DNA in horses refers to the genetic material that carries the hereditary information necessary for the growth, development, functioning, and reproduction of equine species. It consists of sequences of nucleotides that encode the genetic instructions used in the development and functioning of horses. DNA analysis in horses can provide insights into genetic diversity, lineage, and breed characteristics. It is also utilized in identifying genetic disorders, understanding hereditary traits, and assisting in selective breeding programs. This page compiles peer-reviewed research studies and scholarly articles that explore the structure, function, and applications of DNA analysis in equine genetics and breeding.
Stefaniuk M, Kaczor U, Augustyn R, Gurgul A, Kulisa M, Podstawski Z.Myostatin (GDF-8) encoded by the MSTN gene is a negative regulator of muscle growth and development and belongs to the TGF-β superfamily of secreted growth and differentiation factors. In Thoroughbred horses, an MSTN sequence polymorphism (g.66493737C>T) is associated with optimum race distance. In the present study, a genetic polymorphism of a predicted promoter of the MSTN gene was investigated in 451 horses belonging to five different breeds: Arabian, Thoroughbred, Polish Konik, Hucul and Polish Heavy Draft. Two SNPs located at g.66495826T>C and g.66495696T>C (chr;18 EquCab 2.0) s...
Gambini A, De Stefano A, Bevacqua RJ, Karlanian F, Salamone DF.Embryo aggregation has been demonstrated to improve cloning efficiency in mammals. However, since no more than three embryos have been used for aggregation, the effect of using a larger number of cloned zygotes is unknown. Therefore, the goal of the present study was to determine whether increased numbers of cloned aggregated zygotes results in improved in vitro and in vivo embryo development in the equine. Zona-free reconstructed embryos (ZFRE's) were cultured in the well of the well system in four different experimental groups: I. 1x, only one ZFRE per microwell; II. 3x, three per microwell;...
Fernandes KA, Kittelmann S, Rogers CW, Gee EK, Bolwell CF, Bermingham EN, Thomas DG.The effects of abrupt dietary transition on the faecal microbiota of forage-fed horses over a 3-week period were investigated. Yearling Thoroughbred fillies reared as a cohort were exclusively fed on either an ensiled conserved forage-grain diet ("Group A"; n = 6) or pasture ("Group B"; n = 6) for three weeks prior to the study. After the Day 0 faecal samples were collected, horses of Group A were abruptly transitioned to pasture. Both groups continued to graze similar pasture for three weeks, with faecal samples collected at 4-day intervals. DNA was isolated from the faeces and microbial 16S ...
Schrimpf R, Dierks C, Martinsson G, Sieme H, Distl O.A consistently high level of stallion fertility plays an economically important role in modern horse breeding. We performed a genome-wide association study for estimated breeding values of the paternal component of the pregnancy rate per estrus cycle (EBV-PAT) in Hanoverian stallions. A total of 228 Hanoverian stallions were genotyped using the Equine SNP50 Beadchip. The most significant association was found on horse chromosome 6 for a single nucleotide polymorphism (SNP) within phospholipase C zeta 1 (PLCz1). In the close neighbourhood to PLCz1 is located CAPZA3 (capping protein (actin filam...
Hamond C, Martins G, Bremont S, Medeiros MA, Bourhy P, Lilenbaum W.The purpose of the present study was to detect the presence of DNA of pathogenic Leptospira sp. in vaginal fluids of mares regarding a possible role of the sexual transmission. A total of 134 breeding mares from four troops were studied and sampling was conducted from vaginal fluids and urine for culture and PCR; and blood for serology. From the 134 serum samples tested, 59 (44%) were seroreactive, and serovar Bratislava was the most frequent (54.2%). None positive culture was obtained, but leptospiral DNA was detected by PCR (lipL32 gene) in 45 (33.5%) urine samples and 43 (32%) vaginal fluid...
Ghosh S, Qu Z, Das PJ, Fang E, Juras R, Cothran EG, McDonell S, Kenney DG, Lear TL, Adelson DL, Chowdhary BP, Raudsepp T.We constructed a 400K WG tiling oligoarray for the horse and applied it for the discovery of copy number variations (CNVs) in 38 normal horses of 16 diverse breeds, and the Przewalski horse. Probes on the array represented 18,763 autosomal and X-linked genes, and intergenic, sub-telomeric and chrY sequences. We identified 258 CNV regions (CNVRs) across all autosomes, chrX and chrUn, but not in chrY. CNVs comprised 1.3% of the horse genome with chr12 being most enriched. American Miniature horses had the highest and American Quarter Horses the lowest number of CNVs in relation to Thoroughbred r...
Nergadze SG, Belloni E, Piras FM, Khoriauli L, Mazzagatti A, Vella F, Bensi M, Vitelli V, Giulotto E, Raimondi E.Centromeres are the sites of kinetochore assembly and spindle fiber attachment and consist of protein-DNA complexes in which the DNA component is typically characterized by the presence of extended arrays of tandem repeats called satellite DNA. Here, we describe the isolation and characterization of a 137-bp-long new satellite DNA sequence from the horse genome (EC137), which is also present, even if less abundant, in the domestic donkey, the Grevy's zebra and the Burchelli's zebra. We investigated the chromosomal distribution of the EC137 sequence in these 4 species. Moreover, we analyzed its...
Giara and Sarcidano are 2 of the 15 extant native Italian horse breeds with limited dispersal capability that originated from a larger number of individuals. The 2 breeds live in two distinct isolated locations on the island of Sardinia. To determine the genetic structure and evolutionary history of these 2 Sardinian breeds, the first hypervariable segment of the mitochondrial DNA (mtDNA) was sequenced and analyzed in 40 Giara and Sarcidano horses and compared with publicly available mtDNA data from 43 Old World breeds. Four different analyses, including genetic distance, analysis of molecular...
Marenzoni ML, Sforna M, Stefanetti V, Casagrande Proietti P, Brignone L, Del Sero A, Falcioni F, Orvieto S, Tamantini C, Tiburzi A, Valentini S....In recent years, there has been increasing evidence of the potential pathogenic significance of equine gammaherpesviruses in the horse. In humans, cattle and mice, gammaherpesviruses have already been associated with uterine infection. The aim of the present study was to investigate the presence of gammaherpesviruses in uterine flushings of mares with reproductive problems and to evaluate if there was a possible statistical association with clinical and laboratory findings in these cases. A total of 80 uterine flushings were collected from 61 mares with different reproductive problems and thes...
Herrera C, Morikawa MI, Castex CB, Pinto MR, Ortega N, Fanti T, Garaguso R, Franco MJ, Castañares M, Castañeira C, Losinno L, Miragaya MH, Mutto AA.Normal mammalian early embryonic development involves apoptosis of blastomeres as a remodeling process during differentiation, starting at the blastocyst stage. Genomic DNA has been recently detected in the blastocele fluid of human embryos and has been amplified by real-time polymerase chain reaction (PCR) to diagnose the sex of in vitro-produced human embryos. This new approach varies from conventional preimplantation genetic diagnosis in that no cells are extracted from the embryo and only the blastocele fluid is aspirated and used as a DNA sample for diagnosis. In the present work, we inv...
Balao da Silva CM, Ortega Ferrusola C, Gallardo Bolaños JM, Plaza Dávila M, Martín-Muñoz P, Morrell JM, Rodriguez Martínez H, Peña FJ.Flow cytometry is considered the only reliable method for the separation of X and Y chromosome bearing spermatozoa in equines. The MoFlo SX DP sorter is highly efficient, allowing the production of foals of the desired sex. However, to achieve acceptable pregnancy rates the currently used protocol requires working with fresh semen obtained close to, or at, the sorting facility. An alternative protocol was tested during two consecutive breeding seasons. Fresh stallion semen was cooled for 20 h, during which staining with Hoechst 33342 took place. On the following day, this sample was flow sort...
Sykora S, Brandt S.Equine hoof canker is a chronic pododermatitis of still unknown aetiology. Recent findings reported for 3 canker-bearing individuals are suggestive for Treponema spp. having a role in disease pathogenesis. Objective: Based on this hypothesised association, we assessed a larger number of DNA samples from hooves with canker and normal hooves for the presence of treponemal DNA. Methods: Retrospective survey of archived material. Methods: The study involved 71 archival, PCR-compatible DNA extractions purified from 59 canker samples obtained from 26 equine cases and from 12 hoof biopsies taken from...
Yeste M, Estrada E, Rocha LG, Marín H, Rodríguez-Gil JE, Miró J.Although cryopreservation of stallion spermatozoa allows long-term preservation of spermatozoa from particular stallions and facilitates international trade, it is understood to inflict damages on sperm cells that may finally reduce their fertilizing ability. In addition, individual differences are known to exist in the sperm ability to withstand freeze-thawing protocols. To date, these differences have mainly been reported on the basis of sperm motility and membrane integrity. For this reason, the present work sought to determine differences between good (good freezability ejaculates: GFE) an...
Gil L, Olaciregui M, Luño V, Malo C, González N, Martínez F.In recent years, there has been an increased interest in new preservation techniques that facilitate sperm storage and distribution, with freeze-drying (FD) having been proposed as an alternative method for sperm preservation and maintenance of genetic resources in different animal species. FD is a method in which frozen material is dried by sublimation of ice, thereby involving a direct transition from a solid (ice) to a vapour (gas) phase. One of the main advantages of FD is that nitrogen and dry ice are no longer required for the storage and shipment of frozen sperm, which can be stored at ...
Frischknecht M, Neuditschko M, Jagannathan V, Drögemüller C, Tetens J, Thaller G, Leeb T, Rieder S.A cost-effective strategy to increase the density of available markers within a population is to sequence a small proportion of the population and impute whole-genome sequence data for the remaining population. Increased densities of typed markers are advantageous for genome-wide association studies (GWAS) and genomic predictions. Methods: We obtained genotypes for 54 602 SNPs (single nucleotide polymorphisms) in 1077 Franches-Montagnes (FM) horses and Illumina paired-end whole-genome sequencing data for 30 FM horses and 14 Warmblood horses. After variant calling, the sequence-derived SNP geno...
Legrand R, Tiret L, Abitbol M.Seven donkey breeds are recognized by the French studbook. Individuals from the Pyrenean, Provence, Berry Black, Normand, Cotentin and Bourbonnais breeds are characterized by a short coat, while those from the Poitou breed (Baudet du Poitou) are characterized by a long-hair phenotype. We hypothesized that loss-of-function mutations in the FGF5 (fibroblast growth factor 5) gene, which are associated with a long-hair phenotype in several mammalian species, may account for the special coat feature of Poitou donkeys. To the best of our knowledge, mutations in FGF5 have never been described in Equi...
Finno CJ, Aleman M, Higgins RJ, Madigan JE, Bannasch DL.Genome-wide association (GWA) studies are widely used to investigate the genetic etiology of diseases in domestic animals. In the horse, GWA studies using 40-50,000 single nucleotide polymorphisms (SNPs) in sample sizes of 30-40 individuals, consisting of only 6-14 affected horses, have led to the discovery of genetic mutations for simple monogenic traits. Equine neuroaxonal dystrophy is a common inherited neurological disorder characterized by symmetric ataxia. A case-control GWA study was performed using genotypes from 42,819 SNP marker loci distributed across the genome in 99 clinically phe...
Smet A, Cools P, Krizova L, Maixnerova M, Sedo O, Haesebrouck F, Kempf M, Nemec A, Vaneechoutte M.We previously reported the presence of an OXA-23 carbapenemase in an undescribed species of the genus Acinetobacter isolated from horse dung at the Faculty of Veterinary Medicine, Ghent University, Belgium. Here we include six strains to corroborate the delineation of this taxon by phenotypic characterization, DNA-DNA hybridization, 16S rRNA gene and rpoB sequence analysis, % G+C determination, MALDI-TOF MS and fatty acid analysis. The nearly complete 16S rRNA gene sequence of strain UG 60467(T) showed the highest similarities with those of the type strains of Acinetobacter bouvetii (98.4 %)...
Rakhshandehroo E, Sharifiyazdi H, Shayegh H, Ahmadi A.Habronema muscae is a spirurid nematode that undergoes developmental stages in the stomach of equids, causing chronic catarrhal gastritis. Despite preceding investigations have developed polymerase chain reaction (PCR)-based assays for molecular diagnosis, we aimed to assess the applicability of cytochrome c oxidase subunit 1 (cox1) sequences to identify the H. muscae infection and to assess the level of intraspecific variations in this parasite obtained from affected horses in Southern Iran. According to the morphological characterizations, two different isolates of H. muscae were identified....
Ząbek T, Semik E, Wnuk M, Fornal A, Gurgul A, Bugno-Poniewierska M.DNA methylation patterns and their relation with genetic polymorphisms were determined in the equine OAS1 locus. Genetic variants of OAS1 were previously found to be associated with susceptibility to West Nile virus infections in horses. The subject of the study were white blood cells of 13 juvenile and 13 old horses from AA and HC breed and a set of solid tissues from a single adult horse. The aim was to determine the degree of variation of CpG methylation profiles with concern for tissue type, horse breed and age. Results of direct BSPCR and cloned BSPCR sequencing revealed that all of deter...
Park KD, Kim H, Hwang JY, Lee CK, Do KT, Kim HS, Yang YM, Kwon YJ, Kim J, Kim HJ, Song KD, Oh JD, Kim H, Cho BW, Cho S, Lee HK.Copy number variations (CNVs), important genetic factors for study of human diseases, may have as large of an effect on phenotype as do single nucleotide polymorphisms. Indeed, it is widely accepted that CNVs are associated with differential disease susceptibility. However, the relationships between CNVs and gene expression have not been characterized in the horse. In this study, we investigated the effects of copy number deletion in the blood and muscle transcriptomes of Thoroughbred racing horses. We identified a total of 1,246 CNVs of deletion polymorphisms using DNA re-sequencing data from...
Trewby H, Ayele G, Borzacchiello G, Brandt S, Campo MS, Del Fava C, Marais J, Leonardi L, Vanselow B, Biek R, Nasir L.Papillomaviruses are a family of slowly evolving DNA viruses and their evolution is commonly linked to that of their host species. However, whilst bovine papillomavirus-1 (BPV-1) primarily causes warts in its natural host, the cow, it can also cause locally aggressive and invasive skin tumours in equids, known as sarcoids, and thus provides a rare contemporary example of cross-species transmission of a papillomavirus. Here, we describe the first phylogenetic analysis of BPV-1 in equine sarcoids to our knowledge, allowing us to explore the evolutionary history of BPV-1 and investigate its cross...
Lee JH, Lee T, Lee HK, Cho BW, Shin DH, Do KT, Sung S, Kwak W, Kim HJ, Kim H, Cho S, Park KD.Genetics is important for breeding and selection of horses but there is a lack of well-established horse-related browsers or databases. In order to better understand horses, more variants and other integrated information are needed. Thus, we construct a horse genomic variants database including expression and other information. Horse Single Nucleotide Polymorphism and Expression Database (HSDB) (http://snugenome2.snu.ac.kr/HSDB) provides the number of unexplored genomic variants still remaining to be identified in the horse genome including rare variants by using population genome sequences of...
Wang W, Zhang D, Hu D, Chu H, Cao J, Ente M, Jiang G, Li K.Gasterophilosis is a significant threat to equids in the desert steppe of Xinjiang, China, where Gasterophilus pecorum (Fabricius) (Diptera: Gasterophilidae) is the dominant botfly species. A population analysis was conducted on 195 individual G. pecorum larvae from three host species, Przewalski's horse, the domestic horse and the Asiatic wild ass. The distribution of haplotypes of the maternally inherited mitochondrial cytochrome oxidase subunit I (COI) gene was analysed to assess the population differentiation of G. pecorum. High haplotype diversity was observed among G. pecorum popul...
Tydén E, Dahlberg J, Karlberg O, Höglund J.The development of anthelmintic resistance (AR) to macrocyclic lactones in the equine roundworm Parascaris equorum has resulted in benzimidazoles now being the most widely used substance to control Parascaris infections. However, over-reliance on one drug class is a risk factor for the development of AR. Consequently, benzimidazole resistance is widespread in several veterinary parasites, where it is associated with single nucleotide polymorphisms (SNPs) in drug targets encoded by the β-tubulin genes. The importance of these SNPs varies between different parasitic nematodes, but it has been h...
Esteves CL, Sharma R, Dawson L, Taylor SE, Pearson G, Keen JA, McDonald K, Aurich C, Donadeu FX.Expression of several putative markers of pluripotency (OCT4, SOX2, NANOG, LIN28A, REX1, DNMT3B and TERT) was examined in a range of equine tissues, including early embryos, induced pluripotent stem cells (iPSCs), testis, adipose- and bone marrow-derived mesenchymal stromal cells (MSCs), and keratinocytes. Transcript levels of all markers were highest in embryos and iPSCs and, except for SOX2, were very low or undetectable in keratinocytes. Mean expression levels of all markers were lower in testis than in embryos or iPSCs and, except for DNMT3B, were higher in testis than in MSCs. Expression ...
Boeer U, Buettner FF, Klingenberg M, Antonopoulos GC, Meyer H, Haverich A, Wilhelmi M.The limited biocompatibility of decellularized scaffolds is an ongoing challenge in tissue engineering. Here, we demonstrate the residual immunogenicity of an extensively decellularized equine carotid artery (dEAC(intens)) and identify the involved immunogenic components. EAC were submitted to an elaborated intensified decellularization protocol with SDS/sodium desoxycholate for 72 h using increased processing volumes (dEAC(intens)), and compared to dEAC(ord) prepared by an ordinary protocol (40 h, normal volumes). Matrix integrity was checked via correlative volumetric visualization which rev...
Abitbol M, Legrand R, Tiret L.The seven donkey breeds recognised by the French studbook are characterised by few coat colours: black, bay and grey. Normand bay donkeys seldom give birth to red foals, a colour more commonly seen and recognised in American miniature donkeys. Red resembles the equine chestnut colour, previously attributed to a mutation in the melanocortin 1 receptor gene (MC1R). We used a panel of 124 donkeys to identify a recessive missense c.629T>C variant in MC1R that showed a perfect association with the red coat colour. This variant leads to a methionine to threonine substitution at position 210 in th...
Sano Y, Matsuda K, Osaki K, Miyasho T, Tsuda T, Taniyama H.A male Thoroughbred fetus was aborted on day 251 of pregnancy. Gross and histological examinations detected systemic granulomatous lesions in many superficial and visceral lymph nodes and organs including the liver, tonsils, lungs, thymus, spleen, right thyroid gland and gastrointestinal tract, and suppurative placentitis, pyogranulomatous amnionitis and intralesional acid-fast bacilli were also detected. An examination of the DNA base sequence of the β subunit of RNA polymerase demonstrated that Mycobacterium avium strain 104 had infected several organs. To the best of our knowledge, this is...
Sadet-Bourgeteau S, Philippeau C, Dequiedt S, Julliand V.The horse's hindgut bacterial ecosystem has often been studied using faecal samples. However few studies compared both bacterial ecosystems and the validity of using faecal samples may be questionable. Hence, the present study aimed to compare the structure of the equine bacterial community in the hindgut (caecum, right ventral colon) and faeces using a fingerprint technique known as Automated Ribosomal Intergenic Spacer Analysis (ARISA). Two DNA extraction methods were also assessed. Intestinal contents and faeces were sampled 3 h after the morning meal on four adult fistulated horses fed mea...
Ohnuma A, Tozaki T, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Takahashi Y, Nagata SI.To ensure fair competition and sports integrity, gene doping is prohibited in horseracing and equine sports. One gene doping method is by administering exogenous genes, called transgenes, to postnatal animals. Although several transgene detection methods have been developed for horses, many are unsuitable for multiplex detection. In this proof-of-concept study, we developed a highly sensitive and multiplex transgene detection method using multiple πCode with identification patterns printed on the surface. The following steps were employed: (1) multiplex polymerase chain reaction amplification...
Gastal GDA, Aguiar FLN, Alves BG, Alves KA, de Tarso SGS, Ishak GM, Cavinder CA, Feugang JM, Gastal EL.Ovarian tissue cryopreservation allows the preservation of the female fertility potential for an undetermined period. The objectives of this study were to compare the efficiency of cryoprotective agents (CPAs; dimethyl sulfoxide, DMSO; ethylene glycol, EG; and propylene glycol, PROH) using slow-freezing and vitrification methods, and evaluate the viability of cryopreserved equine ovarian tissue after 7 days of culture. Fresh and cryopreserved ovarian fragments were evaluated for preantral follicle morphology, stromal cell density, EGFR, Ki-67, Bax, and Bcl-2 protein expression, and DNA fragmen...
Abdullah HHAM, Elbayoumy MK, Allam AM, Ashry HM, Abdel-Shafy S.Vector-borne bacterial diseases (VBBD) are a diverse group of tropical and subtropical zoonotic diseases. This study investigated the possibility of domestic animals to carry certain vector-borne bacterial microorganisms (VBBMs), as well as the presence of these targeted DNAs in their ectoparasites in different localities of Egypt using molecular analyses. For this study, 234 animal hosts (112 cattle, 38 sheep, 28 goats, 26 buffaloes, 22 donkeys, and 8 horses) in addition to 115 ectoparasites (95 ticks and 20 lice) were investigated for the molecular detection of Bartonella spp., Borrelia spp....
Nowicki ST, Minning-Wenz D, Johnston KH, Lottenberg R.Streptokinases are proteins with plasminogen activator activity produced by certain hemolytic streptococci. We previously identified equine streptococcal isolates which produced streptokinases (ESKs) that bound both human and equine plasminogen but only readily activated equine plasminogen (14). This property was exploited to purify a representative ESK produced by Streptococcus equisimilis strain 87-542-W. Affinity chromatography with human plasminogen resulted in the isolation of a M(r) approximately 49,000 molecule with two isoforms. This ESK was subsequently compared to well characterized ...
Hoyles L, Falsen E, Foster G, Rogerson F, Collins MD.A polyphasic taxonomic study was performed on a previously unidentified gram-positive, facultatively anaerobic, diphtheroid-shaped organism isolated from a vaginal discharge of a horse. Comparative 16S rRNA gene sequencing demonstrated that the strain was a member of the genus Arcanobacterium, but sequence divergence values of >4% with described species of this genus (viz: Arcanobacterium haemolyticum, Arcanobacterium bernardiae, Arcanobacterium phocae, Arcanobacterium pluranimalium and Arcanobacterium pyogenes) demonstrated that the isolate represented a novel species. The unknown bacteriu...
Borkent D, Reardon RJM, McLACHLAN G, Glendinning L, Dixon PM.Although, peripheral caries (PC) affects almost half of UK horses, no comprehensive microbiological study has been performed on this disorder. As a high proportion of oral bacteria cannot be conventionally cultured, molecular microbiological techniques such as Next Generation Sequencing are required to examine the complex oral bacteria community. Objective: To identify the microbiota involved in equine PC, including comparing microbiota at the more commonly and severely affected three caudal cheek teeth with the less commonly affected three rostral cheek teeth. Methods: Equine dental plaque sa...
Grønbaek LM, Angen O, Vigre H, Olsen SN.Streptococcus equi is the cause of strangles in horses. To improve diagnostic sensitivity, development and evaluation of DNA-based methods are necessary. Objective: To evaluate diagnostic methods and observe the pattern of bacterial shedding during natural outbreaks. Methods: Two herds with natural outbreaks of strangles were visited over a period of 15 weeks and 323 samples originating from 35 horses investigated. The diagnostic use of a nested PCR test was evaluated using a collection of 165 isolates of Lancefield group C streptococci (species specificity) and swabs from nasal passages or fr...
Park T, Yoon J, Kim A, Unno T, Yun Y.(1) Background: The large intestine of horses is an anaerobic fermentative chamber filled with fibrolytic bacteria that play essential roles in digesting and absorbing nutrients for energy production. Although Jeju horses are a prominent local breed in Korea, few studies have investigated the gut microbiota of Jeju horses; (2) Methods: This study performed sequencing of V3 and V4 hypervariable regions of the partial 16S rRNA genes obtained from horse fecal samples and compared the gut microbiota between Jeju and Thoroughbred horses. Thirty and 24 fecal samples were obtained from Jeju and Thoro...
Shen Z, Zhang MZ, Stich RW, Mitchell WJ, Zhang S.Anaplasma spp., Ehrlichia spp., Rickettsia spp., and Lyme disease associated Borrelia spp. are the most common tick-borne pathogens reported to infect human beings worldwide and other animals, such as dogs and horses. In the present study, we developed a broad-coverage SYBR Green QPCR panel consisting of four individual assays for the detection and partial differentiation of the aforementioned pathogens. All assays were optimized to the same thermocycling condition and had a detection limit of 10 copies per reaction. The assays remained sensitive when used to test canine and equine blood DNA s...
Zylicz-Stachula A, Zolnierkiewicz O, Jasiecki J, Skowron PM.Genomics and metagenomics are currently leading research areas, with DNA sequences accumulating at an exponential rate. Although enormous advances in DNA sequencing technologies are taking place, progress is frequently limited by factors such as genomic contig assembly and generation of representative libraries. A number of DNA fragmentation methods, such as hydrodynamic sharing, sonication or DNase I fragmentation, have various drawbacks, including DNA damage, poor fragmentation control, irreproducibility and non-overlapping DNA segment representation. Improvements in these limited DNA scissi...
Horín P, Matiasovic J.More than two nucleotide sequences of the second exon of the ELA-DQB region retrieved from a single animal and two different sequences isolated from horses homozygous in the major histocompatibility complex (MHC) region by descent indicated the existence of at least two ELA-DQB loci at the genomic level. New alleles detected by polymerase chain reaction single strand conformation polymorphism (SSCP) and defined by nucleotide sequencing of the second exon of the DQB gene(s) were described. Based on the level of nucleotide sharing, at least two groups of alleles were shown to exist. The newly de...
Frontiers in geneticsJanuary 23, 2023
Volume 14 1099896 doi: 10.3389/fgene.2023.1099896
Capomaccio S, Ablondi M, Colombi D, Sartori C, Giontella A, Cappelli K, Mancin E, Asti V, Mantovani R, Sabbioni A, Silvestrelli M. The Italian peninsula is in the center of the Mediterranean area, and historically it has been a hub for numerous human populations, cultures, and also animal species that enriched the hosted biodiversity. Horses are no exception to this phenomenon, with the peculiarity that the gene pool has been impacted by warfare and subsequent "colonization". In this study, using a comprehensive dataset for almost the entire Italian equine population, in addition to the most influential cosmopolitan breeds, we describe the current status of the modern Italian gene pool. The Italian dataset comprised 1,3...
Hammerschmitt ME, Henker LC, Lichtler J, da Costa FVA, Soares RM, Llano HAB, Pavarini SP.Sarcocystis neurona is the main agent associated with equine protozoal myeloencephalitis (EPM). Apart from horses, S. neurona has been occasionally described causing neurologic disease in several other terrestrial animals as well as mortality in marine mammals. Herein, we describe the clinical, pathological, and molecular findings of a fatal case of S. neurona-associated meningoencephalitis in a domestic cat. The causing agent was analyzed by multilocus genotyping, confirming the presence of S. neurona DNA in the tissue samples of the affected animal. Significant molecular differences were fou...
Corapi WV, Snowden KF, Rodrigues A, Porter BF, Buote MA, Birch SM, Jackson ND, Eden KB, Whitley DB, Mansell J, Edwards JF, Hardy J, Chaffin MK.The schistosome Heterobilharzia americana infects dogs, raccoons, and other mammals in the southeastern United States. Migration of eggs into the liver results in parasitic granulomas with varying degrees of fibrosis and inflammation. Recently, hepatic parasitic granulomas in horses were shown to be caused by H. americana infection. In the present study, samples of liver from 11 of 12 horses with hepatic granulomas identified at necropsy (n = 11) or surgical biopsy (n = 1) were used for DNA extraction, polymerase chain reaction amplification and sequencing using primers specific for a portion ...
Rui BR, Angrimani D, Bicudo LC, Losano J, Nichi M, Pereira R.Sperm DNA fragmentation is a condition that interferes directly in the reproductive efficiency. Currently, there are several methods for assessing the sperm DNA integrity, such as Alkaline Comet, TUNEL and Sperm Chromatin Structure Assay. However, many of these techniques are laborious and require high-precision equipment. Thus, the development of new techniques can optimize the evaluation of sperm DNA damage. Therefore, the aim of this study was to standardize the toluidine blue (TB) stain technique for the analysis of DNA fragmentation of dog, cat, bull, stallion and ram spermatozoa. For thi...
Klonisch T, Steger K, Kehlen A, Allen WR, Froehlich C, Kauffold J, Bergmann M, Hombach-Klonisch S.We employed molecular and immunological techniques to investigate the expression of INSL3, a member of the insulin-like superfamily, in prepubertal testis, postpubertal testes exhibiting normal and disturbed spermatogenesis, and cryptorchid testes of male horses. In addition, the partial cDNA coding sequences of the equine homologue of the human relaxin/INSL3-receptor Lgr8 were determined. Nonradioactive in-situ hybridization with a cRNA probe for equine Insl3 and immunohistochemistry with a specific rabbit INSL3 antiserum localized Insl3 transcripts and immunoreactive INSL3 ligand to Leydig c...
Bond S, McMullen C, Timsit E, Léguillette R.The lower respiratory tract microbiota of the horse is different in states of health and disease, but the bacterial and fungal composition of the healthy respiratory tract of the horse has not been studied in detail. Objective: The respiratory tract environment contains distinct niche microbiotas, which decrease in species richness at more distal sampling locations. Objective: Characterize the bacterial and fungal microbiotas along the upper and lower respiratory tract of the horse. Methods: Healthy Argentinian Thoroughbred horses (n = 11) from the same client-owned herd. Methods: Prospectiv...
Shojaeian K, Nouri H, Kohram H.Overproduction of reactive oxygen species during sperm freeze-thawing process leads to membrane lipid peroxidation, DNA damage, motility loss, and subsequent death. This oxidative stress can be alleviated by the addition of some antioxidants to semen extenders prior to freezing. This study was performed to evaluate the in vitro effectiveness of MnTBAP (a cell permeable antioxidant) on stallion sperm freezability and in vivo fertility rate. Twenty-one ejaculates were, collected with missouri model artificial vagina (n = 3 stallions, seven ejaculate each), and diluted (1:2 v/v) with phosphoc...
Tozaki T, Mashima S, Hirota K, Miura N, Choi-Miura NH, Tomita M.We performed efficient cloning and genotyping methods for isolation of a large number of polymorphic microsatellites. The methods contain the time-efficient cloning method of constructing microsatellite-enriched libraries and the economic genotyping method of fluorescent labeling of PCR products. Eighty novel equine microsatellites cloned were efficiently isolated from the enrichment library and analyzed for genotype polymorphism. Of these, 72 microsatellites were analyzed with a good resolution. The average heterozygosity of all loci was 0.52, and the number of alleles ranged from one to 9 wi...
Paruch L, Paruch AM.Zoogenic faecal contamination of the environment is one of the indices included in the evaluation of ecological threats, health hazards and adverse impacts on various ecosystems. The risks and environmental concerns are associated with the fact that faeces of wild and domesticated animals constitute the largest source of environmental loading of enteropathogens associated with transmission of zoonotic diseases (enteric zoonoses). Although sick animals are more likely to transmit pathogens, healthy ones can also be the carriers and defecate them into the environment. This is of particular impor...
Cwiklinski K, Kooyman FN, Van Doorn DC, Matthews JB, Hodgkinson JE.Cyathostomins comprise a group of 50 species of parasitic nematodes that infect equids. Ribosomal DNA sequences, in particular the intergenic spacer (IGS) region, have been utilized via several methodologies to identify pre-parasitic stages of the commonest species that affect horses. These methods rely on the availability of accurate sequence information for each species, as well as detailed knowledge of the levels of intra- and inter-specific variation. Here, the IGS DNA region was amplified and sequenced from 10 cyathostomin species for which sequence was not previously available. Also, add...
Feral horses in Theodore Roosevelt National Park (TRNP) represent an iconic era of the North Dakota Badlands. Their uncertain history raises management questions regarding origins, genetic diversity, and long-term genetic viability. Hair samples with follicles were collected from 196 horses in the Park and used to sequence the control region of mitochondrial DNA (mtDNA) and to profile 12 autosomal short tandem repeat (STR) markers. Three mtDNA haplotypes found in the TRNP horses belonged to haplogroups L and B. The control region variation was low with haplotype diversity of 0.5271, nucleotide...
Tozaki T, Hirota K, Hasegawa T, Tomita M, Kurosawa M.Linkage disequilibrium (LD) mapping is often used in searches for genes governing economically significant traits and diseases. The D' coefficient is a commonly used measure of the extent of LD between all possible pairs of alleles at two markers. This study aimed to test the utility of the D' coefficient for LD mapping of a trait in a thoroughbred population. Microsatellite genotype data and grey coat colour as a trait model in a thoroughbred population were used to assess the extent of LD. We demonstrated that LD mapping was a reasonable approach for initial genome-wide scans in a thoroughbr...
Lucchesi PM, Parma AE.Horses infected with Leptospira interrogans present several clinical disorders, one of them being recurrent uveitis. An antigenic relationship between this bacterium and equine cornea has been described in previous studies. With the aim to make progress on defining the molecular basis and pathogenesis of equine recurrent uveitis, here we describe the cloning of one DNA fragment from a Leptospira interrogans serovar pomona genomic lambda gt11 library. Although there are references of transcription of leptospiral genes in E. coli from their own leptospiral promoters, in this recombinant construc...
Tozaki T, Ohnuma A, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Kusano K, Nagata SI.Processed pseudogenes, also known as retrocopy genes, are copies of messenger RNAs that have been reverse transcribed into DNA and inserted into the genome. In this study, we identified 62 processed pseudogene candidates as intron-less genes from whole-genome sequencing (WGS) data of Thoroughbred horses using delly structural variation software. The 62 processed pseudogene candidates were confirmed by PCR amplification of intron-less products. A total of 11 processed pseudogenes were confirmed in the genome of all 23 analysed horses, whereas three processed pseudogenes with structures of ATP11...
Vieira MIB, Costa MM, de Oliveira MT, Gonçalves LR, André MR, Machado RZ.Equine piroplasmosis is a disease caused by the hemoparasites Babesia caballi and Theileria equi and is considered to be the most important parasitic infection affecting Equidae. The objective of the present study was to carry out an epidemiological molecular and serological survey for the presence of these two protozoal organisms in equids from the northwestern region of the State of Rio Grande do Sul (RS), south Brazil. For this purpose, blood samples were collected from 90 equids in the city of Passo Fundo, RS, Brazil. Those were animals used for sport activities, outdoor recreational ridin...
Sild E, Värv S, Kaart T, Kantanen J, Popov R, Viinalass H.The maternal and paternal genetic variation of horse breeds from the Baltic Sea region, including three local Estonian breeds, was assessed and compared with that of Altai and Yakutian horses. In the mtDNA D-loop region, 72 haplotypes assigned to 20 haplogroups in the nine breeds were detected. In Estonian local breeds, 38 mtDNA haplotypes were found, and five of them were shared by the three breeds. More than 60% of all identified haplotypes were rare. Compared with the Estonian Native and Estonian Heavy Draught breeds, a higher haplotypic diversity was found in the Tori breed (h = 0.969). ...
Cheung HW, Wong KS, Lin VYC, Farrington AF, Bond AJ, Wan TSM, Ho ENM.The concern about gene doping has remained high in horseracing and other equestrian competitions. Our laboratory has previously developed a duplex quantitative polymerase chain reaction (qPCR) assay capable of detecting in equine blood the human erythropoietin (hEPO) transgene and equine tubulin α 4a (TUBA4A) gene as an internal control the latter providing quality control over DNA extraction and qPCR. This study aimed to optimize the method for routine testing of regulatory samples. The use of an automated DNA extraction system has increased the sample throughput, consistency of DNA extracti...
Brooks KE, Daughtry BL, Metcalf E, Masterson K, Battaglia D, Gao L, Park B, Chavez SL.The timing of early mitotic events during preimplantation embryo development is important for subsequent embryogenesis in many mammalian species, including mouse and human, but, to date, no study has closely examined mitotic timing in equine embryos from oocytes obtained by ovum pick-up. Here, cumulus-oocyte complexes were collected by transvaginal follicular aspiration, matured invitro and fertilised via intracytoplasmic sperm injection. Each fertilised oocyte was cultured up to the blastocyst stage and monitored by time-lapse imaging for the measurement of cell cycle intervals and identifica...
Borchers K, Wolfinger U, Lawrenz B, Schellenbach A, Ludwig H.Neuronal and lymphoid tissues of 15 randomly selected horses were analysed post mortem by liquid nested-PCR to study the tropism of equine herpesvirus 4 (EHV-4). In four animals the trigeminal ganglia and in one case the lung were positive. Using a direct in situ PCR the EHV-4 genome was localized in the nuclei of neurons and in the bronchiolar as well as alveolar epithelium of the lung. In none of these tissues could infectious virus or viral antigens be detected. Applying the more sensitive liquid RT-PCR, however, an acute infection was demonstrated in one of the trigeminal ganglia by amplif...
