MicroRNA expression profiling of porcine mammary epithelial cells after challenge with Escherichia coli in vitro.

BACKGROUND: Coliform mastitis is a symptom of postpartum dysgalactia syndrome (PDS), a multifactorial infectious disease of sows. Our previous study showed gene expression profile change after bacterial challenge of porcine mammary epithelial cells (PMECs). These mRNA expression changes may be regulated through microRNAs (miRNAs) which play critical roles in biological processes. Therefore, miRNA expression profile was investigated in PMECs. RESULTS: PMECs were isolated from three lactating sows and challenged with heat-inactivated potential mastitis-causing pathogen Escherichia coli (E. coli) for 3 h and 24 h, in vitro. At 3 h post-challenge with E. coli, target gene prediction identified a critical role of miRNAs in regulation of host immune responses and homeostasis of PMECs mediated by affecting pathways including cytokine binding (miR-202, miR-3277, miR-4903); IL-10/PPAR signaling (miR-3277, miR-4317, miR-548); and NF-ĸB/TNFR2 signaling (miR-202, miR-2262, miR-885-3p). Target genes of miRNAs in PMECs at 24 h were significantly enriched in pathways associated with interferon signaling (miR-210, miR-23a, miR-1736) and protein ubiquitination (miR-125, miR-128, miR-1280). CONCLUSIONS: This study provides first large-scale miRNA expression profiles and their predicted target genes in PMECs after contact with a potential mastitis-causing E. coli strain. Both, highly conserved miRNAs known from other species as well as novel miRNAs were identified in PMECs, representing candidate predictive biomarkers for PDS. Time-dependent pathogen clearance suggests an important role of PMECs in inflammatory response of the first cellular barrier of the porcine mammary gland.

Methylating micronutrient supplementation during pregnancy influences foetal hepatic gene expression and IGF signalling and increases foetal weight.

PURPOSE: Maternal diet during pregnancy impacts foetal growth and development. In particular, dietary levels of methylating micronutrients (methionine, folate, choline, vitamins B6, and B12) interfere with the availability and allocation of methyl groups for methylation reactions, thereby influencing normal transcription. However, the currently recommended methylating micronutrient supplementation regimen is haphazard and arbitrary at best. METHODS: To investigate the effects of a methylating micronutrient-rich maternal diet, pregnant Pietrain sows were fed either a standard diet (CON) or a diet supplemented with methionine, folate, choline, B6, B12, and zinc (MET). Foetal liver and muscle (M. longissimus dorsi) tissues were collected at 35, 63, and 91 days post-conception. Transcriptional responses to diet were assessed in foetal liver. Altered insulin-like growth factor (IGF) signalling in transcriptome analyses prompted investigation of IGF-2 and insulin-like growth factor binding proteins (IGFBPs) levels in muscle and liver. RESULTS: Maternal diet enriched with methylating micronutrients was associated with increased foetal weight in late gestation. Hepatic transcriptional patterns also revealed differences in vitamin B6 and folate metabolism between the two diets, suggesting that supplementation was effective. Additionally, shifts in growth-supporting metabolic routes of the lipid and energy metabolism, including IGF signalling, and of cell cycle-related pathways were found to occur in liver tissue in supplemented individuals. Weight differences and modulated IGF pathways were also reflected in the muscle content of IGF-2 (increased in MET) and IGFBP-2 (decreased in MET). CONCLUSIONS: Maternal dietary challenges provoke stage-dependent and tissue-specific transcriptomic modulations in the liver pointing to molecular routes contributing to the organismal adaptation. Subtle effects on late foetal growth are associated with changes in the IGF signalling mainly in skeletal muscle tissue that is less resilient to dietary stimuli than liver.

Analysis of non-synonymous SNPs of the porcine SERPINA6 gene as potential causal variants for a QTL affecting plasma cortisol levels on SSC7.

Recently, the SERPINA6 gene encoding corticosteroid-binding globulin (CBG) has been proposed as a candidate gene for a quantitative trait locus (QTL) affecting cortisol level on pig chromosome 7. The QTL was repeatedly detected in different lines, including a Piétrain × (German Landrace × German Large White) cross (PiF1) and purebred German Landrace (LR). In this study, we investigated whether the known non-synonymous polymorphisms c.44G>T, c.622C>T, c.770C>T, c.793G>A, c.832G>A and c.919G>A of SERPINA6 are sufficient to explain the QTL in these two populations. Our investigations revealed that SNPs c.44G>T, c.622C>T, c.793G>A and c.919G>A are associated with cortisol level in PiF1 (P < 0.01). Haplotype analysis showed that these associations are largely attributable to differences between a major haplotype carrying SNPs c.793G>A and c.919G>A and a haplotype carrying SNPs c.44G>T and c.622C>T. Furthermore, some SNPs, particularly c.44G>T and c.622C>T and the carrier haplotype, showed association with meat quality traits including pH and conductivity (P < 0.05). In LR, the non-synonymous SNPs segregate at very low frequency (<5%) and/or show only weak association with cortisol level (SNPs c.832G>A and c.919G>A; P < 0.05). These findings suggest that the non-synonymous SNPs are not sufficient to explain the QTL across different breeds. Therefore, we examined whether the expression of SERPINA6 is affected by cis-regulatory polymorphisms in liver, the major organ for CBG production. We found allelic expression imbalance of SERPINA6, which suggests that its expression is indeed affected by genetic variation in cis-acting elements. This represents candidate causal variation for future studies of the molecular background of the QTL.

Association of TLR5 sequence variants and mRNA level with cytokine transcription in pigs.

The Toll-like receptor 5 (TLR5) plays a crucial role in host defense against flagellated bacteria by recognizing flagellin. Accumulating evidence suggests that single nucleotide polymorphisms (SNPs) in TLR5 have an effect on flagellin recognition and are associated with susceptibility/resistance to disease. In this study, we analyzed association of SNPs, including c.834T>G, c.1065T>C, c.1205C>T, c.1246A>T, c.1269G>A, and c.1398C>T, as well as mRNA level of TLR5 with the abundance of transcripts of cytokines in pigs. SNPs c.1246A>T and c.1269G>A were significantly associated with the transcript abundance of interleukin (IL)-2, and SNPs c.834T>G and c.1398C>T with IL-10 (P < 0.05); the haplotypes showed a tendency to affect the transcript abundance of IL-10 (P = 0.0660) and significantly associated with the transcription of TLR5 (P < 0.01); the abundance of transcripts of TLR5 and IL-10 were strongly correlated (P < 0.01). The results indicated that the SNPs, associated with the transcript abundance of cytokines, were related to immune responsiveness mediated by cytokine, which, in turn, would have a role in pig breeding for disease resistance. Furthermore, the positive correlation between the abundance of TLR5 and IL10 suggest a link between TLR5 activation and IL-10 expression in porcine.

Transcriptional profiling and miRNA-dependent regulatory network analysis of longissimus dorsi muscle during prenatal and adult stages in two distinct pig breeds.

MicroRNAs (miRNAs) and mRNAs establish a complex regulatory network influencing diverse biological pathways including muscle development and growth. Elucidating miRNA-dependent regulatory networks involved in muscle development could provide additional insights into muscle traits largely predefined during prenatal development. The present study aimed to determine differentially expressed transcripts and functional miRNA-mRNA relationships associated with different stages of skeletal muscle development in two pig breeds, German Landrace and Pietrain, distinct in muscle characteristics. A comparative transcriptional profiling of longissimus dorsi muscle tissues from fetuses at 35, 63 and 91 days post-conception as well as adult pigs (180 days postnatum) was performed using the Affymetrix GeneChip porcine genome microarray. Differential expression patterns were identified to be associated with muscularly developmental stages and breed types. The integration of miRNA expression data and ingenuity pathways analysis (ipa) pathway analysis revealed several miRNA-dependent regulatory networks related to muscle growth and development. The present results provide insights into muscle biology for further improvement of porcine meat quality.

QTL region-specific microarrays reveal differential expression of positional candidate genes of signaling pathways associated with the liability for the inverted teat defect.

The inverted teat defect is the most common disorder of the mammary complex in pigs. It is characterized by the failure of teats to protrude from the udder surface, preventing normal milk flow and thus limiting the rearing capacity and increasing the risk of mastitis. The inverted teat defect is a liability trait with a complex mode of inheritance. We previously identified QTL for inverted teats. As a complementary approach that integrates map-based efforts to identify candidate genes for the inverted teat defect with function-driven expression analysis, application-specific microarrays were constructed that cover 1525 transcripts mapping in QTL regions on pig chromosomes 2, 3, 4, 6 and 11. About 950 transcripts were expressed in epithelial and mesenchymal teat tissue. The expression of three categories of teats was compared: normal teats of both non-affected and affected animals and inverted teats of affected animals. In epithelium and mesenchyme, 62 and 24 genes respectively were significantly differentially expressed (DE). The majority of biofunctions to which a significant number of DE genes were assigned are related to the following: (1) cell maintenance, proliferation, differentiation and replacement; (2) organismal, organ and tissue development; or (3) genetic information and nucleic acid processing. Moreover, the DE genes belong almost exclusively to canonical pathways related to signaling rather than metabolic pathways. This is in line with findings obtained by genome-wide catalogue microarrays. This study adds another piece to the puzzle of the etiology of inverted teats by indicating that causal genetic variation leading to the disorder is likely among the genes encoding for members of the signaling cascades of growth factors.

Association of TLR4 polymorphism with cytokine expression level and pulmonary lesion score in pigs.

The toll-like receptor 4 (TLR4), recognizing lipopolysaccharide of gram-negative bacteria, plays an essential role in immune responses. Variation in TLR4 alters host immune responses to pathogen and is associated with resistance/susceptibility to infectious diseases, as suggested by studies in humans and agricultural species, including cattle and chicken. In this study, we analyzed association of single nucleotide polymorphisms (SNPs) of TLR4 with cytokine expression level and pulmonary lesion score in swine. The SNP c.611 T>A showed significant association with the transcription levels of IFNG, TNFA, and IL-6 (P < 0.05); the SNP c.962 G>A showed significant association with the transcription of IFNG, IL-2, and IL-4 (P < 0.05); the SNP c.1,027 C>A showed significant association with the transcription of IFNG and IL-6 (P < 0.05); the haplotypes showed significant association with the transcription of IFNG, IL-2, IL-4, IL-6, and TNFA (P < 0.05). Both SNPs c.611 T>A and c.962 G>A showed significant association with pulmonary lesion scores (P < 0.01); and the combination genotypes of 3 polymorphic sites were also significantly associated with pulmonary lesion scores (P < 0.01). The observed relationship between TLR4 polymorphism and the transcription levels of cytokines indicate that these SNPs are related to the modulation of the cytokine mediated immune response.

Genes with expression levels correlating to drip loss prove association of their polymorphism with water holding capacity of pork.

Six genes that were known to exhibit expression levels that are correlated to drip loss BVES, SLC3A2, ZDHHC5, CS, COQ9, and EGFR have been for candidate gene analysis. Based on in silico analysis SNPs were detected, confirmed by sequencing, and used for genotyping. The SNPs were genotyped in about 1,800 animals from six pig populations including commercial herds of Pietrain (PI) and German Landrace (DL), different commercial herds of Pietrain×(German Large White×German Landrace) (PIF1(a/b/c)), and one experimental F2-population Duroc×Pietrain (DUPI). Comparative and genetic mapping established the location of BVES on SSC1, of SLC3A2 and ZDHHC5 on SSC2, of CS on SSC5, of COQ9 on SSC6 and of EGFR on SSC9, respectively, coinciding with QTL regions for carcass and meat quality traits. BVES, SLC3A2, and CS revealed association at least with drip loss and with several other measures of water holding capacity (WHC). Moreover, COQ9 and EGFR were associated with several meat quality traits such as meat color and/or thawing loss. This study reveals statistic evidence in addition to the functional relationship of these genes to WHC previously evidenced by expression analysis. This study reveals positional and genetic statistical evidence for a link of genetic variation at these loci or close to them and promotes those six candidate genes as functional and/or positional candidate genes for meat quality traits.

Integrating expression profiling and whole-genome association for dissection of fat traits in a porcine model.

Traits related to fatness, important as economic factors in pork production, are associated with serious diseases in humans. Genetical genomics is a useful approach for studying the effects of genetic variation at the molecular level in biological systems. Here we applied a whole-genome association analysis to hepatic gene expression traits, focusing on transcripts with expression levels that correlated with fatness traits in a porcine model. A total of 150 crossbred pigs [Pietrain × (German Large White × German Landrace)] were studied for transcript levels in the liver. The 24K Affymetrix expression microarrays and 60K Illumina single nucleotide polymorphism (SNP) chips were used for genotyping. A total of 663 genes, whose expression significantly correlated with the trait "fat area," were analyzed for enrichment of functional annotation groups as defined in the Ingenuity Pathways Knowledge Base (IPKB). Genes involved in metabolism of various macromolecules and nutrients as well as functions related to dynamic cellular processes correlated with fatness traits. Regions affecting the transcription levels of these genes were mapped and revealed 4,727 expression quantitative trait loci (eQTL) at P < 10⁻5, including 448 cis-eQTL. In this study, genome-wide association analysis of trait-correlated expression was successfully used in a porcine model to display molecular networks and list genes relevant to fatness traits.

Expression quantitative trait loci analysis of genes in porcine muscle by quantitative real-time RT-PCR compared to microarray data.

Genetic analysis of transcriptional profiling is a promising approach for identifying biological pathways and dissecting the genetics of complex traits. Here, we report on expression quantitative trait loci (eQTL) that were estimated from the quantitative real-time RT-PCR data of 276 F(2) animals and compared with eQTL identified using 74 microarrays. In total, 13 genes were selected that showed trait-dependent expression in microarray experiments and exhibited 21 eQTL. Real-time RT-PCR and microarray data revealed seven cis eQTL in total, of which one was only detected by real-time RT-PCR, one was only detected by microarray analysis, three were consistently found in overlapping intervals and two were in neighbouring intervals on the same chromosome; whereas no trans eQTL was confirmed. We demonstrate that cis regulation is a stable characteristic of individual transcripts. Consequently, a global microarray eQTL analysis of a limited number of samples can be used for exploring functional and regulatory gene networks and scanning for cis eQTL, whereas the subsequent analysis of a subset of likely cis-regulated genes by real-time RT-PCR in a larger number of samples is relevant to narrow down a QTL region by targeting these positional candidate genes. In fact, when modelling SNPs of six genes as fixed effects in the eQTL analysis, eQTL peaks were shifted downwards, experimentally confirming the impact of the respective polymorphic genes, although these SNPs were not located in the regulatory sequence and these shifts occur as a result of linkage disequilibrium in the F(2) population.

Porcine muscle sensory attributes associate with major changes in gene networks involving CAPZB, ANKRD1, and CTBP2.

Principal component analysis of traits related to carcass and meat properties were combined with microarray expression data for the identification of functional networks of genes and biological processes taking place during the conversion of muscle to meat. Principal components (PCs) with high loadings of meat quality traits were derived from phenotypic data of 572 animals of a porcine crossbreed population. Microarray data of 74 M. longissimus dorsi samples were correlated with PC datasets. Lists of significantly correlated genes were analyzed for enrichment of functional annotation groups as defined in the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes databases as well as the Ingenuity Pathways Analysis library. Ubiquitination, phosphorylation, mitochondrion dysfunction, actin, integrin, platelet-derived growth factor, epidermal growth factor, vascular endothelial growth factor, and Ca signaling pathways are correlated with meat quality. Among the significantly trait-associated genes, CAPZB, ANKRD1, and CTBP2 are promoted as candidate genes for meat quality that provide a link between the highlighted pathways. Knowledge of the relevant biological processes and the differential expression of members of the pathway will provide tools that are predictive for traits related to meat quality and that may also be diagnostic for many muscle defects or damages including muscle atrophy, dystrophy, and hypoxia.

Evidence for association of lymphoid enhancer-binding factor-1 (LEF1) with the number of functional and inverted teats in pigs.

The lymphoid enhancer-binding factor-1 (LEF1) belongs to a family of regulatory proteins that share homology with the high mobility group protein-1 (HMG1). The LEF1 gene is a mediator in the canonical Wnt-signalling pathway required for morphogenesis of early mammary gland during embryogenesis. Here we describe the molecular characterisation of the porcine LEF1 gene and its association with number of teats and inverted teats in experimental and commercial populations. The 2357-bp cDNA sequence contains an 1197-bp open reading frame encoding a protein of 398 amino acids. The porcine LEF1 protein shares high identity with LEF1 in other mammalian species. The LEF1 gene contains 12 exons and maps to pig chromosome 8 (SSC8). We identified two single nucleotide polymorphisms (SNPs), a T1351C transition and an A1666C transversion, in the 3' end of LEF1. Associations of the SNP A1666C with presence of inverted teats (P

QTL for traits related to humoral immune response estimated from data of a porcine F2 resource population.

This study aimed to map quantitative trait loci (QTL) for traits related to humoral innate immune defence. Therefore, haemolytic complement activity in the alternative and the classical pathway, serum concentration of C3c and of haptoglobin (HP) were measured in blood samples obtained from F2 piglets (n = 457) of a porcine F2 resource population before and after Mycoplasma hyopneumoniae, Aujeszky's disease virus (Suid herpesvirus I, SuHVI) and porcine reproductive and respiratory syndrome virus (PRRSV) vaccination at 6, 14 and 16 weeks of age. Animals were genotyped at 88 autosomal markers. QTL analysis was performed under the line cross and the half sib. Phenotypic data were adjusted for systematic effects by mixed models with and without repeated measures statement. In total, 46 and 21 estimated QTL positions were detected with genome-wide significance at the 0.05 and 0.01 level, respectively. The proximal region of SSC2 (orthologous to HSA11 0-70 Mb), the distal region of SSC4 (HSA1 95-155 Mb), and the intermediate region of SSC16 (HSA5 0-73 Mb and 150-174 Mb) showed a clustering of estimated QTL positions for complement activity based on the different models. A common genetic background, i.e. a single true QTL, might underlie these QTL positions for related traits. In addition, QTL for antibody titres were detected on SSC1, 2, 6 and 7. With regard to number and magnitude of their impact, QTL for humoral innate immune traits behave like those for other quantitative traits. Discovery of such QTL facilitates the identification of candidate genes for disease resistance and immune competence that are applicable in selective breeding and further research towards improving therapeutic and prophylactic measures.

Expression of the porcine adrenergic receptor beta 2 gene in longissimus dorsi muscle is affected by cis-regulatory DNA variation.

The beta-2 adrenergic receptor (AR) mediates metabolic actions of catecholamines, including glycogenolysis, lipolysis and proteolysis, in muscle and adipose tissue. Factors influencing the density of beta-2 ARs thus might affect carcass composition and meat quality. One such factor might represent cis-regulatory DNA variation affecting mRNA expression of the adrenergic receptor beta 2 (ADRB2) gene in relevant tissues. To identify potential cis-regulatory DNA variation of porcine ADRB2, we comparatively sequenced part of the 5' flanking region and identified 10 single nucleotide polymorphisms (SNPs). The SNP at position g.673C>T (AF000134) resides in an evolutionarily conserved region (ECR) in an in silico predicted androgen response element. Quantification of total transcript levels and allelic expression imbalance (AEI) revealed significant variability in mRNA expression of ADRB2 in longissimus dorsi muscle of slaughter pigs, partly attributable to cis-regulatory DNA variation. However, the g.673C>T SNP has, in the given temporo-spatial context, no significant effect but is apparently in linkage disequilibrium with the causal cis-regulatory DNA variant. We used the g.673C>T SNP as a marker to study the association of ADRB2 variation with carcass and meat quality in four commercial lines. We found association with the pH of loin at 45 min and 24 h postmortem (p.m.) and with the pH of ham at 24 h p.m. Supporting evidence for ADRB2 as a candidate gene for pork quality is provided by our assignment of the gene to the telomeric end of the q arm of porcine chromosome 2, where several quantitative trait loci for meat quality were reported.

Association of parathyroid hormone-like hormone (PTHLH) and its receptor (PTHR1) with the number of functional and inverted teats in pigs.

Parathyroid hormone-like hormone gene (PTHLH) and its receptor, parathyroid hormone/ parathyroid hormone-like hormone receptor 1 (PTHR1), play a role in epithelial mesenchymal interactions during growth and differentiation of different tissues and anatomic structures, including teats. Therefore, PTHLH and PTHR1 were evaluated as functional candidate genes for their effects on number and shape of teats in pigs. In particular, focus was on the occurrence and number of inverted teats, the most frequent and economically relevant teat developmental defect in pigs. For this purpose, association and linkage of the PTHLH gene and the PTHR1 gene with inverted teat defect and the total number of teats and inverted teats were studied in an experimental Duroc and Berlin Miniature pig (DUMI) population. Polymorphism C1819T of PTHR1 was significantly associated with inverted teat phenotype (p = 0.014), total number of teats (p = 0.047) and was close to significance with the number of inverted teats (p = 0.078). Polymorphism C375T of PTHLH was close to significance with the inverted teat phenotype (p = 0.122) and showed no significant association with the total number of teats (p = 0.621) and the number of inverted teats (p = 0.256) in the DUMI population. Association analyses were also performed for combined effects of PTHLH and PTHR1 in order to address potential interaction, however, revealed no indication of effects of interaction. The function, position and the association shown here promote PTHR1 as a candidate gene for number of teats and in particular for affection by and number of inverted teats.

Porcine IL12A and IL12B gene mapping, variation and evidence of association with lytic complement and blood leucocyte proliferation traits.

Interleukin-12, a heterodimeric cytokine consisting of glycosylated subunits of 35 and 40 kDa, is a central molecule in controlling innate as well as adaptive immunity. This study was aimed to investigate the role of IL12A and IL12B as candidate genes for immune competence in pigs. The porcine genes were screened for polymorphism and association analysis was carried out by mixed model analysis with parameters of innate immunity, in vitro haemolytic complement activity in the classical and alternative pathways, in vivo complement activation expressed as C3c serum concentration, and blood leucocyte proliferation measured in F2 animals of a pig resource population based on cross of Duroc and Berlin miniature pig (DUMI resource population). A single nucleotide polymorphism (SNP) in the promoter region (C > A) of IL12A was identified. Two SNPs were detected in intron 4 of IL12B at positions 192 (A > G) and 437 (C > T). Significant effects of IL12 genotypes on complement activity traits and mitogen-induced leucocyte proliferation were found. The IL12A and IL12B genes were assigned to chromosome13 and 16, respectively, by using radiation hybrid analysis and genetic mapping in the DUMI resource population. Mapping and association analyses promote the IL12 genes as functional and positional candidate gene for disease resistance in pigs.

Porcine genome-wide gene expression in response to tetanus toxoid vaccine.

The very early in vivo response to immune stimuli was studied using tetanus toxoid as a model antigen known to induce Th1 and Th2 responses. Eighteen weaning piglets were vaccinated subcutaneously with tetanus toxoid. Leukocyte RNAs were isolated from samplings before and 2, 4, 8, and 24 hours after vaccination. After competitive hybridization of a 13297 porcine 70-mer oligo microarray (Qiagen-Operon NRSP8), subsequent image analysis and normalization, the data was analysed using analyses of variance (ANOVA) to identify genes regulated due to vaccination (ANOVA, p < or = 0.05; corresponding to false discovery q < or = 0.12). Of 8240 probes representing genes expressed in leukocytes, 1289 genes showed differential expression. Results were exemplarily confirmed by real-time PCR. Holistic expression data was clustered to six prominent groups of genes with similar changes in expression patterns using a k-means algorithm. To get more insight into functional and structural components and impact of the genes represented in each cluster, the EASE score was used to identify Gene Ontology functional categories. The results showed that in vivo significant changes of expression profiles of peripheral blood mononuclear cells (PBMCs) occurred very early after immune stimuli. These alterations concerned genes of pathways related to immune response as well as other metabolic and regulatory pathways including ATP/energy metabolism, transcription, structural molecule activity, biosynthesis, and metabolism. The analysis reveals new functional candidate genes for traits related to immune responsiveness and also provides new insight into the interaction of immune response, and metabolic and endocrine status. This will facilitate a better understanding of the relationship between immune and performance traits.

Verification of chromosomal regions affecting the innate immunity in pigs using linkage mapping.

Understanding the genetic control of innate immunity in pigswould offerthe opportunity to utilize natural variation and improve selective breeding programmes. As part of our porcine genome scan to identify quantitative trait loci (QTL) we examined immune response traits in a Duroc x Berlin miniature pig resource family (DUMI). Complement activity via classical (CH50) and alternative (AH50) pathways, antibody response to Mycoplasma hyopneumoniae (Mh), tetanus toxoid (TET) and PRRS virus (PRRSV), the complement component (C3c), and Haptoglobin serum concentration (HP) were used as phenotypes for linkage mapping. A total of 220 backcross animals were used for the QTL analysis. Blood was collected six times from each animal prior to and after vaccinations against Mycoplasma hyopneumoniae, tetanus toxoid and PRRS, respectively. Seventy-four microsatellites from 18 autosomes were used for QTL mapping. The analyses were performed treating the measurements of phenotypes at different time points as single traits. Forty-two significant and 24 highly significant QTL were detected, using the program QTL Express, for all immune traits using the single traits. Most QTL were detected on SSC3, SSC16, and SSC18. No significant F-value corresponded to data for SSC12 and SSC13.

Physiological interactions between the endocrine and immune systems shown in gene analysis in pigs.

A number of observations support the concept of important physiological interactions between the endocrine and immune systems. It could be confirmed that hormones secreted by the neuroendocrine system play an important role in communication and regulation of the cells of the immune system. Among protein hormones, this has been most clearly documented for prolactin (PRL), growth hormone (GH), and insulin-like growth factor 1 (IGF-1). A number of traits of the immune response in a Duroc x Berlin Miniature pig family (DUMI) were examined. The haemolytic complement activity in the classical complement pathway (CH50) and the alternative pathway (AH50)was examined at eight different time points in pigs that were vaccinated with different vaccines. Single nucleotide polymorphism (SNP) genotyping was employed to genotype the DUMI F2 animal for growth hormone (Gh), growth hormone releasing hormone (Ghrh), pituitary-specific transcription factor 1 (Pit1), and prolactin receptor (Prlr) loci, and also a microsatellite within insulin-like growth factor 1 (Igf1). Using a family-based association test (FBAT) program, a highly significant association of Gh, Pit1, and Prlr to AH50 (p < or = 0.01) and a significant association of Pit1 to CH50 (p < or = 0.05) were found. Using the SAS system for mixed model, a highly significant association of Gh, and Igf1 to AH50 and CH50 (p < or = 0.01) was detected, while Prlr and Ghrh had a highly significant association (p < or = 0.01) with CH50 only.

Structural and functional genomics to elucidate the genetic background of microstructural and biophysical muscle properties in the pig.

Linkage analyses enable identifying genomic regions that exhibit quantitative trait loci (QTL) without prior hypothesis on the physiology of a trait. Function-oriented expression analyses are a complementary approach to derive hypothesis on the genetic background of phenotypic variation. Muscle fibre types and size affect body composition and meat quality traits. The number and proportion of muscle fibres are to a large extent determined during the prenatal development. Consequently, QTL for muscle fibre, meat quality and carcass traits were detected in a porcine experimental population based on Duroc and Berlin Miniature Pig. Regions with either significant QTL for muscle fibre traits or significant QTL for meat quality and muscularity or both were detected on SSC1, 2, 3, 4, 5, 13, 14, 15 and 16. Here, effects on the complex traits of muscularity and meat quality might be the result of genetic variation primarily affecting fibre type distribution traits. To complement the QTL study expression profiling of prenatal muscle tissue of Duroc and Pietrain was conducted that revealed a list of functional candidate genes for meat quality and carcass traits of various physiological networks. Assignments of these genes to QTL regions highlight them as positional functional candidates. Exemplarily, five genes were analysed further and shown to be associated with meat quality and carcass traits. Further, the relative MYH isotype transcript abundance was found to be associated with muscularity. Relative MYH isotype transcript abundance is proposed as a new phenotype to unravel the genetic background of variation in traits related to muscle and meat properties.