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Topic:Molecular biology
Molecular biology in horses involves the study of molecular processes and genetic mechanisms that underpin equine physiology and health. This field encompasses the analysis of DNA, RNA, proteins, and other biomolecules to understand gene expression, genetic variation, and cellular functions in horses. Techniques such as genomic sequencing, gene expression profiling, and molecular diagnostics are employed to explore topics like hereditary diseases, performance traits, and immune responses in equines. This page assembles peer-reviewed research studies and scholarly articles that investigate the molecular biology of horses, focusing on genetic research, molecular techniques, and their applications in equine science.
Isolation and identification of Mycobacterium avium subspecies silvaticum from a horse. Routine cultivation methods are able to distinguish between isolates of the Mycobacterium avium and the Mycobacterium tuberculosis complex. However, molecular tools are needed to further identify the several subspecies in the M. avium complex, especially for the subspecies avium and silvaticum. A rapid technique using HhaI restriction digestion of a 349 bp amplification product of the 85B antigen (α-antigen) gene was used for the identification of M. avium subsp. silvaticum in a three-year-old gelding presenting with caseous, necrotizing, granulomatous lesions. The result was confirmed by seq...
Monoclonal antibodies to equine CD23 identify the low-affinity receptor for IgE on subpopulations of IgM+ and IgG1+ B-cells in horses. CD23, also called FcεRII, is the low-affinity receptor for IgE and has first been described as a major receptor regulating IgE responses. In addition, CD23 also binds to CD21, integrins and MHC class II molecules and thus has a much wider functional role in immune regulation ranging from involvement in antigen-presentation to multiple cytokine-like functions of soluble CD23. The role of CD23 during immune responses of the horse is less well understood. Here, we expressed equine CD23 in mammalian cells using a novel IL-4 expression system. Expression resulted in high yield of recombinant IL-4/...
Cytokine production and proliferation upon in vitro oligodeoxyribonucleotide stimulation of equine peripheral blood mononuclear cells. Synthetic oligodeoxyribonucleotides (ODN) may prove useful immune modulators in equine medicine. It is however important to assess the effects of each specific ODN in the species it is intended to be used in. The present study therefore aimed to evaluate some ODN for induction of cytokine production; i.e. type I interferons (IFN), IFN-γ, tumor necrosis factor-α (TNF-α) and transforming growth factor-β (TGF-β), and proliferation of equine peripheral blood mononuclear cells (PBMC). A panel of four ODN containing unmethylated cytosine-guanosine sequences (CpG) was used: ODN 1 and ODN 8 repre...
Equine peripheral blood-derived mesenchymal stem cells: isolation, identification, trilineage differentiation and effect of hyperbaric oxygen treatment. Two studies report variability in proliferation and limited adipocyte differentiation of equine peripheral blood-derived adult mesenchymal stem cells, thus casting doubt on their adipogenic potential. Peripheral blood can be a valuable source of adult mesenchymal stem cells if cell culture conditions permissive for their adherence, proliferation and differentiation are defined. Hyperbaric oxygen treatment has been reported to mobilise haematopoietic progenitor stem cells into the peripheral blood in humans and mice, but similar experiments have not been done in horses. Objective: To optimise c...
Isolation and characterization of a novel indigenous intestinal N4-related coliphage vB_EcoP_G7C. 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...
Mass spectral measurements of the apoHDL in horse (Equus caballus) cerebrospinal fluid. As a continuation of our proteogenomic studies of equine apolipoproteins, we have obtained molecular masses for several of the apolipoproteins associated with the HDL in horse cerebrospinal fluid (CSF). Using electrospray-ionization mass spectrometry (ESI-MS), we report on values for apolipoproteins, A-I and A-II, as well as acylated apoA-I. In comparison with our previously published data on equine plasma apolipoproteins, there appears to be a higher percentage of acylated apoA-I in the CSF than in plasma. As was the case in plasma, apoA-II circulates as a homodimer. These studies also reveal...
EcPV2 DNA in equine genital squamous cell carcinomas and normal genital mucosa. 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...
Direct electrochemistry of novel affinity-tag immobilized recombinant horse heart cytochrome c. During the last decade protein electrochemistry at miniaturized electrodes has become important not only for functional studies of the charge transfer properties of redox proteins but also for fostering the development of sensitive biosensor and bioelectronic devices. One of the major challenges in this field is the directed coupling between electronic and biologically active components. A prerequisite for a fast and reversible electron transfer between electrode and protein is that the protein can be bound to the electrode in a favourable orientation. We examined electrostatic and bioaffinity...
Mitochondrial genomes from modern horses reveal the major haplogroups that underwent domestication. 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 ...
Differential gene expression of CYP3A isoforms in equine liver and intestines. Recently, seven CYP3A isoforms - CYP3A89, CYP3A93, CYP3A94, CYP3A95, CYP3A96, CYP3A97 and CYP129 - have been isolated from the horse genome. In this study, we have examined the hepatic and intestinal gene expression of these CYP3A isoforms using TaqMan probes. We have also studied the enzyme activity using luciferin-isopropyl acetal (LIPA) as a substrate. The results show a differential gene expression of the CYP3A isoforms in the liver and intestines in horses. In the liver, CYP3A89, CYP3A94, CYP3A96 and CYP3A97 were highly expressed, while in the intestine there were only two dominating isof...
Evaluation of tyrosinase expression in canine and equine melanocytic tumors. 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...
Altered expression of talin 1 in peripheral immune cells points to a significant role of the innate immune system in spontaneous autoimmune uveitis. The molecular mechanism which enables activated immune cells to cross the blood-retinal barrier in spontaneous autoimmune uveitis is yet to be unraveled. Equine recurrent uveitis is the only spontaneous animal model allowing us to investigate the autoimmune mediated transformation of leukocytes in the course of this sight threatening disease. Hypothesizing that peripheral blood immune cells change their protein expression pattern in spontaneous autoimmune uveitis, we used DIGE to detect proteins with altered abundance comparing peripheral immune cells of healthy and ERU diseased horses. Among ...
Replication and fine-mapping of a QTL for recurrent airway obstruction in European Warmblood horses. Recurrent airway obstruction (RAO), or 'heaves', is a common performance-limiting allergic respiratory disease of mature horses. It is related to sensitization and exposure to mouldy hay and has a familial basis with a complex mode of inheritance. In a previous study, we detected a QTL for RAO on ECA 13 in a half-sib family of European Warmblood horses. In this study, we genotyped additional markers in the family and narrowed the QTL down to about 1.5 Mb (23.7-25.2 Mb). We detected the strongest association with SNP BIEC2-224511 (24,309,405 bp). We also obtained SNP genotypes in an independent...
Identification and characterization of equine herpesvirus type 1 pUL56 and its role in virus-induced downregulation of major histocompatibility complex class I. Major histocompatibility complex class I (MHC-I) molecules play an important role in host immunity to infection by presenting antigenic peptides to cytotoxic T lymphocytes (CTLs), which recognize and destroy virus-infected cells. Members of the Herpesviridae have developed multiple mechanisms to avoid CTL recognition by virtue of downregulation of MHC-I on the cell surface. We report here on an immunomodulatory protein involved in this process, pUL56, which is encoded by ORF1 of equine herpesvirus type 1 (EHV-1), an alphaherpesvirus. We show that EHV-1 pUL56 is a phosphorylated early protein w...
The genetic origin and history of speed in the Thoroughbred racehorse. Selective breeding for speed in the racehorse has resulted in an unusually high frequency of the C-variant (g.66493737C/T) at the myostatin gene (MSTN) in cohorts of the Thoroughbred horse population that are best suited to sprint racing. Here we show using a combination of molecular- and pedigree-based approaches in 593 horses from 22 Eurasian and North-American horse populations, museum specimens from 12 historically important Thoroughbred stallions (b.1764-1930), 330 elite-performing modern Thoroughbreds and 42 samples from three other equid species that the T-allele was ancestral and there...
Detection, molecular characterization and phylogenetic analysis of full-length equine infectious anemia (EIAV) gag genes isolated from Shackleford Banks wild horses. The genetically distinct wild horse herds inhabiting Shackleford Banks, North Carolina are probably the direct descendents of Spanish stock abandoned after failed attempts to settle mid-Atlantic coastal regions of North America in the Sixteenth Century. In a 1996 island survey, 41% of the gathered horses were discovered seropositive for Equine Infectious Anemia Virus (EIAV) with additional cases identified in 1997 and 1998. As a result of their unique genetic heritage, EIAV seropositive individuals identified in the two latter surveys were transferred to a quarantine facility on the mainland. ...
Identification of gonadal tissue in cryptorchid stallion can be improved by molecular biological analysis – a case report. Surgically removed testicular tissue in cryptorchid stallions is sometimes difficult to identify because of morphological and histological malformation. Therefore, a sure method to characterise the removed tissue is required. A 2-year-old Haflinger stallion was castrated after diagnosis of cryptorchidism to remove the left intra-abdomnial testis. Intra-operative exploration of the abdominal cavity revealed a firm, dysmorphic structure, which could not be identified as testis based on macroscopic anatomy. The removed tissue was Bouin-fixed and paraffin-embedded for histological examination. We ...
Sequence variations in equine candidate genes For XX and XY inherited disorders of sexual development. Inherited disorders of sexual development (DSD) cause sterility and infertility in horses. Mutations causing such disorders have been identified in other mammals, but there is little information on the molecular causes in horses. While the equine genome sequence has made it possible to identify candidate genes, additional tools are needed to routinely screen them for causative mutations. In this study, we designed a screening panel of polymerase chain reaction primer pairs for 15 equine genes. These are the candidate genes for testicular or ovotesticular XX DSD and XY DSD, the latter of which ...
Antibodies to a novel leptospiral protein, LruC, in the eye fluids and sera of horses with Leptospira-associated uveitis. Screening of an expression library of Leptospira interrogans with eye fluids from uveitic horses resulted in identification of a novel protein, LruC. LruC is located in the inner leaflet of the leptospiral outer membrane, and an lruC gene was detected in all tested pathogenic L. interrogans strains. LruC-specific antibody levels were significantly higher in eye fluids and sera of uveitic horses than healthy horses. These findings suggest that LruC may play a role in equine leptospiral uveitis.
Genomic characterization of the Taylorella genus. 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...
Characterization of the cDNA and genomic DNA sequence encoding for the platelet integrin alpha IIB and beta III in a horse with Glanzmann thrombasthenia. 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...
The biology of equine mesenchymal stem cells: phenotypic characterization, cell surface markers and multilineage differentiation. Mesenchymal stem cells (MSCs) are multipotent stem cells that can give rise to a range of connective tissue cells including osteoblasts, chondrocytes and adipocytes. MSCs have been isolated from humans and a variety of animal species including rodents, dogs, horses and rabbits. There is currently no consensus on how these cells are identified and characterized. This is partly due to the lack of standardized specific cell surface markers for MSCs. The aim of this review is to examine the literature on equine MSCs and establish whether there is a well-defined phenotype for these cells. Equine MS...
Comparison of equine tendon- and bone marrow-derived cells cultured on tendon matrix with or without insulin-like growth factor-I supplementation. To compare in vitro expansion, explant colonization, and matrix synthesis of equine tendon- and bone marrow-derived cells in response to insulin-like growth factor-I (IGF-I) supplementation. Methods: Cells isolated from 7 young adult horses. Methods: Tendon- and bone marrow-derived progenitor cells were isolated, evaluated for yield, and cultured on autogenous cell-free tendon matrix for 7 days. Samples were analyzed for cell viability and expression of collagen type I, collagen type III, and cartilage oligomeric matrix protein mRNAs. Collagen and glycosaminoglycan syntheses were quantified ov...
Emergence of novel equine arteritis virus (EAV) variants during persistent infection in the stallion: origin of the 2007 French EAV outbreak was linked to an EAV strain present in the semen of a persistently infected carrier stallion. During the summer of 2007, an outbreak of equine viral arteritis (EVA) occurred in Normandy (France). After investigation, a link was suggested between an EAV carrier stallion (A) and the index premise of the outbreak. The full-length nucleotide sequence analysis of a study reference strain (F27) isolated from the lung of a foal revealed a 12,710 nucleotides EAV genome with unique molecular hallmarks in the 5'UTR leader sequence and the ORF1a sequence encoding the non-structural protein 2. The evolution of the viral population in the persistently infected Stallion A was then studied by cloning...
Advanced glycation endproducts in horses with insulin-induced laminitis. Advanced glycation endproducts (AGEs) have been implicated in the pathogenesis of cancer, inflammatory conditions and diabetic complications. An interaction of AGEs with their receptor (RAGE) results in increased release of pro-inflammatory cytokines and reactive oxygen species (ROS), causing damage to susceptible tissues. Laminitis, a debilitating foot condition of horses, occurs in association with endocrine dysfunction and the potential involvement of AGE and RAGE in the pathogenesis of the disease has not been previously investigated. Glucose transport in lamellar tissue is thought to be l...
Expression, refolding and preliminary X-ray crystallographic analysis of equine MHC class I molecule complexed with an EIAV-Env CTL epitope. In order to clarify the structure and the peptide-presentation characteristics of the equine major histocompatibility complex (MHC) class I molecule, a complex of equine MHC class I molecule (ELA-A1 haplotype, 7-6 allele) with mouse β(2)-microglobulin and the cytotoxic T lymphocyte (CTL) epitope Env-RW12 (RVEDVTNTAEYW) derived from equine infectious anaemia virus (EIAV) envelope protein (residues 195-206) was refolded and crystallized. The crystal, which belonged to space group P2(1), diffracted to 2.3 Å resolution and had unit-cell parameters a = 82.5, b = 71.4, c = 99.8 Å, β = 102.9°. T...
Characterization of cytochrome b diversity in Chinese domestic horses. 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...
Complete molecular genome analyses of equine rotavirus A strains from different continents reveal several novel genotypes and a largely conserved genotype constellation. In this study, the complete genome sequences of seven equine group A rotavirus (RVA) strains (RVA/Horse-tc/GBR/L338/1991/G13P[18], RVA/Horse-wt/IRL/03V04954/2003/G3P[12] and RVA/Horse-wt/IRL/04V2024/2004/G14P[12] from Europe; RVA/Horse-wt/ARG/E30/1993/G3P[12], RVA/Horse-wt/ARG/E403/2006/G14P[12] and RVA/Horse-wt/ARG/E4040/2008/G14P[12] from Argentina; and RVA/Horse-wt/ZAF/EqRV-SA1/2006/G14P[12] from South Africa) were determined. Multiple novel genotypes were identified and genotype numbers were assigned by the Rotavirus Classification Working Group: R9 (VP1), C9 (VP2), N9 (NSP2), T12 (NSP3), ...
Comparison of cytokine mRNA expression in the bronchoalveolar lavage fluid of horses with inflammatory airway disease and bronchoalveolar lavage mastocytosis or neutrophilia using REST software analysis. The pathophysiology of inflammatory airway disease (IAD) is unknown, but in some cases involves the accumulation of mast cells, neutrophils, or both in the bronchoalveolar lavage fluid (BALF). The objective of this study was to characterize cytokine gene expression in the BALF cells of horses with IAD, including a comparison of cytokine gene expression between IAD horses with increased BALF mast cells (IAD-Mast) or neutrophils (IAD-Neutro). Methods: The mRNA expression of IL-4, IFN-γ, IL-17, IL-8, IL-1β, IL-5, IL-6, IL-10, IL-12p35, and eotaxin-2 was studied by quantitative polymerase chain ...
