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.
Doan R, Cohen N, Harrington J, Veazey K, Juras R, Cothran G, McCue ME, Skow L, Dindot SV.Copy number variants (CNVs) represent a substantial source of genetic variation in mammals. However, the occurrence of CNVs in horses and their subsequent impact on phenotypic variation is unknown. We performed a study to identify CNVs in 16 horses representing 15 distinct breeds (Equus caballus) and an individual gray donkey (Equus asinus) using a whole-exome tiling array and the array comparative genomic hybridization methodology. We identified 2368 CNVs ranging in size from 197 bp to 3.5 Mb. Merging identical CNVs from each animal yielded 775 CNV regions (CNVRs), involving 1707 protein- and...
Malik P, Bálint A, Dán A, Pálfi V.Equine herpesvirus-1 (EHV-1) can be classified into distinct groups by single nucleotide polymorphisms (SNPs) in their genomes. Only a few of these can be associated with a special attribute of the virus. Differences in the ORF30 region can determine the neuropathogenic potential, while by substitutions in the ORF68 region several strain groups can be made. In previous studies no connection was found between the neuropathogenic potential and the SNPs in ORF68, but the occurrence of members of distinct groups in different outbreaks can facilitate epidemiological investigations because the geogr...
Doan R, Cohen ND, Sawyer J, Ghaffari N, Johnson CD, Dindot SV.The catalog of genetic variants in the horse genome originates from a few select animals, the majority originating from the Thoroughbred mare used for the equine genome sequencing project. The purpose of this study was to identify genetic variants, including single nucleotide polymorphisms (SNPs), insertion/deletion polymorphisms (INDELs), and copy number variants (CNVs) in the genome of an individual Quarter Horse mare sequenced by next-generation sequencing. Results: Using massively parallel paired-end sequencing, we generated 59.6 Gb of DNA sequence from a Quarter Horse mare resulting in an...
Kulikov E, Kropinski AM, Golomidova A, Lingohr E, Govorun V, Serebryakova M, Prokhorov N, Letarova M, Manykin A, Strotskaya A, Letarov A.Lytic coliphage vB_EcoP_G7C and several other highly related isolates were obtained repeatedly from the samples of horse feces held in the same stable thus representing a component of the normal indigenous intestinal communities in this population of animals. The genome of G7C consists of 71,759 bp with terminal repeats of about 1160 bp, yielding approximately 73 kbp packed DNA size. Seventy-eight potential open reading frames, most of them unique to N4-like viruses, were identified and annotated. The overall layout of functional gene groups was close to that of the original N4 phage, with som...
Sykora S, Samek L, Schönthaler K, Palm F, Borzacchiello G, Aurich C, Brandt S.Squamous cell carcinomas (SCC) are malignant tumours arising from keratinocytes. In horses, there is increasing evidence for Equus caballus papillomavirus type 2 (EcPV-2) being causally involved in SCC development. However, only little is known regarding intralesional transcription of the virus, and sparse information on the incidence of EcPV-2 infection in healthy equids is available so far. Using RT-PCR, total mRNA from 8 EcPV-2 DNA-positive and 1 EcPV-2 negative SCC/SCC precursor lesions was screened for the presence of EcPV-2 E6 and E1 transcripts. Using PCR, we tested 193 sample specimens...
Bogaert L, Willemsen A, Vanderstraeten E, Bracho MA, De Baere C, Bravo IG, Martens A.Squamous cell carcinoma (SCC) represents the most common genital malignant tumor in horses. Similar to humans, papillomaviruses (PVs) have been proposed as etiological agents and recently Equine papillomavirus type 2 (EcPV2) has been identified in a subset of genital SCCs. The goals of this study were (1) to determine the prevalence of EcPV2 DNA in tissue samples from equine genital SCCs, penile intraepithelial neoplasia (PIN) and penile papillomas, using EcPV2-specific PCR, (2) to examine the prevalence of latent EcPV2 infection in healthy genital mucosa and (3) to determine genetic variabili...
Achilli A, Olivieri A, Soares P, Lancioni H, Hooshiar Kashani B, Perego UA, Nergadze SG, Carossa V, Santagostino M, Capomaccio S, Felicetti M....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 ...
Phillips JC, Lembcke LM, Noltenius CE, Newman SJ, Blackford JT, Grosenbaugh DA, Leard AT.To determine the tissue-restricted expression pattern of tyrosinase mRNA in canine and equine melanocytic tumors and relative tyrosinase and major histocompatibility complex (MHC) I mRNA expression in variants of melanocytic tumors. Methods: 39 canine and 8 equine tumor samples and 10 canine and 6 equine normal tissue samples. Methods: RNA was isolated from formalin-fixed, paraffin-embedded tissues. Real-time PCR assays were designed to amplify canine and equine tyrosinase, S18 ribosomal RNA, and major histocompatibility complex I transcripts. Relative expression was determined by use of S18 a...
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 ...
Nam GH, Ahn K, Bae JH, Cho BW, Park KD, Lee HK, Yang YM, Kim TH, Seong HH, Han K, Kim HS.In the mitochondrial matrix, the OXCT1 gene catalyzes the reversible transfer of coenzyme A from succinyl-CoA to acetoacetate in a reaction related to energy production from ketone bodies. Here, horse OXCT1 gene containing coenzyme A transferase domain was identified in the transcriptome analysis of cDNAs derived from skeletal muscles. Horse OXCT1 gene consisted of 1761 [corrected] nucleotide sequences with an open reading frame of 1560 nucleotides encoding a protein of 520 putative amino acid residues.The number of non-synonymous substitutions was lower than the number of synonymous substitut...
Steinman A, Zimmerman T, Klement E, Lensky IM, Berlin D, Gottlieb Y, Baneth G.The prevalence of Theileria equi infection as well as the environmental and demographic risk factors for infection was studied in 590 healthy horses from 46 farms in Israel. The prevalence of T. equi DNA was assessed using a polymerase chain reaction for a segment of the Theileria 18S rRNA gene. The overall prevalence was 26.4% (156/590). There was a significant geographical variation in the prevalence of T. equi infection, ranging from 9.3% (25/270) in the central lowlands to 81.7% (49/60) in the Golan Heights. The prevalence of T. equi infection was found to be significantly associated with ...
McCue ME, Bannasch DL, Petersen JL, Gurr J, Bailey E, Binns MM, Distl O, Guérin G, Hasegawa T, Hill EW, Leeb T, Lindgren G, Penedo MC, Røed KH....An equine SNP genotyping array was developed and evaluated on a panel of samples representing 14 domestic horse breeds and 18 evolutionarily related species. More than 54,000 polymorphic SNPs provided an average inter-SNP spacing of ∼43 kb. The mean minor allele frequency across domestic horse breeds was 0.23, and the number of polymorphic SNPs within breeds ranged from 43,287 to 52,085. Genome-wide linkage disequilibrium (LD) in most breeds declined rapidly over the first 50-100 kb and reached background levels within 1-2 Mb. The extent of LD and the level of inbreeding were highest in the ...
Hébert L, Moumen B, Pons N, Duquesne F, Breuil MF, Goux D, Batto JM, Laugier C, Renault P, Petry S.The Taylorella genus comprises two species: Taylorella equigenitalis, which causes contagious equine metritis, and Taylorella asinigenitalis, a closely-related species mainly found in donkeys. We herein report on the first genome sequence of T. asinigenitalis, analyzing and comparing it with the recently-sequenced T. equigenitalis genome. The T. asinigenitalis genome contains a single circular chromosome of 1,638,559 bp with a 38.3% GC content and 1,534 coding sequences (CDS). While 212 CDSs were T. asinigenitalis-specific, 1,322 had orthologs in T. equigenitalis. Two hundred and thirty-four T...
Macieira S, Lussier J, Bédard C.Glanzmann thrombasthenia (GT) is characterized by a defect of platelet aggregation. This autosomal recessive genetic disorder is caused by an abnormality of the platelet glycoprotein receptors alpha IIb or beta III. Recently, we identified a horse with clinical and pathological features of GT. The aim of this study was to describe this case of GT at the molecular level. A point mutation from G to C in exon 2 of ITGA2B causing a substitution of the expected amino acid arginine 72 (Arg(72)) by a proline (Pro(72)) was encountered. This amino acid change may result in abnormal structural conformat...
Aleman M, Pickles KJ, Simonek G, Madigan JE.Trigeminal neuralgia or neuropathic pain has been regarded as a putative cause of idiopathic headshaking in horses. Equine herpesvirus-1 (EHV-1) infection and resultant postherpetic pain have been suggested as a possible cause of such neuropathic pain. Objective: To determine the presence of EHV-1 in the trigeminal ganglia of horses with idiopathic headshaking. Methods: Nineteen horses: control (n = 11, 9 geldings, 2 mares, median age 11 years) and headshaking (n = 8, all geldings, median age 11.5 years) horses were sourced from the equine research herd and caseload at the Veterinary Medical T...
Yue XP, Qin F, Campana MG, Liu DH, Mao CC, Wang XB, Lan XY, Chen H, Lei CZ.Previous mitochondrial DNA (mtDNA) D-loop and microsatellite studies have shown that Chinese horses have multiple maternal origins and high genetic diversity. To better characterize maternal genetic origins and diversity of Chinese domestic horses, we conducted a comprehensive analysis of 407 complete 1140 bp sequences of the horse mitochondrially encoded cytochrome b (CYTB) gene, including 323 horses from 13 Chinese indigenous breeds and 84 reference sequences from GenBank. A total of 114 haplotypes were identified, of which 73 appeared among the 13 Chinese horse breeds. The high mitochondria...
Wobeser BK, Hill JE, Jackson ML, Kidney BA, Mayer MN, Townsend HG, Allen AL.Equine sarcoids are the most common tumor of horses. Bovine papillomavirus (BPV) has been suggested as the cause of sarcoids. Studies have shown that BPV is present in swabs or biopsies from nonsarcoid-bearing equine skin. Skin biopsies from a variety of different conditions and normal skin from horses with no reported history of sarcoids were examined by polymerase chain reaction (PCR) for the presence of BPV, which was found in all different types of skin conditions as well as normal skin. Forty-one out of 86 skin biopsies from horses without sarcoids were found to contain BPV DNA. Laser mic...
Qablan MA, Sloboda M, Jirků M, Oborník M, Dwairi S, Amr ZS, Hořín P, Lukeš J, Modrý D.DNA of two species of piroplasmids was detected in dromedaries during a survey of blood protozoans in Jordan between 2007 and 2009. Ten clinically healthy camels (10%) originating from three Jordanian districts were found, using a PCR assay, to harbor Theileria or Babesia species in their blood and no mix infection was determined. Analysis of the partial 18S rRNA gene sequences of these parasites allowed their unambiguous identification as equine piroplasmids Babesia caballi (n=6) and Theileria equi (n=4). In case of latter species, a novel genotype was found in horses. This first molecular-ba...
Feofilov AV, Bardukov NV, Glazko VI.Using ISSR-PCR marker data, comparative analysis of the gene pools of Altaic and trotting horse breeds was carried out. Horse groups of different origin demonstrated differences in amplification spectra of DNA fragments flanked by inverted repeats of four microsatellites. Combinations of certain DNA fragments present in these profiles reproducibly distinguished genomes of the Altaic breed from the trotting breeds. Genetic differentiation between some trotting breeds, based on Nei genetic distance values, was found to be comparable to that between the groups of horses of Altaic breed from two d...
Leon A, Richard E, Fortier C, Laugier C, Fortier G, Pronost S.Abortion, stillbirth and neonatal death are major causes of equine mortality and cause severe economic loss to the equine industry. The present study was based on a complete necropsy protocol associated with classical microbiological examinations and molecular biology on 407 cases of abortion, stillbirths and neonate death. Based on this retrospective survey, "less common" abortive infectious agents were characterised by molecular tools in nine independent cases of abortion or neonate mortality. Among others, Chlamydophila abortus (1 case), Coxiella burnetii (6 cases) and Neospora caninum (3 c...
Edmond AJ, Brinsko SP, Love CC, Blanchard TL, Teague SR, Varner DD.Centrifugal fractionation of semen is commonly done to improve quality of human semen in assisted-reproduction laboratories, allowing sperm separation based on their isopycnic points. Sperm with morphologic abnormalities are often more buoyant, promoting their retention above defined density media, with structurally normal sperm passing through the media following centrifugation. Three experiments were conducted to evaluate the effects of density-medium type, centrifuge-tube size, sperm number, and density-medium volume (column height) on stallion sperm quality and recovery rate in sperm pelle...
Révay T, Villagómez DA, Brewer D, Chenier T, King WA.Genetic sex in mammals is determined by the sex chromosomal composition of the zygote. The X and Y chromosomes are responsible for numerous factors that must work in close concert for the proper development of a healthy sexual phenotype. The role of androgens in case of XY chromosomal constitution is crucial for normal male sex differentiation. The intracellular androgenic action is mediated by the androgen receptor (AR), and its impaired function leads to a myriad of syndromes with severe clinical consequences, most notably androgen insensitivity syndrome and prostate cancer. In this paper, w...
Lippold S, Matzke NJ, Reissmann M, Hofreiter M.DNA target enrichment by micro-array capture combined with high throughput sequencing technologies provides the possibility to obtain large amounts of sequence data (e.g. whole mitochondrial DNA genomes) from multiple individuals at relatively low costs. Previously, whole mitochondrial genome data for domestic horses (Equus caballus) were limited to only a few specimens and only short parts of the mtDNA genome (especially the hypervariable region) were investigated for larger sample sets. Results: In this study we investigated whole mitochondrial genomes of 59 domestic horses from 44 breeds an...
Sun X, Yao H, Zhang C, Lu C.Equine herpesvirus type 1 (EHV-1) is a major cause of respiratory and reproductive diseases in horses worldwide. The genome of EHV-1 strain 438/77 (isolated from an aborted equine fetus) was cloned as a bacterial artificial chromosome (BAC) in E. coli without any gene deletions. The mini-F plasmid sequence was inserted in the middle of ORF19 and 20 via homologous recombination following co-transfection of viral DNA and plasmid pE19_20/HA into RK13 cells. Circular viral DNA was extracted from RK13 cells infected with purified recombinant virus expressing green fluorescent protein (GFP) and elec...
Binns MM, Boehler DA, Bailey E, Lear TL, Cardwell JM, Lambert DH.Changes in the inbreeding coefficient, F, in the Thoroughbred horse over the past 45 years have been investigated by genotyping 467 Thoroughbred horses (born between 1961 and 2006) using the Illumina Equine SNP50 bead chip, which comprises 54,602 SNPs uniformly distributed across the equine genome. The Spearman rank correlation coefficient, r, between the year of birth and F was estimated. The results indicate that inbreeding in Thoroughbreds has increased over the past 40 years, with r = 0.24, P < 0.001 demonstrating that there is a highly significant, though relatively weak correlation be...
Wnuk M, Villagómez DA, Bugno-Poniewierska M, Tumidajewicz P, Carter TF, Slota E.Nucleolar organizing regions (NORs) containing rDNA gene clusters have been assigned to the equine autosomes ECA1, ECA28, and ECA31. Active NORs (Ag-NORs) are associated with argyrophilic proteins, which allow them to be readily identified using silver staining techniques. Fluorescence in situ hybridization (FISH) for rDNA can also be used to visualize all NOR clusters in the nucleus, regardless of whether they are active or inactive. The present study analyzed the distribution and behavior of equine Ag-NOR and NOR clusters in horse spermatozoa and during male meiosis by FISH and silver staini...
Müller JM, Wissemann J, Meli ML, Dasen G, Lutz H, Heinzerling L, Feige K.Whole blood pharmacokinetics of intratumourally injected naked plasmid DNA coding for equine Interleukin 12 (IL-12) was assessed as a means of in vivo gene transfer in the treatment of melanoma in grey horses. The expression of induced interferon gamma (IFN-g) was evaluated in order to determine the pharmacodynamic properties of in vivo gene transduction. Seven grey horses bearing melanoma were injected intratumourally with 250 µg naked plasmid DNA coding for IL-12. Peripheral blood and biopsies from the injection site were taken at 13 time points until day 14 post injection (p.i.). Samples w...
Shepherd ML, Swecker WS, Jensen RV, Ponder MA.The diversity of the equine fecal bacterial community was evaluated using pyrosequencing of 16S rRNA gene amplicons. Fecal samples were obtained from horses fed cool-season grass hay. Fecal bacteria were characterized by amplifying the V4 region of bacterial 16S rRNA gene. Of 5898 mean unique sequences, a mean of 1510 operational taxonomic units were identified in the four fecal samples. Equine fecal bacterial richness was higher than that reported in humans, but lower than that reported in either cattle feces or soil. Bacterial classified sequences were assigned to 16 phyla, of which 10 were ...
Sundararaman B, Vershinina AO, Hershauer S, Kapp JD, Dunn S, Shapiro B, Green RE.Hybridization capture approaches allow targeted high-throughput sequencing analysis at reduced costs compared to shotgun sequencing. Hybridization capture is particularly useful in analyses of genomic data from ancient, environmental, and forensic samples, where target content is low, DNA is fragmented and multiplex PCR or other targeted approaches often fail. Here, we describe a DNA bait synthesis approach for hybridization capture that we call Circular Nucleic acid Enrichment Reagent, or CNER (pronounced 'snare'). The CNER method uses rolling-circle amplification followed by restriction dige...
Goh G, Raudsepp T, Durkin K, Wagner ML, Schäffer AA, Agarwala R, Tozaki T, Mickelson JR, Chowdhary BP.High-resolution physically ordered gene maps for equine homologs of human chromosome 5 (HSA5), viz., horse chromosomes 14 and 21 (ECA14 and ECA21), were generated by adding 179 new loci (131 gene-specific and 48 microsatellites) to the existing maps of the two chromosomes. The loci were mapped primarily by genotyping on a 5000-rad horse x hamster radiation hybrid panel, of which 28 were mapped by fluorescence in situ hybridization. The approximately fivefold increase in the number of mapped markers on the two chromosomes improves the average resolution of the map to 1 marker/0.9 Mb. The improv...
Wang X, Bou G, Zhang X, Tao L, Shen Y, Na R, Liu G, Ren H, Ren X, Song L, Su S, Bai D, Zhao Y, Li B, Dugarjaviin M.Having considered that the current methods are costly and time-consuming, we designed an only 3 pairs primer-based PCR test to accurately identify the species and gender in horses, donkeys, mules and hinnies. Through a thorough sequence comparison between horse and donkey's highly similar genomes, and a vast amount of preliminary confirmation, we found that three fragments, CNGB3 gene on an autosome, displacement loop region on mitochondrial DNA and SRY genes on chromosome Y, within these equine DNA, are enough to enable us achieving our goal. The PCR test described here would be an economical...
Drebert Z, Golke A, Cymerys J, Słońska A, Chmielewska A, Tucholska A, Bańbura MW.Equid herpesvirus type 1 (EHV-1) is a prevalent causative agent of equine diseases worldwide. After primary replication in the respiratory epithelium the virus disseminates systemically through a peripheral blood mononuclear cell (PBMC)-associated viraemia. EHV-1 is the only alphaherpes- virus known so far which is capable of establishing latent infection not only in neurons but also in immune system cells (mainly in lymphocytes and macrophages). Since leukocytes are not the target cells for viral replication but are used to transport EHV-1 to the internal organs, the questionremains how the v...
Drogemuller M, Schnieder T, von Samson-Himmelstjerna G.P-glycoproteins (Pgps) are adenosine triphosphate-binding transporter proteins thought to be associated with multi-drug resistance in mammals and protozoans and have been suggested to be involved in the mechanism of ivermectin (IVM) resistance in Haemonchus contortus. Until now, resistance to IVM has not been reported in cyathostomins in horses in spite of its widespread and frequent use. Reasons for this might be differences in the molecular mechanism of the development of resistance. Based on this hypothesis, the present study was carried out to find homologues of Pgp in cyathostomins. A 416...
Hinshelwood MM, Liu Z, Conley AJ, Simpson ER.Conversion of androgens to estrogens is catalyzed by aromatase P450 (P450arom; the product of the CYP19 gene). Regulation of tissue-specific expression of P450arom in humans is due, in part, to alternative transcriptional start sites that arise as a consequence of the use of granulosa cells and placental tissue from cows, horses, and pigs (ungulates) in order to determine whether these species, like the human, utilize tissue-specific promoters to drive P450arom expression. The majority of transcripts in the placenta have 5'-termini that differ from those in the ovary upstream of a common site ...
Carvalho R, Passos LM, Martins AS.In this study, a multiplex polymerase chain reaction (PCR) procedure was developed for differentiation of strains and field isolates of equine herpesvirus type 1 (EHV-1) and type 4 (EHV-4). Specific oli-gonucleotide primers were combined to amplify the thymidine kinase (TK) gene region of EHV-1 and EHV-4, which would yield fragments of different lengths for each virus in the same amplification reaction. The specificity of the largest PCR amplicon for EHV-4 was confirmed by restriction digestion with HindIII. The multiplex PCR proved to be a fast and sensitive method for typing EHV-1 and EHV-4 ...
RANDALL CC, WALKER BM.Randall, Charles C. (University of Mississippi School of Medicine, Jackson) and Barbara M. Walker. Degradation of deoxyribonucleic acid and alteration of nucleic acid metabolism in suspension cultures of L-M cells infected with equine abortion virus. J. Bacteriol. 86:138-146. 1963.-Metabolic alterations in log-phase suspension cultures infected with equine abortion virus (EAV) were determined in L-M cells simultaneously labeled or prelabeled with H(3)- or C(14)-thymidine. Although infection produced an early stimulation of the uptake of labeled thymidine (TdR) into the acid-soluble fraction of...
Wierdl M, Morton CL, Danks MK, Potter PM.Butyrylcholinesterases (BuChEs; acylcholine acylhydrolase; EC 3.1.1.8) have been demonstrated to convert the anticancer agent CPT-11 (irinotecan, 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin) into its active metabolite SN-38 (7-ethyl-10-hydroxycamptothecin). In addition, significant differences in the extent of drug metabolism have been observed with BuChEs derived from different species. In an attempt to understand these differences, we have isolated the cDNA encoding a horse BuChE. Based upon the NH2-terminal amino acid sequence of a purified horse BuChE, we designed deg...
Carpenter S, Baker JM, Bacon SJ, Hopman T, Maher J, Ellis SA, Antczak DF.Sequence and functional analyses were undertaken on two cDNAs and a genomic clone encoding horse major histocompatibility complex (MHC) class I molecules. All of the clones were isolated from a single horse that is homozygous for all known horse MHC class I and class II antigens. The two cDNAs (clones 8-9 and 1-29) were isolated from a lymphocyte library and encode polymorphic MHC antigens from two loci. The genomic cosmid clone, isolated from a sperm library, contains the 8-9 gene. All three genes were expressed in mouse L-cells and were recognized by alloantisera and, for the cDNAs, by allor...
Demyda-Peyrás S, Membrillo A, Bugno-Poniewierska M, Pawlina K, Anaya G, Moreno-Millán M.Chromosomal abnormalities associated to sex chromosomes are reported as a problem more common than believed to be in horses. Most of them remain undiagnosed due to the complexity of the horse karyotype and the lack of interest of breeders and veterinarians in this type of diagnosis. Approximately 10 years ago, the Spanish Purebred Breeders Association implemented a DNA paternity test to evaluate the pedigree of every newborn foal. All candidates who showed abnormal or uncertain results are routinely submitted to cytogenetical analysis to evaluate the presence of chromosomal abnormalities. We s...
Lindquist CD, Evans JJ, Wictum EJ.Accurate DNA quantification is essential for optimizing DNA testing and minimizing sample consumption. Real-time quantitative polymerase chain reaction (qPCR) assays have been published for human and canine nuclear DNA, and the need for quantifying other forensically important species was evident. Following the strategy employed for the canine qPCR assay, we developed individual assays to accurately quantify feline, bovine, equine, and cervid nuclear DNA. Each TaqMan-based assay incorporates a genus-specific probe targeting the Melanocortin-1 Receptor gene and includes a piece of synthetic DNA...
Ayoub C, Arroyo LG, Renaud D, Weese JS, Gomez DE.The objective of this study was to compare the fecal microbiota of two healthy teaching horse herds with that of client-owned horses from the same geographic areas. The fecal microbiota of client-owned horses from Ontario Canada (n = 15) and Florida, USA (n = 11) was compared with that teaching horses from the University of Guelph, Ontario, Canada (n = 10) and the University of Florida, Florida, USA (n = 15). The fecal microbiota was characterized by sequencing of bacterial DNA using the V4 hypervariable region of the 16S rRNA gene. The diversity (inverse Simpson index) of the fecal mi...
Lepage N, St-Laurent G, Carman S, Archambault D.The genetic variation in equine arteritis virus (EAV) Gs protein encoding gene was investigated. Nucleic and deduced amino acid sequences from eight different EAV isolates (one European, two American and five Canadian isolates) were compared with those of the Bucyrus reference strain. Nucleotide and amino acid identities between these isolates and the Bucyrus reference strain ranged from 92.3 to 96.4%, and 93.2 to 95.5%, respectively. However, phylogenetic tree analysis and estimation of genetic distances based on the Gs protein encoding gene sequences showed that the European prototype Vienna...
Muscatello G, Browning GF.Selective agar media have been used for many years to facilitate the isolation of Rhodococcus equi from environmental and clinical samples. However, characterisation of R. equi still requires the use of immunochemical or polymerase chain reaction (PCR) analysis to differentiate between virulent and avirulent isolates. Here, we describe a novel method to detect and differentiate between R. equi isolates using colony blotting and DNA hybridization. Radiolabelled PCR product derived from the R. equi rrnA gene and specific hybridization conditions enabled differentiation of colonies of R. equi fro...
Nasir L, Gault E, Morgan IM, Chambers G, Ellsmore V, Campo MS.BPV-1 DNA is the predominant viral type detected in equine sarcoids and represents the only reported natural cross species infection of papillomaviruses. In this study, nucleotide variations in the LCR and the E2 regions of equine sarcoid-associated BPV-1 were characterised by sequence analysis. Variants particular to sarcoid BPV-1 were identified in both the LCR and E2 sequence. The functionality of the most common LCR variant was examined in equine and bovine cells. These studies showed that the activity of the variant LCR was higher in equine cells than bovine cells; the activity of the var...
von Samson-Himmelstjerna G, Janssen IJI, Ramünke S, Goday C, Borges FA, Koudela B, Niedźwiedź A, Tomczuk K, Studzińska MB, Kornas S, Krücken J.Equines were over decades considered to be infected by two morphologically virtually indistinguishable ascarid species, Parascaris univalens and Parascaris equorum. Reliable species discrimination is only possible using enzyme isoelectric focussing and karyotyping with P. univalens having one and P. equorum two chromosome pairs. However, presumably the complexity of both methods prevented their routine use in nearly all previous studies about prevalence and drug resistance of Parascaris spp. These have barely been performed on the species level although most studies stated presence of one or t...
Oldenhof H, Schütze S, Wolkers WF, Sieme H.Sperm chromatin structure and condensation determine accessibility for damage, and hence success of fertilization and development. The aim of this study was to reveal characteristic spectral features coinciding with abnormal sperm chromatin packing (i.e., DNA-protein interactions) and decreased fertility, using Fourier transform infrared spectroscopy. Chromatin structure in spermatozoa obtained from different stallions was investigated. Furthermore, spermatozoa were exposed to oxidative stress, or treated with thiol-oxidizing and disulfide-reducing agents, to alter chromatin structure and pack...
Smith KC, Borchers K.The polymerase chain reaction and DNA in-situ hybridization were used to study sections of uterine tissue collected from mares near the time of abortion due to equid herpesvirus-1 (EHV-1) infection. These techniques revealed viral nucleic acids in endothelial cells of endometrial arterioles, in accordance with previously published immunohistological data. In addition, however, they revealed nucleic acids in cellular debris within endometrial glands and diffusing across the placenta at sites of microcotyledonary infarction. Perivascular leucocytes were generally negative for viral DNA, despite ...
Savill MG, Murray SR, Scholes P, Maas EW, McCormick RE, Moore EB, Gilpin BJ.Rhodococcus coprophilus, a natural inhabitant of herbivore faeces, has been suggested as a good indicator of animal (as opposed to human) faecal contamination of aquatic environments. However, conventional detection methods limit its use for this as they require up to 21 days to obtain a result. In this paper an optimised method for extracting R. coprophilus DNA from faecal samples is described. PCR and 5'-nuclease (TaqMan) PCR methods were developed to allow the detection and enumeration of R. coprophilus in faecal samples within 2-3 days. Both PCR methods targeted the 16S rRNA gene, producin...
Looft C, Paul S, Philipp U, Regenhard P, Kuiper H, Distl O, Chowdhary BP, Leeb T.Defensins are a family of evolutionary ancient antimicrobial peptides consisting of three sub-families: alpha-, beta- and theta-defensins. This investigation was focused on the genomic characterization of equine beta-defensins and the investigation of the potential clustering of beta-defensin genes in the equine genome. Six genomic BAC clones were isolated from the CHORI-241 library and one of these was mapped by FISH to ECA 27q17. This location was confirmed by RH-mapping. The contiguous 212 kb sequence of this clone was determined. Sequence analysis revealed the identification of ten pseudog...
Phumoonna T, Barton MD, Vanniasinkam T, Heuzenroeder MW.Rhodococcus equi remains a significant bacterial pathogen, causing severe pyogranulomatous pneumonia in foals aged 1-3 months. There is no effective vaccine currently available for the prevention of R. equi pneumonia. DNA vaccines are known to offer specific advantages over conventional vaccines. The aim of this study was to demonstrate efficacy of our recombinant DNA vaccine candidates, namely pcDNA3-Re1, pcDNA3-Re3 and pcDNA3-Re5 by combining a heat shock protein GroEL2 to a virulence-associated protein A (VapA) from R. equi to protect C3H/He mice against the R. equi infection. VapA was show...
Adaszek Ł, Winiarczyk S, Łukaszewska J.The study was aimed at determining the cause of a disease in a horse exhibiting symptoms of fever, joint effusion, weakness, and extravasations on the mucous membranes. Blood was drawn from the animal for haematological and biochemical molecular tests. The PCR technique revealed the presence of 16S RNA Ehrlichia spp. genetic material in the blood samples. DNA amplification by means of primers EHR 521 and EHR 747 gave a product with a volume of 247 bp.The sequence of the PCR product obtained showed a 97.6% similarity with a sequence of a fragment of 16S RNA Ehrlichia phagocytophila, gene number...
Studdert MJ, Crabb BS, Ficorilli N.The restriction endonuclease DNA fingerprints of 57 isolates of equine herpesvirus 1 (EHV1; equine abortion virus) from abortion, perinatal foal mortalities and encephalitis from 15 epidemics that occurred in Australasia between 1975 and 1989 were examined using the enzymes Bam HI, EcoRI and Bgl II. There was a remarkable degree of uniformity in the restriction patterns; mobility differences were observed in only 14 of 52 (27%) of the fragments. Twelve of these 14 fragments were located within the repeat structures that bracket the unique short region of the genome or were located at the left ...
Borchers K, Frölich K, Ludwig H.In blood samples of seven captive equid species from four German zoos EHV-1 specific antibodies were detected in 76% and EHV-4 specific antibodies in 73% of the 55 animals, whereas 93% were tested positive for EHV-2 and EHV-5, respectively. In only one blood sample from a Przewalski's wild horse EHV-4 DNA was amplified by PCR. From seven Przewalski's wild horses EHV-2, and from another one EHV-5 was isolated by cocultivation. The identity of the virus isolates was verified by PCR and restriction enzyme digestion.
Wilkin T, Hamilton NA, Cawley AT, Bhat S, Baoutina A.The term 'gene doping' is used to describe the use of any unauthorized gene therapy techniques. We developed a test for five likely candidate genes for equine gene doping: , , , and . The test is based on real-time polymerase chain reaction (PCR) and includes separate screening and confirmation assays that detect different unique targets in each transgene. For doping material, we used nonviral (plasmid) and viral (recombinant adeno-associated virus) vectors carrying complementary DNA for the targeted genes; the vectors were accurately quantified by digital PCR. To reduce non-specific amplific...
Flack N, Hughes L, Cassens J, Enriquez M, Gebeyehu S, Alshagawi M, Hatfield J, Kauffman A, Brown B, Klaeui C, Mabrouk IF, Walls C, Yeater T, Rivas A....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 f...
Diekmann I, Krücken J, Kuzmina TA, Bredtmann CM, Louro M, Kharchenko VA, Tzelos T, Matthews JB, Madeira de Carvalho LM, von Samson-Himmelstjerna G.Morphologically, 64 strongylid species have been described in equines. Co-infections are common, with up to 29 species reported in a single horse. Morphological identification of these species is time consuming and requires expert knowledge due to their similar appearance. Therefore, non-invasive identification methods are needed. DNA barcoding offers a rapid and reliable tool for species identification and the discovery of cryptic species for these most common parasitic nematodes of equines. In total, 269 COI and 312 ITS-2 sequences from 27 equine Strongylidae species, including sequences fro...
Ahedor B, Otgonsuren D, Zhyldyz A, Guswanto A, Ngigi NMM, Valinotti MFR, Kothalawala H, Kalaichelvan N, Silva SSP, Kothalawala H, Acosta TJ....Theileria equi causes equine piroplasmosis, an economically significant disease that affects horses and other equids worldwide. Based on 18S ribosomal RNA (18S rRNA sequences), T. equi can be classified into five genotypes: A, B, C, D, and E. These genotypes have implications for disease management and control. However, no conventional polymerase chain reaction (PCR) assays are available to differentiate the genotypes of T. equi. To overcome this limitation, we developed and evaluated PCR assays specific for the detection of each T. equi genotype. Methods: A pair of forward and reverse primers...
Bull KE, Hodgkinson J, Allen K, Poissant J, Peachey LE.Cyathostomins are the most abundant equid endoparasites globally. There are approximately fifty cyathostomin species and, whilst they occupy distinct niches within the large intestine, they are generally considered to share similar characteristics in terms of pathogenicity and response to drug treatment. There are three classes of anthelmintic licensed in the UK to treat cyathostomins (benzimidazoles, tetrahydropyrimidines and macrocyclic lactones) and cases of resistance have been documented for all classes. Previously, faecal egg count reduction tests (FECRT) on four UK Thoroughbred studs re...
