Genetic diversity and signatures of selection in Icelandic horses and Exmoor ponies.

BACKGROUND: The Icelandic horse and Exmoor pony are ancient, native breeds, adapted to harsh environmental conditions and they have both undergone severe historic bottlenecks. However, in modern days, the selection pressures on these breeds differ substantially. The aim of this study was to assess genetic diversity in both breeds through expected (HE) and observed heterozygosity (HO) and effective population size (Ne). Furthermore, we aimed to identify runs of homozygosity (ROH) to estimate and compare genomic inbreeding and signatures of selection in the breeds. RESULTS: HO was estimated at 0.34 and 0.33 in the Icelandic horse and Exmoor pony, respectively, aligning closely with HE of 0.34 for both breeds. Based on genomic data, the Ne for the last generation was calculated to be 125 individuals for Icelandic horses and 42 for Exmoor ponies. Genomic inbreeding coefficient (FROH) ranged from 0.08 to 0.20 for the Icelandic horse and 0.12 to 0.27 for the Exmoor pony, with the majority of inbreeding attributed to short ROHs in both breeds. Several ROH islands associated with performance were identified in the Icelandic horse, featuring target genes such as DMRT3, DOCK8, EDNRB, SLAIN1, and NEURL1. Shared ROH islands between both breeds were linked to metabolic processes (FOXO1), body size, and the immune system (CYRIB), while private ROH islands in Exmoor ponies were associated with coat colours (ASIP, TBX3, OCA2), immune system (LYG1, LYG2), and fertility (TEX14, SPO11, ADAM20). CONCLUSIONS: Evaluations of genetic diversity and inbreeding reveal insights into the evolutionary trajectories of both breeds, highlighting the consequences of population bottlenecks. While the genetic diversity in the Icelandic horse is acceptable, a critically low genetic diversity was estimated for the Exmoor pony, which requires further validation. Identified signatures of selection highlight the differences in the use of the two breeds as well as their adaptive trait similarities. The results provide insight into genomic regions under selection pressure in a gaited performance horse breed and various adaptive traits in small-sized native horse breeds. This understanding contributes to preserving genetic diversity and population health in these equine populations.

Using high-density SNP data to unravel the origin of the Franches-Montagnes horse breed.

BACKGROUND: The Franches-Montagnes (FM) is the last native horse breed of Switzerland, established at the end of the 19th century by cross-breeding local mares with Anglo-Norman stallions. We collected high-density SNP genotype data (Axiom™ 670 K Equine genotyping array) from 522 FM horses, including 44 old-type horses (OF), 514 European Warmblood horses (WB) from Sweden and Switzerland (including a stallion used for cross-breeding in 1990), 136 purebred Arabians (AR), 32 Shagya Arabians (SA), and 64 Thoroughbred (TB) horses, as introgressed WB stallions showed TB origin in their pedigrees. The aim of the study was to ascertain fine-scale population structures of the FM breed, including estimation of individual admixture levels and genomic inbreeding (FROH) by means of Runs of Homozygosity. RESULTS: To assess fine-scale population structures within the FM breed, we applied a three-step approach, which combined admixture, genetic contribution, and FROH of individuals into a high-resolution network visualization. Based on this approach, we were able to demonstrate that population substructures, as detected by model-based clustering, can be either associated with a different genetic origin or with the progeny of most influential sires. Within the FM breed, admixed horses explained most of the genetic variance of the current breeding population, while OF horses only accounted for a small proportion of the variance. Furthermore, we illustrated that FM horses showed high TB admixture levels and we identified inconsistencies in the origin of FM horses descending from the Arabian stallion Doktryner. With the exception of WB, FM horses were less inbred compared to the other breeds. However, the relatively few but long ROH segments suggested diversity loss in both FM subpopulations. Genes located in FM- and OF-specific ROH islands had known functions involved in conformation and behaviour, two traits that are highly valued by breeders. CONCLUSIONS: The FM remains the last native Swiss breed, clearly distinguishable from other historically introgressed breeds, but it suffered bottlenecks due to intensive selection of stallions, restrictive mating choices based on arbitrary definitions of pure breeding, and selection of rare coat colours. To preserve the genetic diversity of FM horses, future conservation managements strategies should involve a well-balanced selection of stallions (e.g., by integrating OF stallions in the FM breeding population) and avoid selection for rare coat colours.

Comparative transcriptome analysis of slow-twitch and fast-twitch muscles in Kazakh horses.

This study conducted a thorough analysis of the myofiber type composition in the extensor digitorum longus muscle (EDL) and soleus muscle (SOL) of Kazakh horses, across different genders (male and female). The results showed significant differences in myofiber type composition between EDL and SOL, with a higher proportion of Type I fibers in SOL muscles and a greater prevalence of Type II fibers in EDL muscles. Additionally, the myofiber diameter in Kazakh horses was relatively small, potentially related to the tenderness and edible quality of their muscles. Using high-throughput sequencing technology, we constructed 32 cDNA sequencing libraries and obtained high-quality read data. Gene expression analysis revealed 278 and 372 differentially expressed genes (DEGs) in EDL and SOL muscles, respectively, including genes related to muscle contraction, metabolism, and development. Intersection analysis of DEGs between genders showed that 60 DEGs were significantly different in both male and female horses. GO annotation and KEGG analysis further elucidated the roles of these DEGs in muscle structure, function, and cellular signaling. Protein-protein interaction (PPI) network analysis and identification of hub genes provided new insights into the molecular mechanisms underlying muscle growth and development. Finally, the reliability of the DEGs data was validated through quantitative real-time PCR (qRT-PCR). This study not only enhances our understanding of the biological characteristics of horse muscles but also provides potential molecular targets for improving horse muscle performance and health.

CRISPR-Cas guide RNA indel analysis using CRISPResso2 with Nanopore sequencing data.

OBJECTIVE: Insertion and deletion (indel) analysis of CRISPR-Cas guide RNAs (gRNAs) is crucial in gene editing to assess gRNA efficiency and indel frequency. This study evaluates the utility of CRISPResso2 with Oxford Nanopore sequencing data (nCRISPResso2) for gRNA indel screening, compared to two common Sanger sequencing-based methods, TIDE and ICE. To achieve this, sheep and horse fibroblasts were transfected with Cas9 and a gRNA targeting the myostatin (MSTN) gene. DNA was subsequently extracted, and PCR products exceeding 600 bp were sequenced using both Sanger and Nanopore sequencing. Indel profiling was then conducted using TIDE, ICE, and nCRISPResso2. RESULTS: Comparison revealed close correspondence in indel formation among methods. For the sheep MSTN gRNA, indel percentages were 52%, 58%, and 64% for TIDE, ICE, and nCRISPResso2, respectively. Horse MSTN gRNA showed 81%, 87%, and 86% edited amplicons for TIDE, ICE, and nCRISPResso2. The frequency of each type of indel was also comparable among the three methods, with nCRISPResso2 and ICE aligning the closest. nCRISPResso2 offers a viable alternative for CRISPR-Cas gRNA indel screening, especially with large amplicons unsuitable for Illumina sequencing. CRISPResso2's compatibility with Nanopore data enables cost-effective and efficient indel profiling, yielding results comparable to common Sanger sequencing-based methods.

DNA Methylation and Its Effects on TRIM29 Gene Expression in the Equine Sarcoid Tissue.

Sarcoids are the most frequently diagnosed dermatological tumour in horses. It is a disease that can affect various species of equids, such as donkeys, mules and zebras. This type of tumour can develop in all horse breeds, regardless of age and gender. Treatment options depend on many factors, such as the type of lesion, location, extent, owner preference and financial considerations. In the present study, we investigated the TRIM29 expression, the methylation status of its first exon and its involvement in the formation of equine sarcoids. Bisulfite sequencing PCR (BSP) was used to determine DNA methylation at CpG sites and real-time quantitative polymerase chain reaction (qPCR) was used to detect TRIM29 expression level. Our results showed that TRIM29 is significantly downregulated in lesional samples (FC = -3.72; p < 0.001). Furthermore, TRIM29 expression was significantly correlated (R = -0.73; p < 0.001) with hypermethylation of its specific CpG sites in the first exon of this gene. Our research has demonstrated that the identification of increased methylation of CpG sequences in horse sarcoids, along with the decreased expression of the TRIM29 gene, is an important step towards understanding the molecular mechanisms underlying the disease. These findings can serve in the future as a diagnostic biomarker for horse sarcoids and help in detecting the disease.

An endothelial regulatory module links blood pressure regulation with elite athletic performance.

The control of transcription is crucial for homeostasis in mammals. A previous selective sweep analysis of horse racing performance revealed a 19.6 kb candidate regulatory region 50 kb downstream of the Endothelin3 (EDN3) gene. Here, the region was narrowed to a 5.5 kb span of 14 SNVs, with elite and sub-elite haplotypes analyzed for association to racing performance, blood pressure and plasma levels of EDN3 in Coldblooded trotters and Standardbreds. Comparative analysis of human HiCap data identified the span as an enhancer cluster active in endothelial cells, interacting with genes relevant to blood pressure regulation. Coldblooded trotters with the sub-elite haplotype had significantly higher blood pressure compared to horses with the elite performing haplotype during exercise. Alleles within the elite haplotype were part of the standing variation in pre-domestication horses, and have risen in frequency during the era of breed development and selection. These results advance our understanding of the molecular genetics of athletic performance and vascular traits in both horses and humans.

Non-Synonymous Substitutions in Cadherin 13, Solute Carrier Family 6 Member 4, and Monoamine Oxidase A Genes are Associated with Personality Traits in Thoroughbred Horses.

Retraining retired racehorses for various purposes can help correct behavioral issues. However, ensuring efficiency and preventing accidents present global challenges. Based on the hypothesis that a simple personality assessment could help address these challenges, the present study aimed to identify genetic markers associated with personality. Eight genes were selected from 18 personality-related candidate genes that are orthologs of human personality genes, and their association with personality was verified based on actual behavior. A total of 169 Thoroughbred horses were assessed for their tractability (questionnaire concerning tractability in 14 types of situations and 3 types of impressions) during the training process. Personality factors were extracted from the data using principal component analysis and analyzed for their association with single nucleotide variants as non-synonymous substitutions in the target genes. Three genes, CDH13, SLC6A4, and MAOA, demonstrated significant associations based on simple linear regression, marking the identification of these genes for the first time as contributors to temperament in Thoroughbred horses. All these genes, as well as the previously identified HTR1A, are involved in the serotonin neurotransmitter system, suggesting that the tractability of horses may be correlated with their social personality. Assessing the genotypes of these genes before retraining is expected to prevent problems in the development of a racehorse's second career and shorten the training period through individual customization of training methods, thereby improving racehorse welfare.

Genomic analyses of withers height and linear conformation traits in German Warmblood horses using imputed sequence-level genotypes.

BACKGROUND: Body conformation, including withers height, is a major selection criterion in horse breeding and is associated with other important traits, such as health and performance. However, little is known about the genomic background of equine conformation. Therefore, the aim of this study was to use imputed sequence-level genotypes from up to 4891 German Warmblood horses to identify genomic regions associated with withers height and linear conformation traits. Furthermore, the traits were genetically characterised and putative causal variants for withers height were detected. RESULTS: A genome-wide association study (GWAS) for withers height confirmed the presence of a previously known quantitative trait locus (QTL) on Equus caballus (ECA) chromosome 3 close to the LCORL/NCAPG locus, which explained 16% of the phenotypic variance for withers height. An additional significant association signal was detected on ECA1. Further investigations of the region on ECA3 identified a few promising candidate causal variants for withers height, including a nonsense mutation in the coding sequence of the LCORL gene. The estimated heritability for withers height was 0.53 and ranged from 0 to 0.34 for the conformation traits. GWAS identified significantly associated variants for more than half of the investigated conformation traits, among which 13 showed a peak on ECA3 in the same region as withers height. Genetic parameter estimation revealed high genetic correlations between these traits and withers height for the QTL on ECA3. CONCLUSIONS: The use of imputed sequence-level genotypes from a large study cohort led to the discovery of novel QTL associated with conformation traits in German Warmblood horses. The results indicate the high relevance of the QTL on ECA3 for various conformation traits, including withers height, and contribute to deciphering causal mutations for body size in horses.

Ancient Mitochondrial Genomes Provide New Clues in the History of the Akhal-Teke Horse in China.

This study analyzed ancient DNA from the remains of horses unearthed from the Shihuyao tombs. These were found to date from the Han and Tang Dynasties in Xinjiang (approximately 2200 to 1100 years ago). Two high-quality mitochondrial genomes were acquired and analyzed using next-generation sequencing. The genomes were split into two maternal haplogroups, B and D, according to a study that included ancient and contemporary samples from Eurasia. A close genetic affinity was observed between the horse of the Tang Dynasty and Akhal-Teke horses according to the primitive horse haplotype G1. Historical evidence suggests that the ancient Silk Road had a vital role in their dissemination. Additionally, the matrilineal history of the Akhal-Teke horse was accessed and suggested that the early domestication of the breed was for military purposes.

Widespread horse-based mobility arose around 2200 BCE in Eurasia.

Horses revolutionized human history with fast mobility1. However, the timeline between their domestication and their widespread integration as a means of transport remains contentious2-4. Here we assemble a collection of 475 ancient horse genomes to assess the period when these animals were first reshaped by human agency in Eurasia. We find that reproductive control of the modern domestic lineage emerged around 2200 BCE, through close-kin mating and shortened generation times. Reproductive control emerged following a severe domestication bottleneck starting no earlier than approximately 2700 BCE, and coincided with a sudden expansion across Eurasia that ultimately resulted in the replacement of nearly every local horse lineage. This expansion marked the rise of widespread horse-based mobility in human history, which refutes the commonly held narrative of large horse herds accompanying the massive migration of steppe peoples across Europe around 3000 BCE and earlier3,5. Finally, we detect significantly shortened generation times at Botai around 3500 BCE, a settlement from central Asia associated with corrals and a subsistence economy centred on horses6,7. This supports local horse husbandry before the rise of modern domestic bloodlines.

Mitochondrial DNA and Y chromosome reveal the genetic structure of the native Polish Konik horse population.

Polish Konik remains one of the most important horse breeds in Poland. The primitive, native horses with a stocky body and mouse-like coat color are protected by a conservation program, while their Polish population consists of about 3,480 individuals, representing 16 dam and six sire lines. To define the population's genetic structure, mitochondrial DNA and Y chromosome sequence variables were identified. The mtDNA whole hypervariable region analysis was carried out using the Sanger sequencing method on 233 Polish Koniks belonging to all dam lines, while the Y chromosome analysis was performed with the competitive allele-specific PCR genotyping method on 36 horses belonging to all sire lines. The analysis of the mtDNA hypervariable region detected 47 SNPs, which assigned all tested horses to 43 haplotypes. Most dam lines presented more than one haplotype; however, five dam lines were represented by only one haplotype. The haplotypes were classified into six (A, B, E, J, G, R) recognized mtDNA haplogroups, with most horses belonging to haplogroup A, common among Asian horse populations. Y chromosome analysis allocated Polish Koniks in the Crown group, condensing all modern horse breeds, and divided them into three haplotypes clustering with coldblood breeds (28 horses), warmblood breeds (two horses), and Duelmener Pony (six horses). The clustering of all Wicek sire line stallions with Duelmener horses may suggest a historical relationship between the breeds. Additionally, both mtDNA and Y chromosome sequence variability results indicate crossbreeding before the studbooks closure or irregularities in the pedigrees occurred before the DNA testing introduction.

Genomic tools for early selection among Thoroughbreds and Polo Argentino horses for practicing polo.

The Polo Argentino (PA) horse is a recognized breed, developed originally by mixing crossbred and Thoroughbred (TB) horses to play polo. Early PA selection is difficult due to unreliable performance estimations. This study investigated the usefulness of genomic markers previously linked to morphological and functional traits as a tool for the early selection of PA. To this, we genotyped 520 PA and 30 TB horses using the Equine GGPArray (Illumina, n = 71,778 SNPs). Analyses included a genetic characterization of six genetic markers associated with behavioral (DRD4), muscular development (MSTN), and body size (LCORL, HMGA6, ZFAT, and LASP1) genes. Genetic differences in the DRD4, MSTN, and LCORL SNP were found between the two breeds, in the last two FST index between breeds was 0.13 and 0.6, respectively (p < 0.01). In DRD4, G allele was the more prevalent in PA (0.56 vs 0.45 in TB, p < 0.05), but no differences were observed between the genotypes associated with phenotypes. In MSTN, heterozygous genotypes were the most common in PA (48 %), with a significant decrease in AA (Hardy-Weinberg p < 0.05), suggesting a negative selection against it in polo horses. In body size, HMGA2 was monomorphic in all horses, while ZFAT and LASP1 SNP showed higher variability. Interestingly, 99 % of PA showed a TT genotype in LCORL (only 66 % in TB), demonstrating selection for smaller horses. Our results suggest that empirical selection in PA has generated an incipient genomic differentiation in discrete traits which could be used as a marker-assisted selection tool for early selection of polo horses.

The genome of Przewalski's horse (Equus ferus przewalskii).

The Przewalski's horse (Equus ferus przewalskii) is an endangered equid native to the steppes of central Asia. After becoming extinct in the wild multiple conservation efforts convened to preserve the species, including captive breeding programs, reintroduction and monitoring systems, protected lands, and cloning. Availability of a highly contiguous reference genome is essential to support these continued efforts. We used Oxford Nanopore sequencing to produce a scaffold-level 2.5 Gb nuclear assembly and 16,002 bp mitogenome from a captive Przewalski's mare. All assembly drafts were generated from 111 Gb of sequence from a single PromethION R10.4.1 flow cell. The mitogenome contained 37 genes in the standard mammalian configuration and was 99.63% identical to the domestic horse (Equus caballus). The nuclear assembly, EquPr2, contained 2,146 scaffolds with an N50 of 85.1 Mb, 43X mean depth, and BUSCO quality score of 98.92%. EquPr2 successfully improves upon the existing Przewalski's horse reference genome (Burgud), with 25-fold fewer scaffolds, a 166-fold larger N50, and phased pseudohaplotypes. Modified basecalls revealed 79.5% DNA methylation and 2.1% hydroxymethylation globally. Allele-specific methylation analysis between pseudohaplotypes revealed 226 differentially methylated regions in known imprinted genes and loci not previously reported as imprinted. The heterozygosity rate of 0.165% matches previous estimates for the species and compares favorably to other endangered animals. This improved Przewalski's horse assembly will serve as a valuable resource for conservation efforts and comparative genomics investigations.

De novo transcriptome assembly database for 100 tissues from each of seven species of domestic herbivore.

Domesticated herbivores are an important agricultural resource that play a critical role in global food security, particularly as they can adapt to varied environments, including marginal lands. An understanding of the molecular basis of their biology would contribute to better management and sustainable production. Thus, we conducted transcriptome sequencing of 100 to 105 tissues from two females of each of seven species of herbivore (cattle, sheep, goats, sika deer, horses, donkeys, and rabbits) including two breeds of sheep. The quality of raw and trimmed reads was assessed in terms of base quality, GC content, duplication sequence rate, overrepresented k-mers, and quality score distribution with FastQC. The high-quality filtered RNA-seq raw reads were deposited in a public database which provides approximately 54 billion high-quality paired-end sequencing reads in total, with an average mapping rate of ~93.92%. Transcriptome databases represent valuable resources that can be used to study patterns of gene expression, and pathways that are related to key biological processes, including important economic traits in herbivores.

Different expression patterns of DNA methyltransferases during horse testis development.

DNA methyltransferases (DNMTs) are important epigenetic modification during spermatogenesis. To further evaluate the pattern of DNMTs in horse testes during development, we investigated the expression and localization of DNMT1, DNMT3a and DNMT3b at different time points. The qRT-PCR results showed that DNMT1 expression was maintained in testes tissue from 6-month-old (0.5y) to 2-year-old (2y) of age and decreased after 3-year-old (3y) (P < 0.01). The expression levels of DNMT3a and DNMT3b peaked in testes tissue at 3y (P < 0.01). At 4-year-old (4y), the expression of DNMT3a and DNMT3b was decreased and became similar to that at 0.5y. Immunofluorescence of DNMT1, DNMT3a and DNMT3b on testis samples confirmed the differential expression and localization of these three DNA methylation transferases during horse development. Further molecular biological studies are needed to understand the implications of the expression patterns of these DNMTs in horse testes.

The monsoon-associated equine South African pointy mosquito 'Aedes caballus'; the first comprehensive record from southeastern Iran with a description of ecological, morphological, and molecular aspects.

The equine South African pointy vector mosquito, Aedes caballus, poses a significant threat to human health due to its capacity for transmitting arboviruses. Despite favorable climate for its existence in southeast Iran, previous records of this species in the area have indicated very low abundance. This comprehensive field and laboratory study aimed to assess its current adult population status in this region, utilizing a combination of ecological, morphological and molecular techniques. Four distinct types of traps were strategically placed in three fixed and two variable mosquito sampling sites in the southern strip of Sistan and Baluchistan Province. Subsequently, DNA was extracted from trapped mosquitoes and subjected to PCR amplification using the molecular markers COI, ITS2, and ANT. In total, 1734 adult Ae. caballus specimens were collected from rural areas, with the majority being captured by CO2-baited bednet traps. A notable increase in the abundance of this species was observed following rainfall in February. The genetic analysis revealed multiple haplotypes based on COI and ITS2 sequences, with COI gene divergence at 0.89%, and ITS2 sequence divergence at 1.6%. This suggests that previous challenges in morphological identification may have led to misidentifications, with many adults previously classified as Ae. vexans potentially being Ae. caballus. The findings of this study hold significant implications for public health authorities, providing valuable insights for integrated and targeted vector control and disease management efforts.

Double-Tube Multiplex TaqMan Real-Time PCR for the Detection of Eight Animal-Derived Dairy Ingredients.

Milk and dairy products represent important sources of nutrition in our daily lives. The identification of species within dairy products holds importance for monitoring food adulteration and ensuring traceability. This study presented a method that integrated double-tube and duplex real-time polymerase chain reaction (PCR) with multiplex TaqMan probes to enable the high-throughput detection of animal-derived ingredients in milk and dairy products. The detection system utilized one pair of universal primers, two pairs of specific primers, and eight animal-derived specific probes for cow, buffalo, goat, sheep, camel, yak, horse, and donkey. These components were optimized within a double-tube and four-probe PCR multiplex system. The developed double-tube detection system could simultaneously identify the above eight targets with a detection limit of 10-0.1 pg/µL. Validation using simulated adulterated milk samples demonstrated a detection limit of 0.1%. The primary advantage of this method lies in the simplification of the multiplex quantitative real-time PCR (qPCR) system through the use of universal primers. This method provides an efficient approach for detecting ingredients in dairy products, providing powerful technical support for market supervision.

Molecular cytogenetic screening of sex chromosome abnormalities in young horse populations.

BACKGROUND: Chromosomal abnormalities occur in the equine population at a rate of approximately 2%. The use of molecular cytogenetic techniques allows a more accurate identification of chromosomal abnormalities, especially those with a low rate of abnormal metaphases, demonstrating that the actual incidence in equine populations is higher. OBJECTIVES: Estimation of the number of carriers of karyotypic abnormalities in a sample from a population of young horses of various breeds, using molecular cytogenetic techniques. STUDY DESIGN: Cross-sectional. METHODS: Venous blood samples were collected from 500 young horses representing 5 breeds (Purebred Arabian, Hucul, Polish primitive horse [Konik], Malopolska, Coldblood, Silesian). Chromosomes and DNA were obtained from blood lymphocytes and evaluated by fluorescence in situ hybridisation (FISH) and PCR, using probes and markers for the sex chromosomes and select autosomes. RESULTS: Nineteen horses, 18 mares and 1 stallion, were diagnosed with different chromosomal abnormalities: 17 cases of mosaic forms of sex chromosome aneuploidies with a very low incidence (0.6%-4.7%), one case of a SRY-negative 64,XY sex reversal mare, and one mare with X-autosome translocation. The percentage of sex chromosomal aberrations was established as 3.8% in the whole population, 6.08% in females and 0.49% in males. MAIN LIMITATIONS: Limited sample size, confined to horses from Poland. CONCLUSIONS: The rate of sex chromosomal abnormalities we identified was almost double that reported in previous population studies that used classical chromosome staining techniques. FISH allowed the detection of aneuploid cell lines which had a very low incidence. The FISH technique is a faster and more precise method for karyotype examination; however, it is usually focused on only one or two chromosomes while banding karyotyping includes the entire chromosome set.

Investigating age-related differences in muscles of Kazakh horse through transcriptome analysis.

This study conducted transcriptome sequencing on the skeletal muscles of three different anatomical locations across various growth stages to investigate the impact of ages on crucial candidate genes and molecular mechanisms associated with muscle development in Kazakh horses. Sixteen Kazakh horses were selected, and they were divided into four age groups, each with four biological replicates. Tissue samples from the longest dorsal muscle, abdominal muscle, and diaphragm muscle were collected for analysis. The results revealed differential mRNA expression in the longest dorsal muscle between the eight-month group (Group O) and the 10-year group (Group F), with 434 up-regulated and 322 down-regulated genes. In the abdominal muscle, there were 125 up-regulated and 127 down-regulated genes, while in the diaphragm muscle, there were 73 up-regulated and 70 down-regulated genes. In this study, GO enrichment analysis focused on biological processes. KEGG pathway analysis highlighted the Oxidative Phosphorylation pathway for the longest dorsal muscle, annotating 37 differentially expressed genes (DEGs), including ATP5PF, NDUFB8, and ATP5MG, all of which were down-regulated. For the abdominal muscle, the ECM-receptor interaction pathway was enriched, annotating 7 DEGs such as COL4A2, COL4A1, and ITGA5. In the diaphragm muscle, the Hippo signaling pathway was enriched, annotating 6 DEGs, including SERPINE1, RASSF1, and FZD10. This study provides robust data support and a theoretical foundation for a comprehensive understanding of the influence of age on skeletal muscle development in horses.

Population structure and genetic diversity of Polish Arabian horses based on pedigree data.

With a history tracing back to at least the 18th century and a substantial global influence on various breeds, Polish Arabian horse population is of paramount importance for both breeders and conservationists. However, its genetic makeup and the population dynamics are still not well understood. This study presents an analysis of the modern Polish Arabian horse population using pedigree data, focusing on the breed's genetic diversity and population structure. Our analysis encompassed 1 498 individuals defined as the reference population (RP) and their 11 065 ancestors, which resulted in the dataset of 12 254 individuals (total population). We traced their genealogy to assess inbreeding coefficients (F), founder effects, and genetic variability measures such as the effective number of founders (fe), ancestors (fa), or founder genome equivalents (fge). The results indicated a good pedigree quality with an average of 28.1 maximum traced generations, revealing high pedigree completeness for initial generations with a decline beyond the seventh generation. The genetic diversity parameters showed a considerable bottleneck effect, with an effective number of founders at 73, which reflects a substantial loss of genetic diversity over time. Despite the vast total number of founders (852), only a few have had a lasting impact on the current population, signaling the need for revised breeding strategies to maintain diversity. The study identified a slight but consistent rise in inbreeding over the last century, with a marginal recent decline, and a significant difference in the contribution of various founders. The average F was 5.8%, with 99.6% of the reference population being inbred. The analysis of effective population size (Ne) highlighted potential risks for genetic diversity, urging for revision of breeding goals to consider a wider array of founder lineages. The study indicated that stallions belonging to RP can be attributed to 15 distinct sirelines, whereas mares to 45 unique damlines, more than considered in the current breeding program (8 and 15, respectively). Conclusively, the study underlines the need for ongoing monitoring and strategic breeding to maintain and enhance the genetic diversity of Polish Arabians, considering the breed's historical significance and contemporary genetic challenges.