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Several genetic markers, previously associated with meat quality traits, have been proposed to be included in Iberian pig breeding programs. However, before being implemented, effects of these candidate SNPs on premium cuts' yield should be evaluated to avoid potential undesirable antagonistic effects. Therefore, the main goals of this study were to evaluate the effects of a set of 26 polymorphisms on premium cuts weights and to corroborate their effects on meat quality in a larger population. Phenotypic data of approximately 1550 Iberian pigs were recorded. The PRKAG3_rs319678464C and PRKAG3_rs1108399077G alleles were associated with an increase of shear force, water losses and color values and a reduction of average daily gain (ADG). The CAST_rs196949783G > A and ADIPOQ_rs3476515794T > G SNPs mainly affected IMF content, with this last SNP being the only one with significant effects on any of the carcass measures, specifically Longissimus thoracis et lumborum (LTL) weight. The ELOVL6_rs3473714672A, FASN_rs331694510A, MTTP_rs335896411C and ACACA_rs340781986C alleles were linked to a higher percentage of oleic acid and monounsaturated FA and a decrease in palmitic, palmitoleic and saturated FA. Besides, suggestive effects were observed between ELOVL6_rs3473714672A > G and ham and shoulder weights, and between MTTP_rs335896411T > C and LTL muscle weight. Finally, the NR6A1_rs326780270T allele was associated with a significant increase in ADG and tended to reduce the ham weight. The SNPs mapped to PRKAG3 (rs319678464G > C), ACACA, FASN and CAST genes could be implemented in breeding programs to improve meat quality traits without undesirable effects on carcass composition. The SNPs mapped to PRKAG3 (rs1108399077G > A) ELOVL6, MTTP and NR6A1 should continue to be tested in a larger number of animals.

Growth is dependent on genotype and diet, even at early developmental stages. In this study, we investigated the effects of genotype, sex, and body weight on the fetal muscle transcriptome of purebred Iberian and crossbred Iberian x Large White pigs sharing the same uterine environment. RNA sequencing was performed on 16 purebred and crossbred fetuses with high body weight (340±14g and 415±14g, respectively) and 16 with low body weight (246±14g and 311±14g, respectively), on gestational day 77. Genotype had the greatest effect on gene expression, with 645 genes identified as differentially expressed (DE) between purebred and crossbred animals. Functional analysis showed differential regulation of pathways involved in energy and lipid metabolism, muscle development, and tissue disorders. In purebred animals, fetal body weight was associated with 35 DE genes involved in development, lipid metabolism and adipogenesis. In crossbred animals, fetal body weight was associated with 60 DE genes involved in muscle development, viability, and immunity. Interestingly, the results suggested an interaction genotype∗weight for some DE genes. Fetal sex had only a modest effect on gene expression. This study allowed the identification of genes, metabolic pathways, biological functions and regulators related to fetal genotype, weight and sex, in animals sharing the same uterine environment. Our findings contribute to a better understanding of the molecular events that influence prenatal muscle development and highlight the complex interactions affecting transcriptional regulation during development., MINECO, FEDER, Depto. de Producción Animal, Fac. de Veterinaria, TRUE, pub

The Iberian pig breed includes several well-differentiated strains. The present study evaluated carcass traits, fat deposition and muscle expression of important lipogenic genes (SCD, ME1, ACACA, FASN, EGR1, ACOX and ACLY) using 65 male pigs of 3 Iberian strains (20 Lampiño, 23 Torbiscal, and 22 Retinto) finished either in a conventional, concentrate-based system (CF) or in montanera (MF), a traditional free-range system with acorn feeding. Torbiscal had the highest ham, Longissimus thoracis and prime cuts yields, and the thinnest subcutaneous adipose tissue (SAT). Retinto had the highest monounsaturated fatty acids (MUFA) and percentage of intramuscular fat (IMF), while Lampiño had the greatest content of saturated fatty acids (SFA), polyunsaturated fatty acids (PUFA), atherogenic (AI) and thrombogenic (TI) indexes in SAT. Conventionally finished pigs had the highest ham, L. thoracis and prime cuts yields, and SFA. Montanera-finished animals had the highest PUFA and MUFA contents, and the lowest AI, TI and n6/n3 ratio in SAT. In relation to gene expression, Retinto had the greatest SCD, FASN and ACLY levels. Most studied genes were overexpressed in CF pigs. In conclusion, MF pigs had healthier fat than CF pigs, and Retinto had the healthiest fat and the greatest lipogenic trend in muscle, supported by IMF and lipogenic gene expression.

An adequate intestinal environment before weaning may contribute to diarrhea predisposition and piglet development. This study evaluates how the dietary supplementation of vitamin E (VE) (100 mg/kg), hydroxytyrosol (HXT) (1.5 mg/kg) or the combined administration (VE + HXT) given to Iberian sows from gestation affects the piglet's faecal characteristics, short chain fatty acids (SCFAs), fatty acid profile or intestinal morphology as indicators of gut health; and quantify the contribution of the oxidative status and colostrum/milk composition to the piglet's SCFAs content and intestinal health. Dietary VE increased isobutyric acid (iC4), butyric acid (C4), isovaleric acid (iC5), and ∑SCFAs, whereas HXT increased iC4 and tended to decrease ∑SCFAs of faeces. Piglets from HXT-supplemented sows also tended to have higher faecal C20:4n-6/C20:2 ratio C22:6 proportion and showed lower occludin gene expression in the duodenum. The combination of both antioxidants had a positive effect on iC4 and iC5 levels. Correlation analyses and regression equations indicate that faecal SCFAs were related to oxidative status (mainly plasma VE) and colostrum and milk composition (mainly C20:2, C20:3, C20:4 n-6). This study would confirm the superiority of VE over HXT supplementation to improve intestinal homeostasis, gut health, and, consequently piglet growth.

The hypothalamus is implicated in controlling feeding and adiposity, besides many other physiological functions, and thus can be of great importance in explaining productive differences between lean and fatty pig breeds. The present study aimed to evaluate the hypothalamic transcriptome of pure Iberian (IBxIB) and Large White x Iberian crossbreds (IBxLW) at 60 days-old, produced in a single maternal environment. Results showed the implication of gender and genotype in the hypothalamic transcriptome, with 51 differentially expressed genes (DEGs) between genotypes and 10 DEGs between genders. Fourteen genotype by sex interactions were found, due to a higher genotype effect on transcriptome found in males. In fact, just 31 DEGs were identified when using only females but 158 using only males. A higher expression of genes related to mitochondrial activity in IBxIB male animals (ND3, ND4, ND5, UQCRC2 and ATP6) was found, which was related to a higher oxidative phosphorylation and greater reactive oxygen species and nitric oxide production. IBxLW male animals showed higher expression of SIRT3 regulator, also related to mitochondrial function. When females were analysed, such differences were not found, since only some differences in genes related to the tricarboxylic acid cycle. Thus, the results indicate a significant effect and interaction of the breed and the sex on the hypothalamic transcriptome at this early age.

Iberian pigs and its crosses are produced to obtain high-quality meat products. The objective of this work was to evaluate a wide panel of DNA markers, selected by biological and functional criteria, for association with traits related to muscle growth, fatness, meat quality and metabolism. We used 18 crossbred Iberian pigs with divergent postnatal growth patterns for whole genome sequencing and SNP discovery, with over 13 million variants being detected. We selected 1023 missense SNPs located on annotated genes and showing different allele frequencies between pigs with makerdly different growth patterns. We complemented this panel with 192 candidate SNPs obtained from literature mining and from muscle RNAseq data. The selected markers were genotyped in 480 Iberian × Duroc pigs from a commercial population, in which phenotypes were obtained, and an association study was performed for the 1005 successfully genotyped SNPs showing segregation. The results confirmed the effects of several known SNPs in candidate genes (such as LEPR, ACACA, FTO, LIPE or SCD on fatness, growth and fatty acid composition) and also disclosed interesting effects of new SNPs in less known genes such as LRIG3, DENND1B, SOWAHB, EPHX1 or NFE2L2 affecting body weight, average daily gain and adiposity at different ages, or KRT10, NLE1, KCNH2 or AHNAK affecting fatness and FA composition. The results provide a valuable basis for future implementation of marker-assisted selection strategies in swine and contribute to a better understanding of the genetic architecture of relevant traits.

Meat color is the first perceived sensory feature and one of the most important quality traits. Myoglobin is the main pigment in meat, giving meat its characteristic cherry-red color, highly appreciated by the consumers. In the current study, we used the RNA-seq technique to characterize the longissimus dorsi muscle transcriptome in two groups of Iberian pigs with divergent breeding values for myoglobin content. As a result, we identified 57 differentially expressed genes and transcripts (DEGs). Moreover, we have validated the RNA-seq expression of a set of genes by quantitative PCR (qPCR). Functional analyses revealed an enrichment of DEGs in biological processes related to oxidation (HBA1), lipid metabolism (ECH1, PLA2G10, PLD2), inflammation (CHST1, CD209, PLA2G10), and immune system (CD209, MX2, LGALS3, LGALS9). The upstream analysis showed a total of five transcriptional regulatory factors and eight master regulators that could moderate the expression of some DEGs, highlighting SPI1 and MAPK1, since they regulate the expression of DEGs involved in immune defense and inflammatory processes. Iberian pigs with high myoglobin content also showed higher expression of the HBA1 gene and both molecules, myoglobin and hemoglobin, have been described as having a protective effect against oxidative and inflammatory processes. Therefore, the HBA1 gene is a very promising candidate gene to harbor polymorphisms underlying myoglobin content, whereby further studies should be carried out for its potential use in an Iberian pig selection program.

[ES] Actualmente existe una emergente preocupación por el aumento de bacterias con resistencia a antibióticos. La introducción de las mismas en la cadena trófica humana resulta alarmante en el ámbito sanitario, por su potencial patogénico y su dificultad de tratamiento con los antibióticos disponibles. Por ello, desde las principales organizaciones internacionales se promueve la vigilancia de las bacterias y genes resistentes a antibióticos tanto en aguas, como en productos vegetales o cárnicos de consumo humano. En concreto, la detección de microorganismos resistentes a antibióticos en vegetales resulta relevante al darse habitualmente su consumo en crudo sin un lavado y/o cocción previos. Dentro de los microorganismos patógenos se consideran prioritarios por la Organización Mundial de la Salud (OMS) los gram negativos resistentes a carbapenémicos al no presentarse alternativas eficaces de tratamiento y estar relacionados con aumentos de la mortalidad por enfermedades infecciosas. En el presente trabajo se analizaron muestras de lechuga, espinaca, col lombarda, col blanca y fresa procedentes de cultivo convencional no ecológico y adquiridos en diferentes comercios minoristas de la ciudad de Valencia. El objetivo de estudio se enfocaba en la detección e identificación de bacterias resistentes a antibióticos y la determinación de los genes que les confieren dicha resistencia. Se aislaron un total de 239 cepas, siguiendo el estudio con 28 de ellas por presentar características de interés (oxidasa negativo, catalasa positivo y gram negativas). A partir de estas, se determinó su sensibilidad a distintos antimicrobianos mediante el método antibiograma disco-placa, confirmándose así sus resistencias tanto intrínsecas como adquiridas a diversos antibióticos resultando todas ellas resistentes a dos o más antibióticos beta-lactámicos. Se identificaron mediante pruebas bioquímicas destacando la detección de cepas de Stenotrophomonas maltophilia y Acinetobacter spp. considerad, [EN] Currently, there is an emerging concern about the increase of antibiotic-resistant bacteria. The introduction of these bacteria into the human food chain is alarming in the healthcare field due to their pathogenic potential and the difficulty in treating them with available antibiotics. For this reason, the main international organizations promote surveillance of antibiotic-resistant bacteria (ARB) and antibiotic-resistant genes (ARG) in water sources, vegetable and meat products consumed by humans. In particular, the detection of antibiotic-resistant microorganisms in vegetables is relevant because we usually consumed them raw without prior washing and/or cooking. Among pathogenic microorganisms, carbapenem-resistant gram-negative microorganisms are considered priority by the World Health Organization (WHO) because of the lack of effective treatment alternatives and because they are associated with the increase in mortality from infectious diseases. In the present study, samples of lettuce, spinach, red cabbage, white cabbage and strawberry from conventional non-organic farming purchased in different retail stores in the city of Valencia were analyzed. The objective of the study focused on the detection and identification of antibiotic-resistant bacteria and the determination of the genes that confer them such resistance. A total of 239 strains were isolated, and further analysis was conducted on 28 of them which presented characteristics of interest (oxidase negative, catalase positive and gram negative). From these strains, we tested their sensitivity to different antibiotics using the disc-plate antibiogram method, confirming their intrinsic and acquired resistance to various antibiotics, all of them being resistant to two or more beta-lactam antibiotics. They were identified by biochemical methods, highlighting the detection of strains of Stenotrophomonas maltophilia and Acinetobacter spp. considered clinically important bacteria causing numerous nosocomia

This work was aimed at evaluating loin transcriptome and metabolic pathway differences between the two main Serbian local pig breeds with divergent characteristics regarding muscle growth and fatness, as well as exploring nutrigenomic effects of tannin supplementation in Mangalitsa (MA) pigs. The study comprised 24 Mangalitsa and 10 Moravka (MO) males, which were kept under identical management conditions. Mangalitsa animals were divided in two nutritional groups (n = 12) receiving a standard (control) or tannin–supplemented diet (1.5%; MAT). Moravka pigs were fed the standard mixture. All animals were slaughtered at a similar age; 120 kg of average live weight (LW) and loin tissue was used for RNA‐seq analysis. Results showed 306 differentially expressed genes (DEGs) according to breed, enriched in genes involved in growth, lipid metabolism, protein metabolism and muscle development, such as PDK4, FABP4, MYOD1 and STAT3, as well as a relevant number of genes involved in mitochondrial respiratory activity (MT‐NDs, NDUFAs among others). Oxidative phosphorylation was the most significantly affected pathway, activated in Mangalitsa muscle, revealing the basis of a different muscle metabolism. Also, many other relevant pathways were affected by breed and involved in oxidative stress response, fat accumulation and development of skeletal muscle. Results also allowed the identification of potential regulators and causal networks such as those controlled by FLCN, PPARGC1A or PRKAB1 with relevant regulatory roles on DEGs involved in mitochondrial and lipid metabolism, or IL3 and TRAF2 potentially controlling DEGs involved in muscle development. The Tannin effect on transcriptome was small, with only 23 DEGs, but included interesting ones involved in lipid deposition such as PPARGC1B. The results indicate a significant effect of the breed on muscle tissue gene expression, affecting relevant biological pathways and allowing the identification of strong regulatory candidate ge

Maternal supplementation with antioxidants and n-3 PUFAs may be a promising strategy to reduce the risk of intrauterine growth restriction and preterm delivery, which may diminish the appearance of low-birth-neonates. A previous studies showed beneficial outcomes of the combination of hydroxytyrosol and linoleic acid, but there is no data of its prenatal effects. The present study aimed to determine the possible prenatal implications of such maternal supplementation at prenatal stages in swine, a model of IUGR pregnancies. Results showed effects on litter size, with treated sows having larger litters and, therefore, smaller fetuses. However, the brain/head weight ratio showed a positive effect of the treatment in development, as well as in some other major organs like lungs, spleen, or kidneys. On the other hand, treated piglets showed better glycemic and lipidemic profiles, which could explain postnatal effects. However, further research on the implications of the treatment on litter size and prenatal and postnatal development must be done before practical recommendation can be given.

ROHs are long stretches of DNA homozygous at each polymorphic position. The proportion of genome covered by ROHs and their length are indicators of the level and origin of inbreeding. Frequent common ROHs within the same population define ROH islands and indicate hotspots of selection. In this work, we investigated ROHs in a total of 1131 pigs from 20 European local pig breeds and in three cosmopolitan breeds, genotyped with the GGP Porcine HD Genomic Profiler. plink software was used to identify ROHs. Size classes and genomic inbreeding parameters were evaluated. ROH islands were defined by evaluating different thresholds of homozygous SNP frequency. A functional overview of breed-specific ROH islands was obtained via over-representation analyses of GO biological processes. Mora Romagnola and Turopolje breeds had the largest proportions of genome covered with ROH (~1003 and ~955 Mb respectively), whereas Nero Siciliano and Sarda breeds had the lowest proportions (~207 and 247 Mb respectively). The highest proportion of long ROH (>16 Mb) was in Apulo-Calabrese, Mora Romagnola and Casertana. The largest number of ROH islands was identified in the Italian Landrace (n = 32), Cinta Senese (n = 26) and Lithuanian White Old Type (n = 22) breeds. Several ROH islands were in regions encompassing genes known to affect morphological traits. Comparative ROH structure analysis among breeds indicated the similar genetic structure of local breeds across Europe. This study contributed to understanding of the genetic history of the investigated pig breeds and provided information to manage these pig genetic resources.

Simple Summary The genetic mechanisms that regulate biological processes, such as skeletal muscle development and growth, or intramuscular fat deposition, have attracted great interest, given their impact on production traits and meat quality. In this sense, a comparison of the transcriptome of skeletal muscle between phenotypically different pig breeds, or along growth, could be useful to improve the understanding of the molecular processes underlying the differences in muscle metabolism and phenotypic traits, potentially driving the identification of causal genes, regulators and metabolic pathways involved in their variability. This experiment was conducted to investigate the effects of developmental stage, breed, and diet energy source on the genome-wide expression, meat quality traits, and tissue composition of biceps femoris muscle in growing pure Iberian and Duroc pigs. The study comprised 59 Iberian (IB) and 19 Duroc (DU) animals, who started the treatment at an average live weight (LW) of 19.9 kg. The animals were kept under identical management conditions and fed two diets with different energy sources (6% high oleic sunflower oil or carbohydrates). Twenty-nine IB animals were slaughtered after seven days of treatment at an average LW of 24.1 kg, and 30 IB animals plus all the DU animals were slaughtered after 47 days at an average LW of 50.7 kg. The main factors affecting the muscle transcriptome were age, with 1832 differentially expressed genes (DEGs), and breed (1055 DEGs), while the effect of diet on the transcriptome was very small. The results indicated transcriptome changes along time in Iberian animals, being especially related to growth and tissue development, extracellular matrix (ECM) composition, and cytoskeleton organization, with DEGs affecting relevant functions and biological pathways, such as myogenesis. The breed also affected functions related to muscle development and cytoskeleton organization, as well as functions related to solute tra

Low protein diets supplied during the growing period of pigs can diminish their growth rate and increase the intramuscular fat (IMF) content which affects the sensorial and technological characteristics of the products. In the present study, the effects of a low protein diet supplied during the growing period of Duroc × Iberian crossbred pigs on several phenotypic traits and on liver and longissimus dorsi transcriptome were analysed at the beginning (EARLY) and at the end (LATE) of the growing period. Two experimental groups of 10 crossbred pigs each were fed two isocaloric diets with different protein content: control diet (C) with 16.5% protein and 0.8% lysine and low protein diet (LP) with 11% CP and 0.6% lysine. Animals fed LP diet have a slower growth than those fed C diet, but no effect of LP diet was observed on the IMF content. The transcriptomes of liver and longissimus dorsi were characterised and quantified through RNA-sequencing (RNA-seq). In liver, 134 and 480 differentially expressed annotated genes and new isoforms (DEGs) were detected between C and LP diets for EARLY and LATE animals, respectively. In muscle, 128 and 68 DEGs were detected at EARLY and LATE time-points. Functional interpretation revealed that LP diet may inhibit immune system molecules and processes in both tissues at EARLY stage. In liver, the DEGs mainly affect lipid and cholesterol metabolic processes, while in muscle, the expression changes would be involved in growth, development and meat quality. In conclusion, a low protein diet supplied during the growing period seems to slow down the growth of Duroc × Iberian crossbred pigs, but it also seems to affect multiple biological processes that could compromise the immune system of Duroc × Iberian crossbred pigs. Therefore, these results question the adequacy of this type of regime in Duroc × Iberian pigs that must be studied in greater depth before being implemented.

Gene expression is one of the main factors to influence meat quality by modulating fatty acid metabolism, composition, and deposition rates in muscle tissue. This study aimed to explore the transcriptomics of the Longissimus lumborum muscle in two local pig breeds with distinct genetic background using next-generation sequencing technology and Real-Time qPCR. RNA-seq yielded 49 differentially expressed genes between breeds, 34 overexpressed in the Alentejano (AL) and 15 in the Bísaro (BI) breed. Specific slow type myosin heavy chain components were associated with AL (MYH7) and BI (MYH3) pigs, while an overexpression of MAP3K14 in AL may be associated with their lower loin proportion, induced insulin resistance, and increased inflammatory response via NFkB activation. Overexpression of RUFY1 in AL pigs may explain the higher intramuscular (IMF) content via higher GLUT4 recruitment and consequently higher glucose uptake that can be stored as fat. Several candidate genes for lipid metabolism, excluded in the RNA-seq analysis due to low counts, such as ACLY, ADIPOQ, ELOVL6, LEP and ME1 were identified by qPCR as main gene factors defining the processes that influence meat composition and quality. These results agree with the fatter profile of the AL pig breed and adiponectin resistance can be postulated as responsible for the overexpression of MAP3K14's coding product NIK, failing to restore insulin sensitivity.

This work was aimed at evaluating loin transcriptome and metabolic pathway differences between the two main Serbian local pig breeds with divergent characteristics regarding muscle growth and fatness, as well as exploring nutrigenomic effects of tannin supplementation in Mangalitsa (MA) pigs. The study comprised 24 Mangalitsa and 10 Moravka (MO) males, which were kept under identical management conditions. Mangalitsa animals were divided in two nutritional groups (n = 12) receiving a standard (control) or tannin-supplemented diet (1.5%; MAT). Moravka pigs were fed the standard mixture. All animals were slaughtered at a similar age; 120 kg of average live weight (LW) and loin tissue was used for RNA-seq analysis. Results showed 306 differentially expressed genes (DEGs) according to breed, enriched in genes involved in growth, lipid metabolism, protein metabolism and muscle development, such as PDK4, FABP4, MYOD1 and STAT3, as well as a relevant number of genes involved in mitochondrial respiratory activity (MT-NDs, NDUFAs among others). Oxidative phosphorylation was the most significantly affected pathway, activated in Mangalitsa muscle, revealing the basis of a different muscle metabolism. Also, many other relevant pathways were affected by breed and involved in oxidative stress response, fat accumulation and development of skeletal muscle. Results also allowed the identification of potential regulators and causal networks such as those controlled by FLCN, PPARGC1A or PRKAB1 with relevant regulatory roles on DEGs involved in mitochondrial and lipid metabolism, or IL3 and TRAF2 potentially controlling DEGs involved in muscle development. The Tannin effect on transcriptome was small, with only 23 DEGs, but included interesting ones involved in lipid deposition such as PPARGC1B. The results indicate a significant effect of the breed on muscle tissue gene expression, affecting relevant biological pathways and allowing the identification of strong regulatory candidate ge

Phenotypic variability for productive and meat quality traits has been largely studied in Iberian pigs, especially in genetic selection and nutritional experiments. Complex interactions among genetic background, diet composition and gut microbiota hinder the correct assessment of each factor's contribution on phenotypes. In order to disentangle these interactions, we evaluated changes in gut microbiota composition comparing 48 Iberian and Duroc pigs fed diets with different energy source (standard diet with carbohydrates vs sunflower oil-enriched diet with high oleic acid content).

Coronaviruses silently circulate in human and animal populations, causing mild to severe diseases. Therefore, livestock are important components of a "One Health" perspective aimed to control these viral infections. However, at present there is no example that considers pig genetic resources in this context. In this study, we investigated the variability of four genes (ACE2, ANPEP and DPP4 encoding for host receptors of the viral spike proteins and TMPRSS2 encoding for a host proteinase) in 23 European (19 autochthonous and three commercial breeds and one wild boar population) and two Asian Sus scrofa populations. A total of 2229 variants were identified in the four candidate genes: 26% of them were not previously described; 29 variants affected the protein sequence and might potentially interact with the infection mechanisms. The results coming from this work are a first step towards a "One Health" perspective that should consider conservation programs of pig genetic resources with twofold objectives: (i) genetic resources could be reservoirs of host gene variability useful to design selection programs to increase resistance to coronaviruses; (ii) the described variability in genes involved in coronavirus infections across many different pig populations might be part of a risk assessment including pig genetic resources.

Natural and artificial directional selection in cosmopolitan and autochthonous pig breeds and wild boars have shaped their genomes and resulted in a reservoir of animal genetic diversity. Signatures of selection are the result of these selection events that have contributed to the adaptation of breeds to different environments and production systems. In this study, we analysed the genome variability of 19 European autochthonous pig breeds (Alentejana, Bísara, Majorcan Black, Basque, Gascon, Apulo-Calabrese, Casertana, Cinta Senese, Mora Romagnola, Nero Siciliano, Sarda, Krškopolje pig, Black Slavonian, Turopolje, Moravka, Swallow-Bellied Mangalitsa, Schwäbisch-Hällisches Schwein, Lithuanian indigenous wattle and Lithuanian White old type) from nine countries, three European commercial breeds (Italian Large White, Italian Landrace and Italian Duroc), and European wild boars, by mining whole-genome sequencing data obtained by using a DNA-pool sequencing approach. Signatures of selection were identified by using a single-breed approach with two statistics [within-breed pooled heterozygosity (HP) and fixation index (FST)] and group-based FST approaches, which compare groups of breeds defined according to external traits and use/specialization/type.

In this study, we identified copy number variants (CNVs) in 19 European autochthonous pig breeds and in two commercial breeds (Italian Large White and Italian Duroc) that represent important genetic resources for this species. The genome of 725 pigs was sequenced using a breed-specific DNA pooling approach (30–35 animals per pool) obtaining an average depth per pool of 42×. This approach maximised CNV discovery as well as the related copy number states characterising, on average, the analysed breeds. By mining more than 17.5 billion reads, we identified a total of 9592 CNVs (~683 CNVs per breed) and 3710 CNV regions (CNVRs; 1.15% of the reference pig genome), with an average of 77 CNVRs per breed that were considered as private. A few CNVRs were analysed in more detail, together with other information derived from sequencing data. For example, the CNVR encompassing the KIT gene was associated with coat colour phenotypes in the analysed breeds, confirming the role of the multiple copies in determining breed-specific coat colours. The CNVR covering the MSRB3 gene was associated with ear size in most breeds. The CNVRs affecting the ELOVL6 and ZNF622 genes were private features observed in the Lithuanian Indigenous Wattle and in the Turopolje pig breeds respectively. Overall, the genome variability unravelled here can explain part of the genetic diversity among breeds and might contribute to explain their origin, history and adaptation to a variety of production systems.

Spanish legislation regulates the labelling of Iberian pig meat and dry-cured products, which are labelled as "Ibérico" or "100% Ibérico" when they come from Duroc x Iberian crossbred or Iberian purebred pigs. Although the analytical authentication of breed origin is not mandatory, a genetic diagnostic tool is demanded by producers and consumers. We have designed a 64 Single Nucleotide Variant genotyping panel displaying extreme allelic frequencies between Duroc and Iberian purebred samples. Average proportions of Iberian alleles of 0.99, 0.01, 0.77 and 0.48 were estimated by admixture clustering analysis of known origin samples, for Iberian and Duroc purebred, 75% Iberian and 50% Iberian classes, respectively. A supervised analysis with 1419 samples showed some overlapping between contiguous classes, but the calculated degrees of separability ranged from 0.800 to 0.996, exceeding the threshold value (0.70) for considering suitable for prediction. Therefore, this panel is a useful genetic tool to infer purebred or crossbred Iberian origin of live animals, meat and dry-cured products.

Tenderness is one of the most important meat quality traits and it can be measured through shear force with the Warner–Bratzler test. In the current study, we use the RNA-seq technique to analyze the transcriptome of Longissimus dorsi (LD) muscle in two groups of Iberian pigs (Tough and Tender) divergent for shear force breeding values. We identified 200 annotated differentially expressed genes (DEGs) and 245 newly predicted isoforms. The RNAseq expression results of 10 genes were validated with quantitative PCR (qPCR). Functional analyses showed an enrichment of DE genes in biological processes related to proteolysis (CTSC, RHOD, MYH8, ACTC1, GADD45B, CASQ2, CHRNA9 and ANKRD1), skeletal muscle tissue development (ANKRD1, DMD, FOS and MSTN), lipid metabolism (FABP3 and PPARGC1A) and collagen metabolism (COL14A1). The upstream analysis revealed a total of 11 transcription regulatory factors that could regulate the expression of some DEGs. Among them, IGF1, VGLL3 and PPARG can be highlighted since they regulate the expression of genes involved in biological pathways that could affect tenderness. The experiment revealed a set of candidate genes and regulatory factors suggestive to search polymorphisms that could be incorporated in a breeding program for improving meat tenderness.

Growth is dependent on genotype and diet, even at early developmental stages. In this study, we investigated the effects of genotype, sex, and body weight on the fetal muscle transcriptome of purebred Iberian and crossbred Iberian x Large White pigs sharing the same uterine environment. RNA sequencing was performed on 16 purebred and crossbred fetuses with high body weight (340±14g and 415±14g, respectively) and 16 with low body weight (246±14g and 311±14g, respectively), on gestational day 77. Genotype had the greatest effect on gene expression, with 645 genes identified as differentially expressed (DE) between purebred and crossbred animals. Functional analysis showed differential regulation of pathways involved in energy and lipid metabolism, muscle development, and tissue disorders. In purebred animals, fetal body weight was associated with 35 DE genes involved in development, lipid metabolism and adipogenesis. In crossbred animals, fetal body weight was associated with 60 DE genes involved in muscle development, viability, and immunity. Interestingly, the results suggested an interaction genotype*weight for some DE genes. Fetal sex had only a modest effect on gene expression. This study allowed the identification of genes, metabolic pathways, biological functions and regulators related to fetal genotype, weight and sex, in animals sharing the same uterine environment. Our findings contribute to a better understanding of the molecular events that influence prenatal muscle development and highlight the complex interactions affecting transcriptional regulation during development.

When compared to modern lean-type breeds, Portuguese local Alentejano (AL) and Bísaro (BI) pig breeds present a high potential for subcutaneous and intramuscular fat (IMF) deposition which contributes for better meat quality. The aim of this work was to explore the genome function to better understand the underlying physiological mechanisms associated with body fat accretion. Dorsal subcutaneous fat samples were collected at slaughter from adult animals (n = 4 for each breed) with ~150 kg body weight. Total RNA was obtained and sequenced for transcriptome analysis using DESeq2. A total of 458 differentially expressed (DE) genes (q-value < 0.05) were identified, with 263 overexpressed in AL and 195 in BI. Key genes involved in de novo fatty acid biosynthesis, elongation and desaturation were upregulated in AL such as ACLY, FASN, ME1, ELOVL6 and SCD. A functional enrichment analysis of the DE genes was performed using Ingenuity Pathway Analysis. Cholesterol synthesis is suggested to be higher in AL via SREBF2, SCAP and PPARG, while lipolytic activity may be more active in BI through GH and AMPK signalling. Increased signalling of CD40 together with the predicted activation of INSIG1 and INSIG2 in BI suggests that this breed is more sensitive to insulin whereas the AL is less sensitive like the Iberian breed.

This experiment was conducted to investigate the effects of developmental stage, breed, and diet energy source on the genome-wide expression, meat quality traits, and tissue composition of biceps femoris muscle in growing pure Iberian and Duroc pigs. The study comprised 59 Iberian (IB) and 19 Duroc (DU) animals, who started the treatment at an average live weight (LW) of 19.9 kg. The animals were kept under identical management conditions and fed two diets with different energy sources (6% high oleic sunflower oil or carbohydrates). Twenty-nine IB animals were slaughtered after seven days of treatment at an average LW of 24.1 kg, and 30 IB animals plus all the DU animals were slaughtered after 47 days at an average LW of 50.7 kg. The main factors affecting the muscle transcriptome were age, with 1832 differentially expressed genes (DEGs), and breed (1055 DEGs), while the effect of diet on the transcriptome was very small. The results indicated transcriptome changes along time in Iberian animals, being especially related to growth and tissue development, extracellular matrix (ECM) composition, and cytoskeleton organization, with DEGs affecting relevant functions and biological pathways, such as myogenesis. The breed also affected functions related to muscle development and cytoskeleton organization, as well as functions related to solute transport and lipid and carbohydrate metabolism. Taking into account the results of the two main comparisons (age and breed effects), we can postulate that the Iberian breed is more precocious than the Duroc breed, regarding myogenesis and muscle development, in the studied growing stage., Comunidad de Madrid/FEDER, Ministerio de Economía y Competitividad, Depto. de Producción Animal, Fac. de Veterinaria, TRUE, pub

The present study aimed to determine benefits and risks of a dietary supplementation combining hydroxytyrosol and n-3 polyunsaturated fatty acids (PUFA) on prenatal development and metabolic traits in swine, a model of intrauterine growth restricted (IUGR) pregnancies. No effects were found regarding sows' weight and adiposity. Treated sows had larger litters, with smaller fetuses. However, these animals had better development of some major organs. Fetuses from the treated group had better glycemic and lipidic indexes, but no effects on anti/prooxidant profiles were found. Maternal supplementation with antioxidants and n-3 PUFAs may be a promising strategy to reduce the risk of intrauterine growth restriction and preterm delivery, which may diminish the appearance of low-birth-neonates. A previous studies showed beneficial outcomes of the combination of hydroxytyrosol and linoleic acid, but there is no data of its prenatal effects. The present study aimed to determine the possible prenatal implications of such maternal supplementation at prenatal stages in swine, a model of IUGR pregnancies. Results showed effects on litter size, with treated sows having larger litters and, therefore, smaller fetuses. However, the brain/head weight ratio showed a positive effect of the treatment in development, as well as in some other major organs like lungs, spleen, or kidneys. On the other hand, treated piglets showed better glycemic and lipidemic profiles, which could explain postnatal effects. However, further research on the implications of the treatment on litter size and prenatal and postnatal development must be done before practical recommendation can be given.

Meat quality depends on tissue composition which is in turn influenced by different factors, such as diet, genotype, age, or sex. We evaluated the effects of breed, 24 h fasting, and dietary energy source (HO: oleic acid versus CH: carbohydrates) on the expression of candidate genes involved in adipogenesis, lipogenesis, and lipolysis in the adipose tissue from Iberian and Duroc growing pigs. The Iberian pigs showed greater feed intake, backfat thickness, and saturated fatty acids (SFA) content in the subcutaneous fat, whereas the Duroc pigs had greater ham weight and polyunsaturated fatty acids (PUFA) content. In both breeds, the diet induced changes in the fatty acid (FA) composition of subcutaneous fat samples. The HO group had higher monounsaturated fatty acids (MUFA) and oleic acid, and lower SFA than the CH group. Regarding gene expression, breed and feeding status (fasting versus postprandial) had significant effects on gene expression, with quantitative interactions between them, while diet showed negligible effects. In general, adipogenic and lipogenic genes were upregulated in the Iberian pigs and in postprandial samples. In contrast, the expression of lipolytic genes showed complex interaction effects. Our results agree with the phenotypic differences between the Iberian and Duroc breeds and with the inhibition of lipogenesis by fasting. Quantitative interactions between breed and feeding status effects were observed, which indicates a different response to fasting of the two breeds, with the obese Iberian breed showing a more stable expression of lipogenic genes. These results highlight the complexity of lipid metabolism regulation, especially in relation to lipolysis processes.

The aim of this work was to analyse the distribution of causal and candidate mutations associated to relevant productive traits in twenty local European pig breeds. Also, the potential of the SNP panel employed for elucidating the genetic structure and relationships among breeds was evaluated. Most relevant genes and mutations associated with pig morphological, productive, meat quality, reproductive and disease resistance traits were prioritized and analyzed in a maximum of 47 blood samples from each of the breeds (Alentejana, Apulo-Calabrese, Basque, Bísara, Majorcan Black, Black Slavonian (Crna slavonska), Casertana, Cinta Senese, Gascon, Iberian, Krškopolje (Krškopoljski), Lithuanian indigenous wattle, Lithuanian White Old Type, Mora Romagnola, Moravka, Nero Siciliano, Sarda, Schwäbisch-Hällisches Schwein (Swabian Hall pig), Swallow-Bellied Mangalitsa and Turopolje). We successfully analyzed allelic variation in 39 polymorphisms, located in 33 candidate genes. Results provide relevant information regarding genetic diversity and segregation of SNPs associated to production and quality traits. Coat color and morphological trait-genes that show low level of segregation, and fixed SNPs may be useful for traceability. On the other hand, we detected SNPs which may be useful for association studies as well as breeding programs. For instance, we observed predominance of alleles that might be unfavorable for disease resistance and boar taint in most breeds and segregation of many alleles involved in meat quality, fatness and growth traits. Overall, these findings provide a detailed catalogue of segregating candidate SNPs in 20 European local pig breeds that may be useful for traceability purposes, for association studies and for breeding schemes. Population genetic analyses based on these candidate genes are able to uncover some clues regarding the hidden genetic substructure of these populations, as the extreme genetic closeness between Iberian and Alentejana breeds and

Growth is dependent on genotype and diet, even at early developmental stages. In this study, we investigated the effects of genotype, sex, and body weight on the fetal muscle transcriptome of purebred Iberian and crossbred Iberian x Large White pigs sharing the same uterine environment. RNA sequencing was performed on 16 purebred and crossbred fetuses with high body weight (340±14g and 415±14g, respectively) and 16 with low body weight (246±14g and 311±14g, respectively), on gestational day 77. Genotype had the greatest effect on gene expression, with 645 genes identified as differentially expressed (DE) between purebred and crossbred animals. Functional analysis showed differential regulation of pathways involved in energy and lipid metabolism, muscle development, and tissue disorders. In purebred animals, fetal body weight was associated with 35 DE genes involved in development, lipid metabolism and adipogenesis. In crossbred animals, fetal body weight was associated with 60 DE genes involved in muscle development, viability, and immunity. Interestingly, the results suggested an interaction genotype∗weight for some DE genes. Fetal sex had only a modest effect on gene expression. This study allowed the identification of genes, metabolic pathways, biological functions and regulators related to fetal genotype, weight and sex, in animals sharing the same uterine environment. Our findings contribute to a better understanding of the molecular events that influence prenatal muscle development and highlight the complex interactions affecting transcriptional regulation during development.