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Archaeological and genetic evidence concerning the time and mode of wild horse (Equus ferus) domestication is still debated. High levels of genetic diversity in horse mtDNA have been detected when analyzing the control region; recurrent mutations, however, tend to blur the structure of the phylogenetic tree. Here, we brought the horse mtDNA phylogeny to the highest level of molecular resolution by analyzing 83 mitochondrial genomes from modern horses across Asia, Europe, the Middle East, and the Americas. Our data reveal 18 major haplogroups (A-R) with radiation times that are mostly confined to the Neolithic and later periods and place the root of the phylogeny corresponding to the Ancestral Mare Mitogenome at ~130-160 thousand years ago. All haplogroups were detected in modern horses from Asia, but F was only found in E. przewalskii--the only remaining wild horse. Therefore, a wide range of matrilineal lineages from the extinct E. ferus underwent domestication in the Eurasian steppes during the Eneolithic period and were transmitted to modern E. caballus breeds. Importantly, now that the major horse haplogroups have been defined, each with diagnostic mutational motifs (in both the coding and control regions), these haplotypes could be easily used to (i) classify well-preserved ancient remains, (ii) (re)assess the haplogroup variation of modern breeds, including Thoroughbreds, and (iii) evaluate the possible role of mtDNA backgrounds in racehorse performance. This research received support from the Italian Ministry of Education, University and Research: Fondo per gli Investimenti della Ricerca di Base (FIRB) Futuro in Ricerca 2008 (to A.A. and A.O.), Progetti Ricerca Interesse Nazionale 2008 (to E.G.) and 2009 (to A.A., O.S., and A.T.); L'ORÉAL Italia per le Donne e la Scienza (to A.O.), Fundação para a Ciência e Tecnologia (FCT), Grant SFRH/BPD/64233/2009 (to P.S.); Consorzio Interuniversitario di Biotecnologie (to M. Santagostino); Fondazione Alma Mater Ticinensis (O.S. and A.T.); and Progetto Ricerca e INNOVAzione nelle attività di miglioramento GENetico animale (INNOVAGEN), Italian Ministry of Agriculture (M. Silvestrelli). Instituto de Patologia e Imunologia Molecular da Universidade do Porto (IPATIMUP) is an Associate Laboratory of the Portuguese Ministry of Science, Technology, and Higher Education and is partially supported by FCT.

Summary A large proportion of mammalian genomes is represented by transposable elements (TE), most of them being long interspersed nuclear elements 1 (LINE-1 or L1). An increased expression of LINE-1 elements may play an important role in cellular stress–related conditions exerting drastic effects on the mammalian transcriptome. To understand the impact of TE on the known horse transcriptome, we masked the horse EST database, pointing out that the amount is consistent with other major vertebrates. A previously developed transcript-derived fragments (TDFs) dataset, deriving from exercise-stimulated horse peripheral blood mononuclear cells (PBMCs), was found to be enriched with L1 (26.8% in terms of bp). We investigated the involvement of TDFs in exercise-induced stress through bioinformatics and gene expression analysis. Results indicate that LINE-derived sequences are not only highly but also differentially expressed during physical effort, hinting at interesting scenarios in the regulation of gene expression in relation to exercise.

It is known that moderate physical activity may have beneficial effects on health, whereas strenuous effort induces a state resembling inflammation. The molecular mechanisms underlying the cellular response to exercise remain unclear, although it is clear that the immune system plays a key role. It has been hypothesized that the physio-pathological condition that develops in athletes subjected to heavy training is caused by derangement of cellular immune regulation. The purpose of the present study was to obtain information on endurance horse gene transcription under strenuous conditions and to identify candidate genes causing immune system derangement. We performed a wide gene expression scan, using microarray technology, on peripheral blood mononuclear cells of ten horses chosen from high-level participants in national and international endurance races. The use of three different timepoints revealed changes in gene expression when post-effort samples (T1, taken immediately after the race; and T2, taken 24 h after the race) were compared with basal sample (T0, at rest). Statistical analysis showed no differences in gene expression between T0 and T2 samples, indicating complete restoration of homeostasis by 24 h after racing, whereas T1 showed strong modulation of expression, affecting 132 genes (97 upregulated, 35 downregulated). Ingenuity pathway analysis revealed that the main mechanisms and biofunctions involved were significantly associated with immunological and inflammatory responses. Real-time PCR was performed on 26 gene products to validate the array data.

Summary The Maremmano is an Italian warmblood horse breed from central Italy. We characterizedthe genetic diversity and the degree of admixture in Maremmano in comparison to 14 otherEuropean horse breeds using 30 microsatellites. Between-breed diversity explained about 9per cent of the total genetic diversity. Cluster analysis, genetic distances and genetic dif-ferentiation coefficients showed a close relationship of Maremmano with Hanoverian andLusitano in accordance with breed history.Keywords clustering analysis, Equus caballus, Maremmano.The Maremmano is an Italian warmblood horse mostly bredin the provinces of Grosseto and Viterbo (Central Italy). It isbelieved that the origin of this horse breed goes back to localEtruscan horse populations which were crossbred withmodern breeds in the last centuries. Pedigree analysis of theMaremmano horses revealed four male lines contributing11.1% to the genetic diversity of all Maremmano stallions(Silvestrelli 1991). In 1980, Maremmano breeders estab-lished a stud book for Maremmano.The objective of this study was to investigate the geneticstructure and the degree of admixture of Maremmano to-gether with the Italian nucleus of Lipizzan and Lusitano aswell as twelve further horse breeds previously characterizedby Aberle et al. (2004). This data set consisted of Hanove-rian, Arabian, Exmoor, Icelandic, Przewalski, Sorraia andall German coldblood horse breeds. We used the samemarker set of 30 autosomal microsatellites and the samePCR conditions and reference samples as described else-where (Aberle et al. 2004).A total of 146 animals that were not closely related weregenotyped in this study: Maremmano (n = 50), the Italiannucleus of Lipizzan (n = 49) and Lusitano (n = 47) fromPortugal. DNA was extracted from EDTA-blood usingstandard methods.Genetic variability of the breeds genotyped in this studywas determined by the mean number of alleles, observedand expected heterozygosities, breed-specific alleles and themolecular variance (

The horse is an optimal model organism for studying the genomic response to exercise-induced stress, due to its natural aptitude for athletic performance and the relative homogeneity of its genetic and environmental backgrounds. Here, we applied RNA-sequencing analysis through the use of SOLiD technology in an experimental framework centered on exercise-induced stress during endurance races in equine athletes. We monitored the transcriptional landscape by comparing gene expression levels between animals at rest and after competition. Overall, we observed a shift from coding to non-coding regions, suggesting that the stress response involves the differential expression of not annotated regions. Notably, we observed significant post-race increases of reads that correspond to repeats, especially the intergenic and intronic L1 and L2 transposable elements. We also observed increased expression of the antisense strands compared to the sense strands in intronic and regulatory regions (1 kb up- and downstream) of the genes, suggesting that antisense transcription could be one of the main mechanisms for transposon regulation in the horse under stress conditions. We identified a large number of transcripts corresponding to intergenic and intronic regions putatively associated with new transcriptional elements. Gene expression and pathway analysis allowed us to identify several biological processes and molecular functions that may be involved with exercise-induced stress. Ontology clustering reflected mechanisms that are already known to be stress activated (e.g., chemokine-type cytokines, Toll-like receptors, and kinases), as well as “nucleic acid binding” and “signal transduction activity” functions. There was also a general and transient decrease in the global rates of protein synthesis, which would be expected after strenuous global stress. In sum, our network analysis points toward the involvement of specific gene clusters in equine exercise-induced stress, including those involved in inflammation, cell signaling, and immune interactions.

In this study, we examine the population structure and inbreeding level of the Italian Lipizzan horse breed by means of AFLP fingerprinting and SSR genotyping. A total of 245 AFLP marker alleles were scored across all horses (n=140) living in the Italian Lipizzan stud in 2007, and microsatellite analysis was performed on all mares (n=42) and stallions (n=7) by genotyping at 30 SSR marker loci. A total of 53 (21.6%) AFLP marker alleles were scored as polymorphic among the animals, with a mean frequency of 0.722. Based on this subset of markers, 123 animals were assigned to three distinct subgroups of 44, 41 and 38 animals, respectively, each one showing a within-subgroup mean Dice's genetic similarity around 86% (standard error

Background Copy number variants (CNVs) have been shown to play an important role in genetic diversity of mammals and in the development of many complex phenotypic traits. The aim of this study was to perform a standard comparative evaluation of CNVs in horses using three different CNV detection programs and to identify genomic regions associated with body size in horses. Results Analysis was performed using the Illumina Equine SNP50 genotyping beadchip for 854 horses. CNVs were detected by three different algorithms, CNVPartition, PennCNV and QuantiSNP. Comparative analysis revealed 50 CNVs that affected 153 different genes mainly involved in sensory perception, signal transduction and cellular components. Genome-wide association analysis for body size showed highly significant deleted regions on ECA1, ECA8 and ECA9. Homologous regions to the detected CNVs on ECA1 and ECA9 have also been shown to be correlated with human height. Conclusions Comparative analysis of CNV detection algorithms was useful to increase the specificity of CNV detection but had certain limitations dependent on the detection tool. GWAS revealed genome-wide associated CNVs for body size in horses.

Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-27T11:28:16Z No. of bitstreams: 0Bitstream added on 2014-05-27T14:32:16Z : No. of bitstreams: 1 2-s2.0-84873867256.pdf: 796974 bytes, checksum: 64f172274307a2abd3e5977743eb1671 (MD5) Made available in DSpace on 2014-05-27T11:28:16Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-01-30 Horses were domesticated from the Eurasian steppes 5,000-6,000 years ago. Since then, the use of horses for transportation, warfare, and agriculture, as well as selection for desired traits and fitness, has resulted in diverse populations distributed across the world, many of which have become or are in the process of becoming formally organized into closed, breeding populations (breeds). This report describes the use of a genome-wide set of autosomal SNPs and 814 horses from 36 breeds to provide the first detailed description of equine breed diversity. FST calculations, parsimony, and distance analysis demonstrated relationships among the breeds that largely reflect geographic origins and known breed histories. Low levels of population divergence were observed between breeds that are relatively early on in the process of breed development, and between those with high levels of within-breed diversity, whether due to large population size, ongoing outcrossing, or large within-breed phenotypic diversity. Populations with low within-breed diversity included those which have experienced population bottlenecks, have been under intense selective pressure, or are closed populations with long breed histories. These results provide new insights into the relationships among and the diversity within breeds of horses. In addition these results will facilitate future genome-wide association studies and investigations into genomic targets of selection. © 2013 Petersen et al. University of Minnesota College of Veterinary Medicine, St Paul, MN Texas AandM University College of Veterinary Medicine and Biomedical Science, College Station, TX Swedish University of Agricultural Sciences Department of Animal Breeding and Genetics, Uppsala University of Kentucky Department of Veterinary Science, Lexington, KY University of California Davis School of Veterinary Medicine, Davis, CA Equine Analysis, Midway, KY University Estadual Paulista Department of Veterinary Clinical Science, Botucatu-SP University College Dublin School of Veterinary Medicine, Dublin University of Azores Institute for Biotechnology and Bioengineering Biotechnology Centre of Azores, Angra do Heroísmo University of Veterinary Medicine Hannover Institute for Animal Breeding and Genetics, Hannover University of Perugia Faculty of Veterinary Medicine, Perugia Animal Health Trust, Lanwades Park, Newmarket, Suffolk French National Institute for Agricultural Research-Animal Genetics and Integrative Biology Unit, Jouy en Josas University of Sydney Veterinary Science, New South Wales Nihon Bioresource College, Koga, Ibaraki University of Helsinki Faculty of Veterinary Medicine, Helsinki University College Dublin College of Agriculture, Food Science and Veterinary Medicine, Belfield, Dublin University of Bern Institute of Genetics, Bern Institute of Cancer Research Breakthrough Breast Cancer Research Centre, London Royal Veterinary College Comparative Neuromuscular Diseases Laboratory, London Swiss National Stud Farm Agroscope Liebefeld-Posieux Research Station, Avenches Norwegian School of Veterinary Science Department of Basic Sciences and Aquatic Medicine, Oslo Animal DNA Diagnostics Ltd, Cambridge Laboratory of Racing Chemistry Department of Molecular Genetics, Utsunomiya, Tochigi

The application of molecular diagnostic techniques along with nucleotide sequence determination to permit contemporary phylogenetic analysis of European field isolates of equine infectious anemia virus (EIAV) has not been widely reported. As a result, of extensive testing instigated following the 2006 outbreak of equine infectious anemia in Italy, 24 farms with a history of exposure to this disease were included in this study. New PCR-based methods were developed, which, especially in the case of DNA preparations from peripheral blood cells, showed excellent correlation with OIE-approved agar gel immunodiffusion (AGID) tests for identifying EIAV-infected animals. In contrast, the OIE-recommended oligonucleotide primers for EIAV failed to react with any of the Italian isolates. Similar results were also obtained with samples from four Romanian farms. In addition, for the first time complete characterization of gag genes from five Italian isolates and one Romanian isolate has been achieved, along with acquisition of extensive sequence information (86% of the total gag gene) from four additional EIAV isolates (one Italian and three Romanian). Furthermore, in another 23 cases we accomplished partial characterization of gag gene sequences in the region encoding the viral matrix protein. Analysis of this information suggested that most Italian isolates were geographically restricted, somewhat reminiscent of the “clades” described for human immunodeficiency virus type 1 (HIV-1). Collectively this represents the most comprehensive genetic study of European EIAV isolates conducted to date.

Background Adequate stress response is a critical factor during athlete horses' training and is central to our capacity to obtain better performances while safeguarding animal welfare. In order to investigate the molecular mechanisms underlying this process, several studies have been conducted that take advantage of microarray and quantitative real-time PCR (qRT-PCR) technologies to analyse the expression of candidate genes involved in the cellular stress response. Appropriate application of qRT-PCR, however, requires the use of reference genes whose level of expression is not affected by the test, by general physiological conditions or by inter-individual variability. Results The expression of nine potential reference genes was evaluated in lymphocytes of ten endurance horses during strenuous exercise. These genes were tested by qRT-PCR and ranked according to the stability of their expression using three different methods (implemented in geNorm, NormFinder and BestKeeper). Succinate dehydrogenase complex subunit A (SDHA) and hypoxanthine phosphoribosyltransferase (HPRT) always ranked as the two most stably expressed genes. On the other hand, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), transferrin receptor (TFRC) and ribosomal protein L32 (RPL32) were constantly classified as the less reliable controls. Conclusion This study underlines the importance of a careful selection of reference genes for qRT-PCR studies of exercise induced stress in horses. Our results, based on different algorithms and analytical procedures, clearly indicate SDHA and HPRT as the most stable reference genes of our pool.