Equine SNP50 BeadChip Research Articles

Genome-wide association study suggests genetic candidate loci of insulin dysregulation in Finnhorses

Equine metabolic syndrome (EMS) is a common welfare problem in horses worldwide. It is characterized by insulin dysregulation (ID), predisposition to laminitis and often obesity. EMS is multifactorial by nature, with both the environment and genetics contributing to the phenotype. Environmental factors, such as feeding and exercise, can be controlled, thus forming the basis for treatment and prevention. Genetic factors, by contrast, are less well-known and not easily controllable. The aim of this study was to identify potential genetic loci influencing ID/EMS in Finnhorses. A single-breed (Finnhorse) case-control genome-wide association study (GWAS) of ID was conducted with controls that included age-appropriate non-ID horses. ID status was determined with an oral sugar test (OST) for fasted horses. Seventy-one Finnhorses participated (n = 34 ID, n = 37 control). DNA samples (hair roots) were genotyped for 65 157 single-nucleotide polymorphisms (SNPs) with the Illumina Equine SNP70 BeadChip, and these data were analysed for association and FST outliers with genomic tools. P-values that exceeded the suggestive threshold (P = 1.00 ×10−5) were found in SNP BIEC2_383954 (P = 3.45 ×10−6) in chromosome 17 and SNP BIEC2_312374 (P = 1.89 ×10−5) in chromosome 15. Hierarchical and Bayesian FST outlier tests also detected these SNPs. Potential candidate genes associated with the ID close to SNP BIEC2_383954, with functions in carbohydrate metabolism, were Arginine and Glutamate Rich 1 (ARGLU1) and Ephrin-B2 (EFNB2).

Genomic inbreeding estimation, runs of homozygosity, and heterozygosity-enriched regions uncover signals of selection in the Quarter Horse racing line.

With the advent of genomics, significant progress has been made in the genetic improvement of livestock species, particularly through increased accuracy in predicting breeding values for selecting superior animals and the possibility of performing a high-resolution genetic scan throughout the genome of an individual. The main objectives of this study were to estimate the individual genomic inbreeding coefficient based on runs of homozygosity (FROH ), to identify and characterize runs of homozygosity and heterozygosity (ROH and ROHet, respectively; length and distribution) throughout the genome, and to map selection signatures in relevant chromosomal regions in the Quarter Horse racing line. A total of 336 animals registered with the Brazilian Association of Quarter Horse Breeders (ABQM) were genotyped. One hundred and twelve animals were genotyped using the Equine SNP50 BeadChip (Illumina, USA), with 54,602 single nucleotide polymorphisms (SNPs; 54K). The remaining 224 samples were genotyped using the Equine SNP70 BeadChip (Illumina, USA) with 65,157 SNPs (65K). To ensure data quality, we excluded animals with a call rate below 0.9. We also excluded SNPs located on non-autosomal chromosomes, as well as those with a call rate below 0.9 or a p-value below 1 × 10-5 for Hardy-Weinberg equilibrium. The results indicate moderate to high genomic inbreeding, with 46,594 ROH and 16,101 ROHet detected. In total, 30 and 14 candidate genes overlap with ROH and ROHet regions, respectively. The ROH islands showed genes linked to crucial biological processes, such as cell differentiation (CTBP1, WNT5B, and TMEM120B), regulation of glucose metabolic process (MAEA and NKX1-1), heme transport (PGRMC2), and negative regulation of calcium ion import (VDAC1). In ROHet, the islands showed genes related to respiratory capacity (OR7D19, OR7D4G, OR7D4E, and OR7D4J) and muscle repair (EGFR and BCL9). These findings could aid in selecting animals with greater regenerative capacity and developing treatments for muscle disorders in the QH breed. This study serves as a foundation for future research on equine breeds. It can contribute to developing reproductive strategies in animal breeding programs to improve and preserve the Quarter Horse breed.

Heritability of insidious uveitis in Appaloosa horses.

Equine recurrent uveitis (ERU) is a blinding ocular disorder among horses, and the Appaloosa horse breed is disproportionally affected by a chronic form of this intraocular inflammatory disease known as insidious uveitis. Strong breed predisposition and previous investigations suggest that there is a genetic component to the pathology of insidious uveitis among Appaloosa horses; however, no estimates of the heritability of the disease have previously been determined. This study aimed to characterize the genetic underpinning of the disease by estimating the heritability for insidious uveitis among Appaloosas. After combining two genotyping array datasets from the Illumina Equine SNP70 BeadChip and the Axiom Equine 670 K Genotyping Array, heritability was estimated for 59 affected and 83 unaffected horses using both restricted maximum likelihood (REML) and phenotype correlation - genotype correlation solvers from the linkage disequilibrium adjusted kinship software. Based on previous research, age and sex were used as covariates, and the locus responsible for the characteristic Appaloosa coat pattern (LP), previously associated with ERU risk, was included as a fixed effect ('top predictor'). Using prevalence values from 0.05 to 0.42, the heritability estimate for insidious uveitis ranged from 0.95 (SE=0.14) to 1.74 (SE=0.25) with LP contributing 0.16-0.33 to the estimate. This study suggests that insidious uveitis is highly heritable (REML 95% CI, h2 =0.68-1.0) and additional loci outside of LP are contributing to the genetic risk for insidious uveitis for Appaloosas. Once identified, these other genetic factors may lead to new disease mitigation efforts in veterinary care and breeding practices.

Selection in the Finnhorse, a native all-around horse breed.

Selection by breeders modifies the morphology, behaviour and performance of domesticated species. Here, we examined signs of selection in Finnhorse, the only native horse breed in Finland. We first searched divergent genomic regions between Finnhorses and other breeds, as well as between different breeding sections of the Finnhorse with data from Illumina Equine SNP70 BeadChip, and then studied several of the detected regions in more detail. We found altogether 35 common outlier SNPs between Finnhorses and other breeds using two different selection tests. Many of the SNPs were located close to genes affecting coat colour, performance, size, sugar metabolism, immune response and olfaction. We selected genes affecting coat colour (KIT, MITF, PMEL), performance (MSTN) and locomotion (DMRT3) for a more detailed examination. In addition, we looked for, and found, associations with height at withers and SNPs located close to gene LCORL. Among the four breeding sections of Finnhorses (harness trotters, riding horses, draught horses and pony‐sized horses), a single SNP located close to the DMRT3 gene was significantly differentiated and only between harness trotters and pony‐sized horses.

Genetic investigation of equine recurrent uveitis in Appaloosa horses.

Equine recurrent uveitis (ERU) is characterized by intraocular inflammation that often leads to blindness in horses. Appaloosas are more likely than any other breed to develop insidious ERU, distinguished by low-grade chronic intraocular inflammation, suggesting a genetic predisposition. Appaloosas are known for their white coat spotting patterns caused by the leopard complex spotting allele (LP) and the modifier PATN1. A marker linked to LP on ECA1 and markers near MHC on ECA20 were previously associated with increased ERU risk. This study aims to further investigate these loci and identify additional genetic risk factors. A GWAS was performed using the Illumina Equine SNP70 BeadChip in 91 horses. Additive mixed model approaches were used to correct for relatedness. Although they do not reach a strict Bonferroni genome-wide significance threshold, two SNPs on ECA1 and one SNP each on ECA12 and ECA29 were among the highest ranking SNPs and thus warranted further analysis (P=1.20×10-5 , P=5.91×10-6 , P=4.91×10-5 , P=6.46×10-5 ). In a second cohort (n=98), only an association with the LP allele on ECA1 was replicated (P=5.33×10-5 ). Modeling disease risk with LP, age and additional depigmentation factors (PATN1 genotype and extent of roaning) supports an additive role for LP and suggests an additive role for PATN1. Genotyping for LP and PATN1 may help predict ERU risk (AUC=0.83). The functional role of LP and PATN1 in ERU development requires further investigation. Testing samples across breeds with leopard complex spotting patterns and a denser set of markers is warranted to further refine the genetic components of ERU.

Expression Quantitative Trait Loci in Equine Skeletal Muscle Reveals Heritable Variation in Metabolism and the Training Responsive Transcriptome.

While over ten thousand genetic loci have been associated with phenotypic traits and inherited diseases in genome-wide association studies, in most cases only a relatively small proportion of the trait heritability is explained and biological mechanisms underpinning these traits have not been clearly identified. Expression quantitative trait loci (eQTL) are subsets of genomic loci shown experimentally to influence gene expression. Since gene expression is one of the primary determinants of phenotype, the identification of eQTL may reveal biologically relevant loci and provide functional links between genomic variants, gene expression and ultimately phenotype. Skeletal muscle (gluteus medius) gene expression was quantified by RNA-seq for 111 Thoroughbreds (47 male, 64 female) in race training at a single training establishment sampled at two time-points: at rest (n = 92) and four hours after high-intensity exercise (n = 77); n = 60 were sampled at both time points. Genotypes were generated from the Illumina Equine SNP70 BeadChip. Applying a False Discovery Rate (FDR) corrected P-value threshold (P FDR < 0.05), association tests identified 3,583 cis-eQTL associated with expression of 1,456 genes at rest; 4,992 cis-eQTL associated with the expression of 1,922 genes post-exercise; 1,703 trans-eQTL associated with 563 genes at rest; and 1,219 trans-eQTL associated with 425 genes post-exercise. The gene with the highest cis-eQTL association at both time-points was the endosome-associated-trafficking regulator 1 gene (ENTR1; Rest: P FDR = 3.81 × 10-27, Post-exercise: P FDR = 1.66 × 10-24), which has a potential role in the transcriptional regulation of the solute carrier family 2 member 1 glucose transporter protein (SLC2A1). Functional analysis of genes with significant eQTL revealed significant enrichment for cofactor metabolic processes. These results suggest heritable variation in genomic elements such as regulatory sequences (e.g. gene promoters, enhancers, silencers), microRNA and transcription factor genes, which are associated with metabolic function and may have roles in determining end-point muscle and athletic performance phenotypes in Thoroughbred horses. The incorporation of the eQTL identified with genome and transcriptome-wide association may reveal useful biological links between genetic variants and their impact on traits of interest, such as elite racing performance and adaptation to training.

A Mechanogenetic Model of Exercise-Induced Pulmonary Haemorrhage in the Thoroughbred Horse.

Exercise-induced pulmonary haemorrhage (EIPH) occurs in horses performing high-intensity athletic activity. The application of physics principles to derive a ‘physical model’, which is coherent with existing physiology and cell biology data, shows that critical parameters for capillary rupture are cell–cell adhesion and cell stiffness (cytoskeleton organisation). Specifically, length of fracture in the capillary is a ratio between the energy involved in cell–cell adhesion and the stiffness of cells suggesting that if the adhesion diminishes and/or that the stiffness of cells increases EIPH is more likely to occur. To identify genes associated with relevant cellular or physiological phenotypes, the physical model was used in a post-genome-wide association study (GWAS) to define gene sets associated with the model parameters. The primary study was a GWAS of EIPH where the phenotype was based on weekly tracheal wash samples collected over a two-year period from 72 horses in a flat race training yard. The EIPH phenotype was determined from cytological analysis of the tracheal wash samples, by scoring for the presence of red blood cells and haemosiderophages. Genotyping was performed using the Illumina Equine SNP50 BeadChip and analysed using linear regression in PLINK. Genes within significant genome regions were selected for sets based on their GeneOntology biological process, and analysed using fastBAT. The gene set analysis showed that genes associated with cell stiffness (cytoskeleton organisation) and blood flow have the most significant impact on EIPH risk.

Selection signatures in four German warmblood horse breeds: Tracing breeding history in the modern sport horse.

The study of selection signatures helps to find genomic regions that have been under selective pressure and might host genes or variants that modulate important phenotypes. Such knowledge improves our understanding of how breeding programmes have shaped the genomes of livestock. In this study, 942 stallions were included from four, exemplarily chosen, German warmblood breeds with divergent historical and recent selection focus and different crossbreeding policies: Trakehner (N = 44), Holsteiner (N = 358), Hanoverian (N = 319) and Oldenburger (N = 221). Those breeds are nowadays bred for athletic performance and aptitude for show-jumping, dressage or eventing, with a particular focus of Holsteiner on the first discipline. Blood samples were collected during the health exams of the stallion preselections before licensing and were genotyped with the Illumina EquineSNP50 BeadChip. Autosomal markers were used for a multi-method search for signals of positive selection. Analyses within and across breeds were conducted by using the integrated Haplotype Score (iHS), cross-population Extended Haplotype Homozygosity (xpEHH) and Runs of Homozygosity (ROH). Oldenburger and Hanoverian showed very similar iHS signatures, but breed specificities were detected on multiple chromosomes with the xpEHH. The Trakehner clustered as a distinct group in a principal component analysis and also showed the highest number of ROHs, which reflects their historical bottleneck. Beside breed specific differences, we found shared selection signals in an across breed iHS analysis on chromosomes 1, 4 and 7. After investigation of these iHS signals and shared ROH for potential functional candidate genes and affected pathways including enrichment analyses, we suggest that genes affecting muscle functionality (TPM1, TMOD2-3, MYO5A, MYO5C), energy metabolism and growth (AEBP1, RALGAPA2, IGFBP1, IGFBP3-4), embryonic development (HOXB-complex) and fertility (THEGL, ZPBP1-2, TEX14, ZP1, SUN3 and CFAP61) have been targeted by selection in all breeds. Our findings also indicate selection pressure on KITLG, which is well-documented for influencing pigmentation.

Genotype imputation accuracy in multiple equine breeds from medium- to high-density genotypes.

Genotype imputation is now a key component of genomic analyses as it increases the density of available genotypes within a population. However, many factors can influence imputation accuracy. The aim of this study was to assess and compare the accuracy of imputation of high-density genotypes (Affymetrix Axiom Equine genotyping array, 670,806 SNPs) from two moderate-density genotypes (Illumina Equine SNP50 BeadChip, 54,602 SNPs and Illumina Equine SNP70 BeadChip, 65,157 SNPs), using single-breed or multiple-breed reference sets. Genotypes were available from five groups of horse breeds: Arab (AR, 1,207 horses), Trotteur Français (TF, 979 horses), Selle Français (SF, 1,979 horses), Anglo-Arab (AA, 229 horses) and various foreign sport horses (FH, 209 horses). The proportions of horses genotyped with the high-density (HD) chip in each breed group were 10% in AA, 15% in AR and FH, 30% in TF and 57% in SF. A validation set consisting of one-third of the horses genotyped with the HD chip was formed and their genotypes deleted. Two imputation strategies were compared, one in which the reference population consisted only of horses from the same breed group as in the validation set, and another with horses from all breed groups. For the first strategy, concordance rates (CRs) ranged from 97.8% (AR) to 99.0% (TF) and correlations (r²) from 0.94 (AR) to 0.99 (TF). For the second strategy, CR ranged from 97.4% (AR) to 98.9% (TF) and r² from 0.93 (AR) to 0.99 (TF). Overall, the results show a small advantage of within-breed imputation compared with multi-breed imputation. Adding horses from different breed groups to the reference population does not improve the accuracy of imputation. Imputation provides an accurate means of combining data sets from different genotyping platforms, now necessary with the increasing use of the recently developed Affymetrix Axiom Equine genotyping array.

Genome-wide analyses of the Jeju, Thoroughbred, and Jeju crossbred horse populations using the high density SNP array.

The Jeju horse is an indigenous Korean horse breed that is currently registered with the Food and Agriculture Organization of the United Nations. However, there is severe lack of genomic studies on Jeju horse. This study was conducted to investigate genetic characteristics of horses including Jeju horse, Thoroughbred and Jeju crossbred (Jeju × Thoroughbred) populations. We compared the genomes of three horse populations using the Equine SNP70 Beadchip array. Short-range Linkage disequilibrium was the highest in Thoroughbred, whereas r2 values were lowest in Jeju horse. Expected heterozygosity was the highest in Jeju crossbred (0.351), followed by the Thoroughbred (0.337) and Jeju horse (0.311). The level of inbreeding was slightly higher in Thoroughbred (- 0.009) than in Jeju crossbred (- 0.035) and Jeju horse (- 0.038). FST value was the highest between Jeju horse and Thoroughbred (0.113), whereas Jeju crossbred and Thoroughbred showed the lowest value (0.031). The genetic relationship was further assessed by principal component analysis, suggesting that Jeju crossbred is more genetically similar to Thoroughbred than Jeju horse population. Additionally, we detected potential selection signatures, for example, in loci located on LCORL/NCAPG and PROP1 genes that are known to influence body. Genome-wide analyses of the three horse populations showed that all the breeds had somewhat a low level of inbreeding within each population. In the population structure analysis, we found that Jeju crossbred was genetically closer to Thoroughbred than Jeju horse. Furthermore, we identified several signatures of selection which might be associated with traits of interest. To our current knowledge, this study is the first genomic research, analyzing genetic relationships of Jeju horse, Thoroughbred and Jeju crossbred.

Effects of Dietary Arginine Supplementation in Pregnant Mares on Maternal Metabolism and Foal Birthweight

Withers height is an important economic trait for the American Miniature Horses (AMHs) as the breeders' primary objective is to produce small and proportionate animals. Identification of Quantitative Trait Loci (QTLs) affecting height in this breed will result in a better understanding of the genetic architecture and biological pathways contributing to skeletal development. In earlier work, we genotyped 48 horses from 16 different breeds that represent extremes in body size, using the Equine SNP50 bead chip (Illumina Inc). In this study, we applied a dominant model Genome-wide association study (GWAS), a genome-wide fixation index (FST) estimation, as well as a cross-population composite likelihood ratio (XP-CLR) test between the eight large and eight small breeds from this data set. Our GWAS implicated the marker at ECA1:37,676,322 bp, positioned within an intron of the ANKRD1 gene, was significantly associated with height variation. An XP-CLR scan further supported a role in height for the ECA1:37,676,322 bp locus and the FST analysis supported correlations to IGF2BP2 at ECA19:23,815,750. To confirm our findings, we genotyped the candidate loci in an independent set of 89 AMHs. This set verified that the locus at ANKRD1 gene indeed correlates to height under a dominant mode of inheritance, but not the locus at the IGF2BP2 gene. ANKRD1, a transcription factor involved in myocyte growth and differentiation, may contribute to height by influencing the overall growth of the horses. This marker will be a valuable tool for the selection of breeding stock in breeds with height restrictions for registration.

Genetic risk factors for osteochondrosis in various horse breeds.

Osteochondrosis (OC) is an injury to cartilage canals with a following necrosis in the growth cartilage, from there it can develop to osteochondrosis dissecans (OCD). Due to its high impact in the equine industry, new insights into predisposing factors and potential high-risk genetic variants are warranted. This article reviews advancements in quantitative and molecular genetics in refining estimation of genetic parameters and identifying predisposing genetic loci. Heritabilities were highest for hock OC with estimates at 0.29-0.46 in Hanoverian warmblood and Norwegian trotters, whereas in Thoroughbreds only very low genetic variation seemed to be present in hock OC lesions. Whole genome scans using the Illumina Equine SNP50 or SNP70 Beadchip were performed in Thoroughbred, Standardbred, French and Norwegian trotter, Hanoverian and Dutch warmblood. Validation studies in Spanish Purebred and Hanoverian warmblood horses corroborated OC risk loci on ECA 3, 14, 27 and 29. Particularly, a strong association with hock-OCD was found for a single nucleotide polymorphism (SNP) on horse chromosome (ECA) 3 upstream to the LCORL gene. Gene expression and microRNA analyses may be helpful to understand pathophysiological processes in equine OC and to connect OCD-associated genomic regions with potential candidate genes. Furthermore progress in elucidating the underlying genetic variants and pathophysiological changes in OC may be expected from whole genome DNA and RNA next-generation sequencing studies.

Detecting selection signatures on the X chromosome of the Chinese Debao pony.

The X chromosome shows a special interaction between demographic factors and genetic variation, and the analysis of X-linked genomic variation can therefore provide insights into the unique effects of demography and selection on the horse genome that cannot be readily detected by autosomal markers. Debao (DB) ponies have experienced intense selective pressure for the development of their small stature (<106cm at adult height). To identify selective sweeps on the X chromosome of the DB pony, we performed a genome-wide scan of three Chinese horse breeds using an Equine SNP70 BeadChip. Using Yili and Mongolian horses (>134cm at adult height) as reference groups, both FST and XP-EHH revealed that five regions on the X chromosome were under strong selection, resulting in 95 overlapping genes. Seven of these genes, SMS, PHEX, ACSL4, CHRDL1, CACNA1F, DKC1 and CDKL5, are involved in bone development, growth hormone secretion and fat deposition. The region showing the strongest selection pressure was located at the position of 86.6-87.5Mb. The subsequent genome-wide association analysis of the adult height of three Chinese horse breeds detected the two most significant SNPs in the same region, and these two SNPs overlapped with the gene CHRDL1. As a member of the bone morphogenetic protein (BMP) superfamily, CHRDL1 antagonizes the function of BMP4 and plays an important role in embryonic bone formation and cartilage generation. Our results provide new insights into the X-linked selection in Chinese Debao pony.

A missense variant in the coil1A domain of the keratin 25 gene is associated with the dominant curly hair coat trait (Crd) in horse

BackgroundCurly horses present a variety of curl phenotypes that are associated with various degrees of curliness of coat, mane, tail and ear hairs. Their origin is still a matter of debate and several genetic hypotheses have been formulated to explain the diversity in phenotype, including the combination of autosomal dominant and recessive alleles. Our purpose was to map the autosomal dominant curly hair locus and identify the causal variant using genome-wide association study (GWAS) and whole-genome sequencing approaches.ResultsA GWAS was performed using a Bayesian sparse linear mixed model, based on 51 curly and 19 straight-haired French and North American horses from 13 paternal families genotyped on the Illumina EquineSNP50 BeadChip. A single strong signal was observed on equine chromosome 11, in a region that encompasses the type I keratin gene cluster. This region was refined by haplotype analysis to a segment including 36 genes, among which are 10 keratin genes (KRT-10, -12, -20, -23, -24, -25, -26, -27, -28, -222). To comprehensively identify candidate causal variants within all these genes, whole-genome sequences were obtained for one heterozygous curly stallion and its straight-haired son. Among the four non-synonymous candidate variants identified and validated in the curly region, only variant g.21891160G>A in the KRT25 gene (KRT25:p.R89H) was in perfect agreement with haplotype status in the whole pedigree. Genetic association was then confirmed by genotyping a larger population consisting of 353 horses. However, five discordant curly horses were observed, which carried neither the variant nor the main haplotype associated with curliness. Sequencing of KRT25 for two discordant horses did not identify any other deleterious variant, which suggests locus rather than allelic heterogeneity for the curly phenotype.ConclusionsWe identified the KRT25:p.R89H variant as responsible for the dominant curly trait, but a second dominant locus may also be involved in the shape of hairs within North American Curly horses.

Genome-Wide Scans Reveal a Quantitative Trait Locus for Withers Height in Horses Near the ANKRD1 Gene

Withers height is an important economic trait for the American Miniature Horses (AMHs) as the breeders' primary objective is to produce small and proportionate animals. Identification of Quantitative Trait Loci (QTLs) affecting height in this breed will result in a better understanding of the genetic architecture and biological pathways contributing to skeletal development. In earlier work, we genotyped 48 horses from 16 different breeds that represent extremes in body size, using the Equine SNP50 bead chip (Illumina Inc). In this study, we applied a dominant model Genome-wide association study (GWAS), a genome-wide fixation index (FST) estimation, as well as a cross-population composite likelihood ratio (XP-CLR) test between the eight large and eight small breeds from this data set. Our GWAS implicated the marker at ECA1:37,676,322 bp, positioned within an intron of the ANKRD1 gene, was significantly associated with height variation. An XP-CLR scan further supported a role in height for the ECA1:37,676,322 bp locus and the FST analysis supported correlations to IGF2BP2 at ECA19:23,815,750. To confirm our findings, we genotyped the candidate loci in an independent set of 89 AMHs. This set verified that the locus at ANKRD1 gene indeed correlates to height under a dominant mode of inheritance, but not the locus at the IGF2BP2 gene. ANKRD1, a transcription factor involved in myocyte growth and differentiation, may contribute to height by influencing the overall growth of the horses. This marker will be a valuable tool for the selection of breeding stock in breeds with height restrictions for registration.

A genome-wide association study for body weight in Japanese Thoroughbred racehorses clarifies candidate regions on chromosomes 3, 9, 15, and 18.

ABSTRACTBody weight is an important trait to confirm growth and development in humans and animals. In Thoroughbred racehorses, it is measured in the postnatal, training, and racing periods to evaluate growth and training degrees. The body weight of mature Thoroughbred racehorses generally ranges from 400 to 600 kg, and this broad range is likely influenced by environmental and genetic factors. Therefore, a genome-wide association study (GWAS) using the Equine SNP70 BeadChip was performed to identify the genomic regions associated with body weight in Japanese Thoroughbred racehorses using 851 individuals. The average body weight of these horses was 473.9 kg (standard deviation: 28.0) at the age of 3, and GWAS identified statistically significant SNPs on chromosomes 3 (BIEC2_808466, P=2.32E-14), 9 (BIEC2_1105503, P=1.03E-7), 15 (BIEC2_322669, P=9.50E-6), and 18 (BIEC2_417274, P=1.44E-14), which were associated with body weight as a quantitative trait. The genomic regions on chromosomes 3, 9, 15, and 18 included ligand-dependent nuclear receptor compressor-like protein (LCORL), zinc finger and AT hook domain containing (ZFAT), tribbles pseudokinase 2 (TRIB2), and myostatin (MSTN), respectively, as candidate genes. LCORL and ZFAT are associated with withers height in horses, whereas MSTN affects muscle mass. Thus, the genomic regions identified in this study seem to affect the body weight of Thoroughbred racehorses. Although this information is useful for breeding and growth management of the horses, the production of genetically modified animals and gene doping (abuse/misuse of gene therapy) should be prohibited to maintain horse racing integrity.

Preliminary analysis of utilization of genomic relationship in mating plan of Old Kladruber horse

The study analyzed 48 Old Kladruber horses genotyped by Illumina Equine SNP70 BeadChip for usefulness of genomic data in determining of mating plan. Totally 12 variants of data filtering and their impact on calculations in dependence of different parameters of GenCall Score, Minor Allele Frequency and assumed average values of loci of ancestors (l) was investigated. For possibility of comparison between genomic and commonly evaluated relationships, pedigree based relationship matrix was constructed and subsequently subtraction of pedigree from genomic matrix was performed. All matrices were thoroughly inspected and most suitable setting of parameters was chosen. Evaluation of genomic relationships can be successfully implemented in more precise method of mating plan design of Old Kladruber horses. Further genotyping and development of method for rescaling of differences between genomic and pedigree relationship matrices´ elements is advised for a purpose of better interpretation of results by breeders.

Genome-wide association study for semen quality traits in German Warmblood stallions

We performed a genome-wide association study for semen quality traits in 139 German Warmblood stallions. Stallions were genotyped using the Illumina equine SNP50 Beadchip. Traits analysed were de-regressed estimated breeding values (EBVs) for gel-free volume, sperm concentration, total number of sperm, progressive motility and the total number of progressively motile sperm. The GWAS revealed 29 SNPs on 12 different chromosomes as genome-wide significantly associated with semen quality traits. For ten genomic regions we could retrieve candidate genes influencing stallion fertility. Among the candidate genes, we could find the genes encoding cysteine-rich secretory proteins (CRISP1, CRISP2 and CRISP3). This was the first GWAS in horses performed for semen quality traits.

Genome-wide association mapping of heritable temperament variation in the Tennessee Walking Horse.

Temperament is a key criterion in the selection of horses for both leisure and competitive riding to ensure optimal performance and safety. The Tennessee Walking Horse (TWH) is described as a calm, docile breed and is often used as a trail, show and pleasure horse. However, among horse owners and caretakers, there are anecdotes supporting familial and disciplinal typical behaviors and personalities. To investigate the contribution of genetics to temperament, we collected a behavior questionnaire, brief training history and identifying information for 276 TWH, as well as blood or hair samples for DNA. Factor analysis was conducted on the 20-item questionnaire for the set of 216 horses that met inclusion thresholds. Factor analysis identified four temperament factors in TWH: 'anxious', 'tractable', 'agonistic' and 'gregarious'. These four factors account for 64% of the total trait variance. DNA from 113 TWHs were selected and genotyped using the Equine SNP70 bead chip for three separate genome-wide association studies (GWAs) using the factor 1-anxious, factor 2-tractable and factor 3-agonistic scores as the phenotype. Quantitative association analysis identified significant candidate loci for each factor that warrant further investigation.

Loci impacting polymorphic gait in the Tennessee Walking Horse.

Following domestication, man selected the horse primarily for the purpose of transportation rather than consumption; this selective strategy created divergent traits for locomotion. At intermediate speeds, beyond the flat walk, the horse can perform a range of diagonal and lateral 2-beat or 4-beat gait patterns. The Tennessee Walking Horse (TWH) is the only U.S. breed able to perform an even-timed 4-beat gait (the "running-walk") at intermediate speeds; however, within the breed, there is remaining variation in gait type. To investigate the contribution of genetics to this unique trait, blood or hair samples for DNA and gait information were collected from 129 TWH and genotyping was performed at approximately 60,000 loci using the Illumina Equine SNP70 beadchip at GeneSeek Inc. (Lincoln, NE). Case-control association tests identified suggestive regions for gait type on equine chromosome (ECA) 19 (-value of 1.50 × 10 after 1 million permutations; PLINK version 1.07). Haplotype analysis identified 2 significant haplotypes on ECA19 and ECA11 (-values of 3.7 × 10 and 3.92 × 10, respectively). Genes within these suggestive regions play roles in developmental processes and biological regulation, indicating there may be variant differences in the neurobiology and regulation of horses with a polymorphic gait.