass and meat quality traits in pigs.

The perilipins (PLIN) belong to a family of structural proteins that play a role regulating intracellular lipid storage and mobilization. Here, PLIN1 and PLIN2 have been evaluated as candidate genes for growth, carcass and meat quality traits in pigs. A sample of 607 Duroc pigs were genotyped for two single-nucleotide polymorphisms, one in intron 2 of the PLIN1 gene (JN860199:g.173G>A) and the other at the 3' untranslated region of the PLIN2 gene (GU461317:g.98G>A). Using a Bayesian approach, we have been able to find evidence of additive, dominant and epistatic associations of the PLIN1 and PLIN2 polymorphisms with early growth rate and carcass length. However, the major effects were produced by the dominant A allele at the PLIN2 polymorphism, which also affected the carcass lean weight. Thus, pigs carrying an additional copy of the A allele at the g.98G>A PLIN2 polymorphism had a probability of at least 98% of producing carcasses with heavier lean weight (+0.41 kg) and ham weight (+0.10 kg). The results obtained indicate that the PLIN2 polymorphism could be a useful marker for lean growth. In particular, it may help to reduce the undesired negative correlated response in lean weight to selection for increased intramuscular fat content, a common scenario in some Duroc lines involved in the production of high quality pork products.

Application of a non-invasive tool and identification of genetic markers in swine to enhance ham quality: a preliminary study.

A genome-wide association study (GWAS) was performed as a preliminary step to identify regions potentially related to ham quality traits. In this research, genomic information was obtained from 238 commercial hybrid pigs utilising the GeneSeek® Genomic Profiler genome-wide porcine genotyping array. Carcasses were tested for hot weight, the thickness of backfat and loin, and lean meat percentage. The corresponding fresh hams were assayed for weight and ultimate pH; the activities of Cathepsin B and Ferrochelatase of Semimembranosus muscle were determined through fluorimetric methods. The lean meat percentage of fresh ham (LMPH), salt absorbed after first (SALT1) and overall salting stages (SALT) were estimated online by the Ham Inspector™ apparatus. Hams were processed in compliance with the procedures established for Protected Designation of Origin Parma ham, and ham weight losses were measured at the main processing stages. Hot carcass weights showed a significant negative correlation with their lean meat percentage and LMPH, while LMPH was correlated positively with carcass lean meat, SALT1, SALT, and weight losses. The GWAS detected genome-wide association for 12 single nucleotide polymorphisms with Ferrochelatase activity. The results obtained in this preliminary study were achieved by combining innovative and non-destructive technologies for screening hams under processing, measures of enzymatic muscle properties relevant to dry-cured ham quality, and genomic information obtained through a GWAS. Additional studies carried out in a larger number of pigs have been planned to investigate the effect of gene variants of Ferrochelatase activity in the dry-cured ham's quality with main reference to colour development and to confirm the GWAS results obtained in this study.

Polymorphisms of beta-lactoglobulin promoter region in three Sicilian goat breeds.

Several beta-lactoglobulin (BLG) polymorphisms have been described within the proximal promoter region and coding region of the caprine gene, although no genetic variants affecting the protein amino acid composition and/or expression level have been characterized so far. Binding sites for several transcription factors (TFs) are present in the BLG promoter region. The aims of this work were to sequence the full-length promoter region of three Sicilian goat breeds in order to identify polymorphisms, analyze the identified haplotypes, search for differences between breeds for the presence of polymorphisms in this gene region, search for putative TFs binding sites, and check if polymorphisms lay within the identified TFs binding sites. The promoter region of BLG gene in Sicilian goat breeds showed high level of polymorphism due to the presence of 36 single nucleotide polymorphisms (SNPs). Association between polymorphic sites was computed within the whole sample analyzed and 18 haplotypes were inferred. Binding sites for three milk protein binding factors (MPBFs) and four nuclear factor-I (NF-I) were found within BLG promoter region based on the ovine sequence. The identification of some SNPs within TFs binding sites allowed hypothesizing the loss of TFs. Further studies are in progress to evaluate the effect of these mutations on binding affinity of TFs, the functional interaction of the TFs with the goat BLG promoter, and the relationship of the polymorphisms with BLG gene expression and milk production and composition.

A whole genome scan for QTL affecting milk protein percentage in Italian Holstein cattle, applying selective milk DNA pooling and multiple marker mapping in a daughter design.

We report on a complete genome scan for quantitative trait loci (QTL) affecting milk protein percentage (PP) in the Italian Holstein-Friesian cattle population, applying a selective DNA pooling strategy in a daughter design. Ten Holstein-Friesian sires were chosen, and for each sire, about 200 daughters, each from the high and low tails of estimated breeding value for PP, were used to construct milk DNA pools. Sires and pools were genotyped for 181 dinucleotide microsatellites covering all cattle autosomes. Sire marker allele frequencies in the pools were obtained by shadow correction of peak height in the electropherograms. After quality control, pool data from eight sires were used for all subsequent analyses. The QTL heterozygosity estimate was lower than that of similar studies in other cattle populations. Multiple marker mapping identified 19 QTL located on 14 chromosomes (BTA1, 2, 3, 4, 5, 6, 8, 9, 12, 14, 17, 20, 23 and 27). The sires were also genotyped for seven polymorphic sites in six candidate genes (ABCG2, SPP1, casein kappa, DGAT1, GHR and PRLR) located within QTL regions of BTA6, 14 and 20 found in this study. The results confirmed or excluded the involvement of some of the analysed markers as the causative polymorphic sites of the identified QTL. The QTL identified, combined with genotype data of these candidate genes, will help to identify other quantitative trait genes and clarify the complex QTL patterns observed for a few chromosomes. Overall, the results are consistent with the Italian Holstein population having been under long-term selection for high PP.

Expression profiling of functional genes in prenatal skeletal muscle tissue in Duroc and Pietrain pigs.

In livestock, skeletal muscle is a tissue of major economic importance for meat production and muscle mass is largely determined during the prenatal period by the number and the size of muscle fibres. The understanding of gene expression changes during prenatal pig muscle development is still limited. In this study, genes identified as differentially expressed in a previous microarray research and chosen for the function of the coded protein as putative candidate involved in myogenesis were considered to analyse their expression profile during foetal growth of Duroc and Pietrain pigs. The eleven genes were considered by real-time PCR for a time-course evaluation of the transcription level at six stages of prenatal longissimus dorsi development. The results suggest that the most relevant variations in mRNA levels of the analysed genes seem to follow temporal waves of gene expression. Significant changes of transcription were observed at 21-35 and 63-91 days, the two main phases of skeletal muscle development. During the early phases of Pietrain embryos' development, 10 of the 11 genes showed an induction. In Duroc embryos, a second phase of gene up-regulation can be identified in the phase 63-77 days. These results provide new data on developmental changes of expression profile of 11 genes involved in different functional pathways related to prenatal myogenic processes in Duroc and Pietrain pigs.

A critical review of the mechanisms involved in the occurrence of growth-related abnormalities affecting broiler chicken breast muscles.

In the past decade, the poultry industry has faced the occurrence of growth-related muscular abnormalities that mainly affect, with a high incidence rate, the Pectoralis major of the fast-growing genotypes selected for their production performances (high growth rate and breast yield). These myopathies are termed as White Striping, Wooden Breast, and Spaghetti Meat and exhibit distinctive phenotypes. A spatiotemporal distribution has been demonstrated for these disorders as in the early stage they primarily affect the superficial area in the cranial portion of the muscle and, as the birds grow older, involve the entire tissue. Aside from their distinctive phenotypes, these myopathies share common histological features. Thus, it might be speculated that common causative mechanisms might be responsible for the physiological and structural perturbations in the muscle associated with these conditions and might underpin their occurrence. The present review paper aims to represent a critical survey of the outcomes of all the histologic and ultrastructural observations carried out on White Striping, Wooden Breast, and Spaghetti Meat affected muscles. Our analysis has been performed by combining these outcomes with the findings of the genetic studies, trying to identify possible initial causative mechanisms triggering the onset and the time-series of the events ultimately resulting in the development and progression of the growth-related myopathies currently affecting broilers Pectoralis major muscles. Several evidences support the hypothesis that sarcoplasmic reticulum stress, primarily induced an accumulation of misfolded proteins (but also driven by other factors including altered calcium homeostasis and accumulation of fatty acids), may be responsible for the onset of these growth-related myopathies in broilers. At the same time, the development of hypoxic conditions, as a direct consequence of an inadequate vascularization, triggers a time-series sequence of events (i.e., phlebitis, oxidative stress, etc.) resulting in the activation of response mechanisms (i.e., modifications in the energetic metabolism, inflammation, degeneration, and regeneration) which are all strictly related to the progression of these myopathic disorders.

Genome-wide association study identifies quantitative trait loci regions involved in muscle acidic profile in Large White heavy pigs.

The widespread use of genome-wide association studies resulted in the discovery of genomic regions associated with fatty acid (FA) composition in different porcine tissues, but little information exists about the genes involved in FA composition of meat obtained from heavy pigs selected for the production of Italian dry-cured hams. To this objective, we genotyped with a single nucleotide polymorphism (SNP) panel 795 Italian Large White heavy pigs to identify the markers and genomic regions associated with Semimembranosus muscle FA profile. Heritability estimates for intramuscular fat FA profile were of low-to-moderate magnitude, suggesting that these traits may be improved with genomic selection. On the whole, 45 SNPs were significantly associated with 14 FAs, and 4 of them (ALGA008109, ALGA0081097, CASI0010164 and SIRI0000267) were associated with more than 1 FA. The palmitoleic : palmitic and oleic : stearic ratios displayed the highest number of significant markers and the most significant associations (Bonferroni adjusted P < 5.00E-07). Of particular interest, the palmitoleic : palmitic ratio was strongly associated with markers located at 111 to 114 Mb on chromosome 14, in the same chromosomal region where Stearoyl-CoA desaturase Δ9 (SCD) gene is located. Several significant chromosomal regions were found; some of them harbour key genes playing pivotal roles in FA desaturation and elongation, such as SCD and some members of the Elongation of Very Long-Chain FA (ELOVL) gene family. The results suggest that the identification of causal mutations in these regions may provide a set of markers useful for selection schemes aimed at improving FA composition in pork products.

Copy number variation and missense mutations of the agouti signaling protein (ASIP) gene in goat breeds with different coat colors.

In goats, classical genetic studies reported a large number of alleles at the Agouti locus with effects on coat color and pattern distribution. From these early studies, the dominant A(Wt) (white/tan) allele was suggested to cause the white color of the Saanen breed. Here, we sequenced the coding region of the goat ASIP gene in 6 goat breeds (Girgentana, Maltese, Derivata di Siria, Murciano-Granadina, Camosciata delle Alpi, and Saanen), with different coat colors and patterns. Five single nucleotide polymorphisms (SNPs) were identified, 3 of which caused missense mutations in conserved positions of the cysteine-rich carboxy-terminal domain of the protein (p.Ala96Gly, p.Cys126Gly, and p.Val128Gly). Allele and genotype frequencies suggested that these mutations are not associated or not completely associated with coat color in the investigated goat breeds. Moreover, genotyping and sequencing results, deviation from Hardy-Weinberg equilibrium, as well as allele copy number evaluation from semiquantitative fluorescent multiplex PCR, indicated the presence of copy number variation (CNV) in all investigated breeds. To confirm the presence of CNV and evaluate its extension, we applied a bovine-goat cross-species array comparative genome hybridization (aCGH) experiment using a custom tiling array based on bovine chromosome 13. aCGH results obtained for 8 goat DNA samples confirmed the presence of CNV affecting a region of less that 100 kb including the ASIP and AHCY genes. In Girgentana and Saanen breeds, this CNV might cause the A(Wt) allele, as already suggested for a similar structural mutation in sheep affecting the ASIP and AHCY genes, providing evidence for a recurrent interspecies CNV. However, other mechanisms may also be involved in determining coat color in these 2 breeds.

The porcine fat mass and obesity associated (FTO) gene is associated with fat deposition in Italian Duroc pigs.

In humans, common variants in the fat mass and obesity associated (FTO) gene are associated with body mass index and obesity. Here we sequenced exon 4, parts of introns 3 and 4 and two portions of the 3'-untranslated region of the porcine FTO gene in a panel of nine pigs of different breeds and identified three SNPs. Allele frequencies of the g.276T>G (AM931150) mutation were studied in seven pig breeds. This mutation was used to linkage-map FTO to SSC6. Association analyses between the g.276T>G polymorphism and several traits [pH of semimembranosus muscle and estimated breeding values (EBV) for average daily gain, back fat thickness, lean cuts, ham weight and feed:gain ratio] were carried out in 257 sib-tested Italian Large White pigs. Only feed:gain ratio showed P<0.05. A selective genotyping approach was applied, analysing two extreme and divergent groups of Italian Large White pigs selected on the basis of back fat thickness EBV (50 with most positive and 50 with most negative values). Fisher's exact test (two-tailed) was not significant when comparing the allele frequencies of these two groups. The same approach was used in the Italian Duroc breed for which two extreme and divergent groups of animals were selected according to visible intermuscular fat EBV. Differences of allele frequencies between these two groups were highly significant (P<0.00001, P<0.001 and P<0.0001, considering all animals or only two- or three-generation unrelated animals respectively), indicating association between the analysed FTO marker and intermuscular fat deposition.

Toll-like receptor 4 genetic diversity among pig populations.

The transmembrane glycoprotein encoded by the Toll-like receptor 4 gene (TLR4) acts as the transducing subunit of the lipopolysaccharide receptor complex of mammals, which is a major sensor of infections by Gram-negative bacteria. As variation in TLR4 may alter host immune response to lipopolysaccharide, the association between TLR4 polymorphisms and immune traits of the respiratory and gut systems has important implications for livestock. Here, a sequence dataset from 259 animals belonging to commercial and traditional European pig populations, consisting of 4305 bp of TLR4, including the full transcribed region, a portion of intron 2 and the putative promoter region, was used to explore genetic variation segregating at the TLR4 locus. We identified 34 single nucleotide polymorphisms, 17 in the coding sequence and 17 in the non-coding region. Five non-synonymous mutations clustered within, or in close proximity to, the hypervariable domain of exon 3. In agreement with studies in other mammals, a major exon 3 haplotype segregated at high frequency in the whole sample of 259 pigs, while variants carrying non-synonymous substitutions showed frequencies ranging between 0.6% and 8.7%. Although results on exon 3 provided suggestive evidence for purifying selection occurring at the porcine TLR4 gene, the analysis of both coding and non-coding regions highlighted the fact that demographic factors strongly influence the tests of departure from neutrality. The phylogenetic analysis of TLR4 identified three clusters of variation (ancestral, Asian, European), supporting the evidence of Asian introgression in European main breeds and the well documented history of pig breed domestication previously identified by mtDNA analysis.

Genetic parameters of backfat fatty acids and carcass traits in Large White pigs.

Subcutaneous fat thickness and fatty acid composition (FAC) play an important role on seasoning loss and organoleptic characteristics of seasoned hams. Dry-cured ham industry prefers meats with low contents of polyunsaturated fatty acids (PUFA) because these negatively affect fat firmness and ham quality, whereas consumers require higher contents in those fatty acids (FA) for their positive effect on human health. A population of 950 Italian Large White pigs from the Italian National Sib Test Selection Programme was investigated with the aim to estimate heritabilities, genetic and phenotypic correlations of backfat FAC, Semimembranosus muscle intramuscular fat (IMF) content and other carcass traits. The pigs were reared in controlled environmental condition at the same central testing station and were slaughtered at reaching 150 kg live weight. Backfat samples were collected to analyze FAC by gas chromatography. Carcass traits showed heritability levels from 0.087 for estimated carcass lean percentage to 0.361 for hot carcass weight. Heritability values of FA classes were low-to-moderate, all in the range 0.245 for n-3 PUFA to 0.264 for monounsaturated FA (MUFA). Polyunsaturated fatty acids showed a significant genetic correlation with loin thickness (0.128), backfat thickness (-0.124 for backfat measured by Fat-O-Meat'er and -0.175 for backfat measured by calibre) and IMF (-0.102). Obviously, C18:2(n-6) shows similar genetic correlations with the same traits (0.211 with loin thickness, -0.206 with backfat measured by Fat-O-Meat'er, -0.291 with backfat measured by calibre and -0.171 with IMF). Monounsaturated FA, except with the backfat measured by calibre (0.068; P<0.01), do not show genetic correlations with carcass characteristics, whereas a negative genetic correlation was found between MUFA and saturated FA (SFA; -0.339; P<0.001). These results suggest that MUFA/SFA ratio could be increased without interfering with carcass traits. The level of genetic correlations between FA and carcass traits should be taken into account in dealing with the development of selection schemes addressed to modify carcass composition and/or backfat FAC.

Ancient vs. recent processes as factors shaping the genetic variation of the European wild boar: are the effects of the last glaciation still detectable?

The European wild boar is an important game species, subjected to local extinctions and translocations in the past, and currently enormously and worryingly expanding in some areas where management is urgently required. Understanding the relative roles of ancient and recent events in shaping the genetic structure of this species is therefore not only an interesting scientific issue, but it represents also the basis for addressing future management strategies. In addition, several pig breeds descend from the European wild boar, but the geographical location of the domestication area(s) and the possible introgression of pig genomes into wild populations are still open questions. Here, we analysed the genetic variation in different wild boar populations in Europe. Ten polymorphic microsatellites were typed in 252 wild boars and the mtDNA control region was sequenced in a subset of 145 individuals. Some samples from different pig breeds were also analysed. Our results, which were obtained considering also 612 published mtDNA sequences, suggest that (i) most populations are similarly differentiated, but the major discontinuity is found along the Alps; (ii) except for the Italian populations, European wild boars show the signature of a postglacial demographic expansion; (iii) Italian populations seem to preserve a high proportion of preglaciation diversity; (iv) the demographic decline which occurred in some areas in the last few centuries did not produce a noticeable reduction of genetic variation; (v) signs of human-mediated gene flow among populations are weak, although in some regions the effects of translocations are detectable and a low degree of pig introgression can be identified; (vi) the hypothesis of an independent domestication centre in Italy is not supported by our data, which in turn confirm that Central European wild boar might have represented an important source for domestic breeds. We can therefore conclude that recent human activities had a limited effect on the wild boar genetic structure. It follows that areas with high variation and differentiation represent natural reservoirs of genetic diversity to be protected avoiding translocations. In this context controlling some populations by hunting is not expected to affect significantly genetic variation in this species.

Investigation of candidate genes for glycolytic potential of porcine skeletal muscle: Association with meat quality and production traits in Italian Large White pigs.

The objective of this study was to investigate the association of DNA markers in candidate genes for glycolytic potential on meat quality parameters (pH(1), pH(u), glycogen and lactate content and glycolytic potential of semimembranosus muscle) and estimated breeding values (EBVs) for average daily gain, lean cuts, back fat thickness, ham weight, and feed:gain ratio in 272 Italian Large White pigs. Three mutations in the PRKAG3 gene (T30N, G52S and I199V) were investigated as well as single nucleotide polymorphisms in two other skeletal muscle genes (PGAM2 and PKM2) involved in the glycolytic pathway. Association analysis with the PRKAG3 markers showed significant results (P<0.05) only for pH(1) (I199V, with significant additive effect) and lactate content (T30N), confirming, at least in part, the effects of this gene on meat quality traits. Significant association (P<0.05) was also observed for PGAM2 and ham weight EBV with significant additive and dominance effects. PKM2 was associated with average daily gain, lean cuts (P<0.001), back fat thickness and feed:gain ratio (P<0.05), with significant additive and/or dominance effects on these traits. PKM2 encodes for a key enzyme of the muscle glycolytic pathway and maps on porcine chromosome 7 where other studies have reported important QTL for the same traits. These data might suggest an important function of this gene in the mechanisms that produce the observed effects. The results will be important to evaluate the inclusion of some of these DNA polymorphisms in marker assisted selection programs.

Investigation of the Perilipin 5 gene expression and association study of its sequence polymorphism with meat and carcass quality traits in different pig breeds.

The growing request for healthier fatty acid composition of animal products is raising the necessity of a deeper knowledge of the main factors controlling fatty acids storage in muscle and backfat. Perilipin (PLIN) 5, and the whole Perilipin family, seem to play a crucial role in the regulation of lipids deposition as code for proteins coating intracellular lipid droplets surface. Nevertheless, the knowledge of these genes in pig is still incomplete. The present research was aimed at investigating in different pig breeds the PLIN5 gene, analysing its expression level and the associations of the variability in its downstream gene region with pork meat and carcass quality traits. Moreover, the PLIN5 protein localisation in porcine skeletal muscle was investigated through immunofluorescence, resulting to be widespread in Semimembranosus muscle (SM) myofibers. The identified single nucleotide polymorphism (SNP) rs327694326 (NC_010444.4:g.74314701T>C) located in PLIN5 downstream region was analysed in different pig populations, represented by 512 Italian Large White (ILW) pigs, 300 Italian Duroc (IDU) samples, 100 Italian Landrace, 100 Pietrain and 60 pigs belonging to three Italian native breeds (20 samples of Cinta Senese, 20 Calabrese and 20 Casertana pigs). The C allele was found in ILW, IDU and Pietrain pigs. In ILW pigs this SNP showed results indicating a possible association with oleic and cis-vaccenic fatty acid contents in backfat tissue. Furthermore, as PLINs are known to regulate lipases activity, we tested if the rs327694326 SNP was associated with differences in Hormone-sensitive lipase (LIPE) gene expression levels. In SM of ILW pigs, PLIN5 C allele was associated with significantly lower LIPE transcription levels than T allele (P=0.02 for Student's t test of TT v. CT samples, P<0.0001 for TT v. CC pigs), whereas in IDU breed no significant difference was noticed in LIPE transcription levels between TT and TC animals. The results may suggest that variations in the PLIN5 sequence may be linked to LIPE expression through a still poorly known regulative molecular process.

Association study highlights the influence of ELOVL fatty acid elongase 6 gene region on backfat fatty acid composition in Large White pig breed.

Dietary fatty acid (FA) composition has an impact on human health. There is an increasing request from consumers for healthier food and pork industry must respond to it without worsening performance and the technological properties of pork products. The inclusion of genetic markers for carcass FA composition in pig selection schemes could be a useful tool to reach the right balance between unsaturated and saturated FAs to satisfy market demands. With the aim of finding genomic regions associated with porcine backfat FA composition, a genome-wide association study was performed on 798 Italian Large White pigs genotyped using Illumina PorcineSNP60 k. The strongest associations with backfat contents of palmitic, palmitoleic, oleic, medium-chain and long-chain FAs were found for the Sus scrofa chromosome (SSC) 8 region located at 119 to 122 Mb, where the gene ELOVL FA elongase 6 is mapped. Palmitic, palmitoleic, stearic and oleic acid contents were also found associated with SSC14, in particular with the genomic region at 121 to 124 Mb, where stearoyl-CoA desaturase Δ9 gene lies. On the other hand, the genomic regions associated with backfat contents of arachidic, arachidonic, n-6 and n-3 FAs showed to harbour mainly genes involved in dietary lipids and carbohydrates digestion, absorption and utilisation. To our knowledge, this is the first study performed in Large White pigs identifying markers and genomic regions associated with backfat FA composition. The results validate in Large White some associations previously detected in other pig breeds and indicate the involvement of distinct metabolic pathways in the deposition pattern of essential and non-essential FAs.