Laboratory methods in equine research encompass a variety of techniques and procedures used to analyze biological samples from horses to study health, disease, genetics, and physiology. These methods include hematological analyses, biochemical assays, molecular biology techniques, and microbiological cultures. Commonly utilized laboratory techniques involve blood tests for complete blood count (CBC) and serum chemistry, polymerase chain reaction (PCR) for genetic and infectious disease studies, and enzyme-linked immunosorbent assays (ELISA) for detecting specific proteins or antibodies. These methods provide valuable data that contribute to understanding equine health and disease mechanisms. This page compiles peer-reviewed research studies and scholarly articles that explore the application, development, and outcomes of laboratory methods in the context of equine research.
Balasuriya UB.The primary goals of this chapter are to discuss common viral RNA isolation and purification methods that are routinely used by various diagnostic laboratories, to highlight the advantages and drawbacks of each method, and to identify the most suitable and reliable method to increase the sensitivity and specificity of RT-PCR assays for the detection of equine influenza virus (EIV) in clinical specimens. Our experiences and review of literature show that magnetic bead-based nucleic extraction methods (manual and automatic) work well for isolation and purification of EIV RNA from nasal swab spec...
Chambers TM, Reedy SE.Equine influenza viruses are cultured in embryonated hen eggs, or in mammalian cells, generally Madin-Darby canine kidney (MDCK) cells, using methods much the same as for other influenza A viruses. Mutations associated with host adaptation occur in both eggs and MDCK cells, but the latter show greater heterogeneity and eggs are the generally preferred host. Both equine-1 H7N7 and equine-2 H3N8 viruses replicate efficiently in 11-day-old eggs, but we find that equine-1 viruses kill the embryos whereas equine-2 viruses do not.
Chambers TM, Reedy SE.Serologic tests for equine influenza virus (EIV) antibodies are used for many purposes, including retrospective diagnosis, subtyping of virus isolates, antigenic comparison of different virus strains, and measurement of immune responses to EIV vaccines. The hemagglutination-inhibition (HI), single radial hemolysis (SRH), and serum micro-neutralization tests are the most widely used for these purposes and are described here. The presence of inhibitors of hemagglutination in equine serum complicates interpretation of HI assay results, and there are alternative protocols (receptor-destroying enzy...
Stirn M, Moritz A, Bauer N.Modern automated haematology instruments are capable of performing leukocyte differentials faster, cheaper and with a higher precision than the traditional 100-cell manual differential count. Thus, in human laboratories, criteria are defined for performing a manual review of the blood smear resulting in a marked reduction of manual differential counts. While common in human laboratories, this approach to reducing the number of manual differentials in veterinary laboratories is still not commonly performed. Thus, our aim was to determine the rate and causes of manual leukocyte differentials in ...
Kranenburg LC, Westermann CM, de Sain-van der Velden MG, de Graaf-Roelfsema E, Buyse J, Janssens GP, van den Broek J, van der Kolk JH.Insulin resistance in horses is an emerging field of interest as it is thought to be a contributing factor in the pathogenesis of many equine conditions. Objective: The objectives of the present study were to determine the effects of long-term oral administration of L-carnitine on insulin sensitivity, glucose disposal, plasma leptin concentrations and acylcarnitine spectrum both in plasma and urine. Methods: Six 3-year-old healthy warmblood geldings were used. In a double blind 2 × 2 Latin square design at a dosage of 100 mg/kg body weight (BW)/day for 28 days the effects of oral supplementat...
Curcio Bda R, Pereira GR, Antunes LI, Boff AN, dos Santos FC, Lucas T, Nogueira CE, Corcini CD, Liu I, Deschamps JC.In vitro fertilization (IVF) procedures are limited by the inability to mature equine oocytes on in vitro methods. Objective: The aim of this study was to evaluate structural integrity of equine oocytes subjected to vitrification with a synthetic polymer (PVA). Methods: The effect of eGH and its relationship with IGF-I on in vitro maturation (IVM) were evaluated. Compact cumulus oocytes complexes (n=122) were cultured in TCM-199 with eGH, IGF-I or eGH+IGF-I for 30h at 38.5C in air with 5 % CO2. Oocytes were fixed after IVM or subjected to the vitrification protocol. Cryopreserved oocytes were ...
Kinoshita Y, Niwa H, Katayama Y.Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) is a dominant pathogenic bacterium in equine pneumonia. We developed a specific loop-mediated isothermal amplification (LAMP) method, which targets the gene encoding sorbitol-6-phosphate 2-dehydrogenase (sorD), for detecting S. zooepidemicus and examined the clinical efficacies of its use in combination with each of 3 DNA extraction methods easily used by veterinary practitioners, namely the Loopamp PURE DNA Extraction Kit, InstaGene Matrix and a conventional boiling method. The LAMP method plus the Loopamp PURE DNA Extraction Kit gave...
Vackova I, Czernekova V, Tomanek M, Navratil J, Mosko T, Novakova Z.This study aimed to determine whether maternal cell contamination exists in cells derived from equine umbilical cord tissue, a perspective material for cell-based therapies in veterinary medicine. Potential maternal cell contamination was analyzed at DNA level via a set of 16 microsatellite markers in cells originating from the cord tissue of 22 foals. In these cells no maternal cell contamination was detected at a sensitivity level of 0.01%. Our results suggest that equine umbilical cord tissue-derived cells are entirely of fetal origin.
Schurer J, Davenport L, Wagner B, Jenkins E.Fecal samples from wild and domestic carnivores are routinely frozen for three days at -80°C to kill eggs of Echinococcus spp., following recommendations from the World Health Organization (WHO) and World Organization for Animal Health (OIE). This is done to decrease the risk of zoonotic infection with these pathogenic cestodes. In addition, it is often necessary to freeze fecal samples collected for research prior to batch processing by a limited number of personnel, especially large numbers of samples or those collected in remote locations. The effect of freezing on the recovery of endopara...
Moreton J, Malla S, Aboobaker AA, Tarlinton RE, Emes RD.The immune system of the horse has not been well studied, despite the fact that the horse displays several features such as sensitivity to bacterial lipopolysaccharide that make them in many ways a more suitable model of some human disorders than the current rodent models. The difficulty of working with large animal models has however limited characterisation of gene expression in the horse immune system with current annotations for the equine genome restricted to predictions from other mammals and the few described horse proteins. This paper outlines sequencing of 184 million transcriptome sh...
Metz GE, Lorenzón EN, Serena MS, Corva SG, Panei CJ, Díaz S, Cilli EM, Echeverría MG.A peptide-based indirect ELISA was developed to detect antibodies against Equine arteritis virus (EAV). Two peptides for epitope C of protein GP5 and fragment E of protein M were designed, synthesized, purified and used as antigens either alone or combined. Ninety-two serum samples obtained from the 2010 Equine viral arteritis outbreak, analyzed previously by virus neutralization, were evaluated by the ELISA here developed. The best resolution was obtained using peptide GP5. The analysis of the inter- and intraplate variability showed that the assay was robust. The results allow concluding tha...
Bohórquez A, Meana A, Pato NF, Luzón M.Current copro-diagnostic tests for Anoplocephala perfoliata show high variation in their sensitivity and given the morphological similarity of Anoplocephala spp. eggs, this could be related to the presence of Anoplocephala magna alone or co-existing with A. perfoliata. In the present study, coprology was significantly more sensitive (p<0.01) at detecting A. magna than A. perfoliata. This difference was independent of the parasite burden and was greater when testing was limited to horses with mature or gravid tapeworms. A. magna infection was strongly linked to young horses (≤ 2 years). The e...
Balssa F, Fischer M, Bonnaire Y.5(10)-Estrene-3β,17α-diol is an essential reference material for doping analysis in horse-racing laboratories. It is used to detect misuse, for doping purpose, of the pregnancy status in the mare. Its stereoselective synthesis from 17β-estradiol-3-methyl ether (prepared from estrone or 17β-estradiol) was performed in four steps: (1) Mitsunobu inversion of the 17β-alcohol; (2) Birch reduction of the aromatic ring; (3) stereoselective reduction of the 3-ketone via Noyori asymmetric transfer hydrogenation; (4) chemoenzymatic purification.
Bujacz A, Zielinski K, Sekula B.Serum albumin, a protein naturally abundant in blood plasma, shows remarkable ligand binding properties of numerous endogenous and exogenous compounds. Most of serum albumin binding sites are able to interact with more than one class of ligands. Determining the protein-ligand interactions among mammalian serum albumins is essential for understanding the complexity of this transporter. We present three crystal structures of serum albumins in complexes with naproxen (NPS): bovine (BSA-NPS), equine (ESA-NPS), and leporine (LSA-NPS) determined to 2.58 Å (C2), 2.42 Å (P61), and 2.73 Å (P2₁2₁...
Tomczuk K, Kostro K, Szczepaniak KO, Grzybek M, Studzińska M, Demkowska-Kutrzepa M, Roczeń-Karczmarz M.The autopsy of 487 slaughter horses revealed the presence of Anoplocephala perfoliata in 36 animals. The invasions varied in the intensity (3 to 2,069 tapeworms) and in the level of tapeworms' proglottid maturity. Twenty nine horses were found to contain tapeworms with gravid proglottid. Fecal samples collected from the rectum were tested using following techniques: flotation with solution-saturated NaCl, decantation, McMaster's, and modified sedimentation-flotation methods (50 g feces samples, flotation solution-saturated NaCl and sucrose, specific gravity 1.25 g/ml). The number of A. perfoli...
Toth B, Slovis NM, Constable PD, Taylor SD.Bacterial sepsis remains a leading cause of morbidity and mortality in neonatal foals, but accurate diagnostic and prognostic markers are lacking. Adrenomedullin (AM) is a polypeptide with diverse biologic effects on the cardiovascular system that increases in septic humans and laboratory animals. Objective: Plasma AM concentration (p[AM]) is increased in septic neonatal foals compared to sick nonseptic and healthy control foals, and p[AM] is predictive of survival in septic neonatal foals. Methods: Ninety critically ill (42 septic, 48 sick nonseptic) and 61 healthy foals <1 week of age. Metho...
Gaivão MM, Rambags BP, Stout TA.Experimental studies and field surveys suggest that embryonic loss during the first 6 weeks of gestation is a common occurrence in the mare. During the first 2 weeks of development, a number of important cell differentiation events must occur to yield a viable embryo proper containing all three major germ layers (ectoderm, mesoderm, and endoderm). Because formation of the mesoderm and primitive streak are critical to the development of the embryo proper, but have not been described extensively in the horse, we examined tissue development and differentiation in early horse conceptuses using a c...
Ing NH, Forrest DW, Love CC, Varner DD.Stallions are unique among livestock in that, like men, they commonly receive medical treatment for subfertility. In both species, about 15% of individuals have normal semen parameters but are subfertile, indicating a need for novel analyses of spermatozoa function. One procedure for improving fertilizing capability of stallions and men is isolation of dense spermatozoa from an ejaculate for use in artificial insemination. In the current study, dense and less dense spermatozoa were purified by density gradient centrifugation from individual ejaculates from seven reproductively normal adult sta...
de Oliveira FS.Anatomical specimens used in human or veterinary anatomy laboratories are usually prepared with formaldehyde (a cancerous and teratogenic substance), glycerin (an expensive and viscous fluid), or ethanol (which is flammable). This research aimed to verify the viability of an aqueous 30% sodium chloride solution for preservation of anatomical specimens previously fixed with formaldehyde. Anatomical specimens of ruminant, carnivorous, equine, swine and birds were used. All were previously fixed with an aqueous 20% formaldehyde solution and held for 7 days in a 10% aqueous solution of the same ...
Menzies-Gow NJ, Wray H, Bailey SR, Harris PA, Elliott J.Insulin and inflammatory cytokines may be involved in equine laminitis, which might be associated with digital vascular dysfunction. This study determined the effects of TNF-α and insulin on the endothelial-dependent relaxant responses of equine digital blood vessels and on equine digital vein endothelial cell (EDVEC) cGMP production. Methods: Isolated rings of equine digital arteries (EDAs) and veins (EDVs) were obtained and EDVECs were cultured from horses euthanized at an abattoir. Methods: The effect of incubation with TNF-α (10 ng/ml) and/or insulin (1,000 μIU/ml) for 1.5 h or overn...
Sessions-Bresnahan DR, Graham JK, Carnevale EM.IVF in horses is rarely successful. One reason for this could be the failure of sperm to fully capacitate or exhibit hyperactive motility. We hypothesized that the zona pellucida (ZP) of equine oocytes prevents fertilization in vitro, and bypassing the ZP would increase fertilization rates. Limited availability of equine oocytes for research has necessitated the use of heterologous oocyte binding assays using bovine oocytes. We sought to validate an assay using bovine oocytes and equine sperm and then to demonstrate that bypassing the ZP using perivitelline sperm injections (PVIs) with equine ...
Barr L, Getgood A, Guehring H, Rushton N, Henson FM.The aim of this in vitro study was to ascertain the effect of recombinant human Fibroblast Growth Factor-18 (rhFGF18) on the repair response of mechanically damaged articular cartilage. Articular cartilage discs were harvested from healthy mature horses (n = 4) and subjected to single impact load (SIL). The impacted explants, together with unimpacted controls were cultured in modified DMEM ± 200 ng/ml rhFGF18 for up to 30 days. Glycosaminoglycan (GAG) release into the media was measured using the dimethylmethylene blue (DMMB) assay. Aggrecan neopepitope CS846, collagen type II synth...
Agrawal K, Ebel JG, Bischoff K.Most antidoping method development in the equine industry has been for plasma and urine, though there has been recent interest in the analysis of synovial fluid for evidence of doping by intra-articular corticosteroid injection. Published methods for corticosteroid analysis in synovial fluid are primarily singleplex methods, do not screen for all corticosteroids of interest and are not adequately sensitive. The purpose of this study is to develop a rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS-MS) screening method for the detection of four of the most common intra-a...
Stojiljkovic N, Paris A, Garcia P, Popot MA, Bonnaire Y, Tabet JC, Junot C.Horse urine is the medium of choice for the implementation of metabolomic approaches aimed at improving horse doping control. However, drug analysis in this biofluid is a challenging task due to the presence of large amounts of interfering compounds. METHODOLOGY & RESULTS: A comparative study of sample preparation has been conducted to evaluate five sample-preparation methods, namely acetonitrile precipitation, proteinase K hydrolysis, membrane filtration and sample dilution with water by factors of five and 20, for metabolome analysis using liquid chromatography coupled to high resolution...
Morrell JM, Georgakas A, Lundeheim N, Nash D, Davies Morel MC, Johannisson A.Removing most of the seminal plasma (SP) from stallion semen has been shown to improve survival during cooled storage, yet adding small quantities of SP may improve pregnancy rates or cryosurvival. Furthermore, there is considerable controversy about whether the stallion's own SP or heterologous SP produces the best effect, possibly because of the variation between stallions in SP proteins or because some homologous SP remained in the sperm preparation. The SP is removed completely from stallion spermatozoa prepared by colloid centrifugation. Thus, the aim of the present study was (1) to inves...
Textor JA, Murphy KC, Leach JK, Tablin F.To compare fiber diameter, pore area, compressive stiffness, gelation properties, and selected growth factor content of platelet-rich fibrin gels (PRFGs) and conventional fibrin gels (FGs). Methods: PRFGs and conventional FGs prepared from the blood of 10 healthy horses. Methods: Autologous fibrinogen was used to form conventional FGs. The PRFGs were formed from autologous platelet-rich plasma of various platelet concentrations (100 × 10³ platelets/μL, 250 × 10³ platelets/μL, 500 × 10³ platelets/μL, and 1,000 × 10³ platelets/μL). All gels contained an identical fibrinogen concentra...
Hdud IM, Mobasheri A, Loughna PT.The metabolic activity of articular chondrocytes is influenced by osmotic alterations that occur in articular cartilage secondary to mechanical load. The mechanisms that sense and transduce mechanical signals from cell swelling and initiate volume regulation are poorly understood. The purpose of this study was to investigate how the expression of two putative osmolyte channels [transient receptor potential vanilloid 4 (TRPV4) and large-conductance Ca(2+)-activated K(+) (BKCa)] in chondrocytes is modulated in different osmotic conditions and to examine a potential role for MAPKs in this process...
Popot MA, Garcia P, Hubert C, Bolopion A, Bailly-Chouriberry L, Bonnaire Y, Thibaud D, Guyonnet J.Tiludronate is a non-nitrogen-containing biphosphonate drug approved in equine veterinary medicine for the treatment of navicular disease and bone sparvin in horse. Its hydrophilic properties and its strong affinity for the bone have made the control of its use quite difficult. After an initial step of method development in plasma and urine, due to a strong matrix effect and erratic detection in urine, the final method development was conducted in plasma. After addition of (3-trifluoromethylphenyl) thiomethylene biphosphonic acid as internal standard, automated sample preparation consisted of ...
Jamila N, Khairí·ªn M, Yeong KK, Osman H, Murugaiyah V.Context: Garcinia hombroniana Pierre, known as manggis hutan in Malaysia is a rich source of xanthones and benzophenones. Objective: This study was aimed to isolate and characterize potential cholinesterase inhibitors from the extracts of G. hombroniana bark and investigate their interactions with the enzymes. Methods: The dichloromethane extract afforded five triterpenoids which were characterized by NMR and mass spectral techniques. Cholinesterase inhibitory assay and molecular docking were performed to get insight of the inhibitory activity and molecular interactions of the compounds. The c...
Doran G, Hughes K, Rendle D, Edwards S.Pergolide is used to treat pituitary pars intermedia dysfunction (equine Cushing's Disease), a neurodegenerative condition associated with loss of dopaminergic inhibition of the pituitary in horses. After oral administration, only low concentrations of the drug are achieved in plasma, making drug detection and quantification difficult. While direct analysis of plasma using sensitive MS/MS techniques is possible, dirty plasma samples and mobile phase buffers can cause instrumentation to become rapidly incapacitated. A method using LC with fluorescence detection was developed for pergolide analy...
De-Simone SG, Napoleão-Pêgo P, Teixeira-Pinto LA, Melgarejo AR, Aguiar AS, Provance DW.The presence of whole horse IgG in therapeutic snake antivenom preparations of high purity is a contamination that can cause IgE-mediated allergic reactions in patients. In this study, the immunodominant IgE and IgG-binding epitopes in horse heavy chain IgG3 were mapped using arrays of overlapping peptides synthesized directly onto activated cellulose membranes. Pooled human sera from patients with and without horse antivenom allergies were used to probe the membrane. We have demonstrated that, for both cases, individuals produce antibodies to epitopes of sequential amino acids of horse heavy ...
Olave C, Alvarez P, Uberti B, Morales N, Henriquez C, Folch H, Sarmiento J, Moran G.Neutrophils play an important role in the exacerbation and maintenance of severe equine asthma; persistent neutrophil activity and delayed apoptosis can be harmful to surrounding tissues. Tamoxifen (TX) is a nonsteroidal estrogen receptor modulator with immunomodulatory effects and induces early apoptosis of blood and bronchoalveolar lavage neutrophils from horses with acute lung inflammation. This study investigated if the in vitro effects of tamoxifen are produced by its action on nuclear (α and β) and membrane (GPR30) estrogen receptors in healthy equine neutrophils. Results showed that T...
Equine germplasm bank management involves not only the conservation and use of semen doses, in addition it can also be a resource to study stallion semen quality and after thawing semen properties for reproductive purposes. A possible criterion to measure quality may be based on differential gene expression of loci involved during spermatogenesis and sperm quality maturation. The rapid degradation of sperm after thawing affects the integrity and availability of RNA. In this study we have analyzed genes expressed in equine cryopreserved sperm, which provided an adequate amplification, specifici...
Wilson SM, Rakhit S, Murdoch R, Pediani JD, Elder HY, Baines DL, Ko WH, Wong PY.Experiments were undertaken using cultured equine sweat gland epithelial cells that express purine receptors belonging to the P2U subclass which allow the selective agonist uridine triphosphate (UTP) to increase the concentration of intracellular free Ca2+ ([Ca2+]i). Experiments using pertussis toxin (Ptx), which inactivates certain guanine-nucleotide-binding proteins (G-proteins), showed that this response consisted of Ptx-sensitive and Ptx-resistant components, and immunochemical analyses of the G-protein alpha subunits present in the cells showed that both Ptx-sensitive (alpha i1-3) and Ptx...
Giordano A, Rossi G, Pieralisi C, Paltrinieri S.The ADVIA 120 is an automated laser cell counter widely used in veterinary medicine. Although specific software for equine samples is available and validated, only a few reports have been published comparing the ADVIA 120 with other methods for equine hemogram evaluation. Objective: The purpose of this study was to compare the hematologic values and reference intervals obtained on the ADVIA 120 with those obtained on an impedance cell counter and manual differential counts in healthy horses. Methods: EDTA-anticoagulated blood samples were obtained from 114 clinically healthy horses of various ...
Claes A, Ball BA, Troedsson MH, Curry TE, Squires EL, Scoggin KE.The wide variation in circulating anti-Müllerian hormone (AMH) concentrations between mares is attributed to differences in antral follicle count (AFC) which may reflect follicular function. There are few data regarding variations in AFC and associated regulatory factors for AMH in the equine follicle during follicular development. Objective: To examine molecular and hormonal differences in the equine follicle in relation to variations in AFC and circulating AMH concentrations during follicular development and to identify genes co-expressed with AMH in the equine follicle. Methods: Observatio...
Hodgkinson JE, Love S, Lichtenfels JR, Palfreman S, Ramsey YH, Matthews JB.Here, we report evaluation of five oligoprobes designed from intergenic spacer (IGS) region sequences for identification of cyathostomin species. Oligoprobes were designed for identification of Cylicocyclus ashworthi, Cylicocyclus nassatus, Cylicostephanus longibursatus, Cylicostephanus goldi and a fifth probe designed to identify all members of this tribe. PCR amplification of IGS DNA from 16 cyathostomin species allowed sequence comparison and identification of four putative species-specific probes. Southern blotting of amplified products from 16 species showed that all probes were species-s...
Kinoshita Y, Niwa H, Katayama Y.Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) is a dominant pathogenic bacterium in equine pneumonia. We developed a specific loop-mediated isothermal amplification (LAMP) method, which targets the gene encoding sorbitol-6-phosphate 2-dehydrogenase (sorD), for detecting S. zooepidemicus and examined the clinical efficacies of its use in combination with each of 3 DNA extraction methods easily used by veterinary practitioners, namely the Loopamp PURE DNA Extraction Kit, InstaGene Matrix and a conventional boiling method. The LAMP method plus the Loopamp PURE DNA Extraction Kit gave...
Liu Y, Uboh CE, Soma LR, Li X, Guan F, You Y, Chen JW.Multiple drug target analysis (MDTA) used in doping control is more efficient than single drug target analysis (SDTA). The number of drugs with the potential for abuse is so extensive that full coverage is not possible with SDTA. To address this problem, a liquid chromatography tandem mass spectrometric method was developed for simultaneous analysis of 302 drugs using a scheduled multiple reaction monitoring (s-MRM) algorithm. With a known retention time of an analyte, the s-MRM algorithm monitors each MRM transition only around its expected retention time. Analytes were recovered from plasma ...
Bousfield GR, Butnev VY, Butnev VY.The O-glycosylation sites for equine LHss (eLHss) and eCGss were identified by solid-phase Edman degradation of four glycopeptides derived from the C-terminal region. Both subunits were O-glycosylated at the same 12 positions, rather than the 4-6 sites anticipated. These sites were partially glycosylated, with carbohydrate attachment ranging from 20% to 100% for eCGss and from 10% to 100% for eLHss. When the C-terminal peptide containing all but one of the O-linked oligosaccharides was removed by mild acid hydrolysis of either eLHss or eCGss, hybrid hormones could be obtained by reassociating ...
Casella S, Giudice E, Giannetto C, Marafioti S, Piccione G.The purpose of this study was to evaluate in vitro the effects of hydrocortisone and aminophylline on adenosine diphosphate (ADP)-induced platelet aggregation in horses. Blood samples from 30 healthy Thoroughbred horses were collected by via jugular venipuncture to assess platelet aggregation. Platelet-rich and platelet-poor plasma were prepared from all samples by centrifugation and divided into three different aliquots. In the first aliquot, platelet aggregation was measured after platelet activation with 1 µM and 0.5 µM ADP (Group A). In the other two aliquots, the effect of a 10 min prei...
Toombs-Ruane LJ, Riley CB, Kendall AT, Bolwell CF, Benschop J, Rosanowski SM.Decreased efficacy of antimicrobials and increased prevalence of multidrug resistance (MDR) is of concern worldwide. Objective: To describe and analyze bacterial culture and antimicrobial susceptibilities from respiratory samples submitted from young horses (4 weeks to 3 years old). Methods: Samples from 289 horses were submitted to a commercial laboratory. Methods: A retrospective database search of submissions made to a New Zealand veterinary laboratory between April 2004 and July 2014. The results of in vitro susceptibility testing by Kirby-Bauer disc diffusion were described and tabulat...
Zak A, Siwinska N, Slowikowska M, Borowicz H, Kubiak K, Hildebrand J, Popiolek M, Niedzwiedz A.There are no available studies describing the possible resistance of strongyles to ivermectin in horses in Poland. One hundred seventy three horses from nine stud farms from South-Western Poland were studied. The effectiveness of ivermectin was studied on the 14th day after ivermectin administration using the fecal egg count reduction test, and a long-term observation of the egg reappearance period was carried out. The fecal study was carried out using a modified McMaster method, which typically detects 20 eggs per gram of stool. The results were subjected to statistical analysis that enabled ...
Saito S, Yokoyama K, Tamagawa T, Ishiwata I.We succeeded in the derivation and maintenance of pluripotent embryonic stem (ES) cells from equine and bovine blastocysts. These cells expressed markers that are characteristics of mouse ES cells, namely, alkaline phosphatase, stage-specific embryonic antigen 1, STAT 3 and Oct 4. We confirmed the pluripotential ability of these cells, which were able to undergo somatic differentiation in vitro to neural progenitors and to endothelial or hematopoietic lineages. We were able to use bovine ES cells as a source of nuclei for nuclear transfer and we generated cloned cattle with a higher frequency ...
Buss T, Aurich J, Aurich C.In horse breeding, quality assessment of semen before insemination is often requested. Non-laboratory-based techniques for objective analysis of sperm motility are thus of interest. The aim of this study was evaluating a portable device for semen analysis (Ongo sperm test) and its comparison with computer-assisted semen analysis (CASA). Semen was collected from 10 stallions, diluted to 100, 50 and 25 × 106 sperm/ml and analysed for total (TM) and progressive motility (PM). The final sperm concentration influenced total motility analysed by Ongo (p < 0.05) which was higher at 100 × 10...
Klein C, Troedsson M.During the second and third week of pregnancy, the equine conceptus is covered by an acellular glycoprotein capsule. This capsule contains glycoproteins resembling those of the mucin family with sialic acid making up a high proportion of the carbohydrate. Coinciding with conceptus fixation, a marked decline in sialic acid content of the capsule occurs, which has been proposed to contribute to cessation of conceptus mobility. Herein, we describe the expression of neuraminidase 2 (NEU2) by pre-implantation stages of equine conceptus development. NEU2 transcript abundance was examined in conceptu...
Pearce JW, Galle LE, Kleiboeker SB, Turk JR, Schommer SK, Dubielizig RR, Mitchell WJ, Moore CP, Giuliano EA.Equine recurrent uveitis (ERU) is the most frequent cause of blindness in horses worldwide. Leptospira has been implicated as an etiologic agent in some cases of ERU and has been detected in fresh ocular tissues of affected horses. The objective of this study was to determine the presence of Leptospira antigen and DNA in fixed equine ocular tissues affected with end-stage ERU. Sections of eyes from 30 horses were obtained. Controls included 1) 10 normal equine eyes and 2) 10 equine eyes with a nonrecurrent form of uveitis. The experimental group consisted of 10 eyes diagnosed with ERU based on...
Wilson KE, Wilcke JR, Crisman MV, Ward DL, McKenzie HC, Scarratt WK.Objective-To determine whether a limited sampling time method based on serum iohexol clearance (Cl(iohexol)) would yield estimates of glomerular filtration rate (GFR) in clinically normal horses similar to those for plasma creatinine clearance (Cl(creatinine)). Animals-10 clinically normal adult horses. Procedures-A bolus of iohexol (150 mg/kg) was administered IV, and serum samples were obtained 5, 20, 40, 60, 120, 240, and 360 minutes after injection. Urinary clearance of exogenous creatinine was measured during three 20-minute periods. The GFR determined by use of serum Cl(iohexol) and plas...
Caillaud M, Gérard N.We analysed the effect of interleukin-1 on oocyte maturation and on steroid and prostaglandin production by equine granulosa and cumulus cells. In Experiment 1, interleukin-1beta (IL-1beta) was injected into the growing dominant follicle, which was punctured 38 h later. Follicular fluid was assayed for steroids and prostaglandin-F2alpha (PGF2alpha). Granulosa cells were analysed for 3beta-hydroxysteroid dehydrogenase (3beta-HSD), progesterone receptor (PR), cyclooxygenase 1 and 2 (Cox 1 and Cox 2) and steroidogenic acute regulatory protein (StAR) mRNAs. In Experiment 2, cumulus-oocyte complexe...
Suganuma K, Yamasaki S, Molefe NI, Musinguzi PS, Davaasuren B, Mossaad E, Narantsatsral S, Battur B, Battsetseg B, Inoue N.Dourine is caused by Trypanosoma equiperdum via coitus with an infected horse. Although dourine is distributed in Equidae worldwide and is listed as an internationally important animal disease by the World Organization for Animal Health (OIE), no effective treatment strategies have been established. In addition, there are no reports on drug discovery, because no drug screening system exists for this parasite. A new T. equiperdum strain was recently isolated from the genital organ of a stallion that showed typical symptoms of dourine. In the present study, we adapted T. equiperdum IVM-t1 from...
Dahlgren LA, Mohammed HO, Nixon AJ.he treatment of overuse tendon injuries with exogenous growth factors such as insulin-like growth factor-I (IGF-I) may facilitate an improved return to sustained athletic function. The biological effects of IGF-I are exerted under the control of a complex of IGF receptors, binding proteins, and proteases. This IGF system includes a family of six structurally related high-affinity IGF binding proteins (IGFBPs) that protect IGF-I from local proteases and restrict access of IGF-I to its receptor. This study describes the expression of the IGFBPs in flexor tendon after acute injury and during heal...
Hoyles L, Ortman K, Cardew S, Foster G, Rogerson F, Falsen E.Three strains of a Gram-positive, catalase-positive, fermentative, non-lipophilic, previously unknown bacterium were isolated from urogenital samples taken from mares in Scotland (M401624/00/1) and Sweden (VM 2074 and VM 2298(T)). All were deposited with the CCUG with tentative identifications as Corynebacterium spp. The strains were characterized using a polyphasic taxonomic approach. Biochemically, the strains were very similar to each other, but phylogenetically distinct from Corynebacterium species with validly published names (≤95% sequence similarity). rpoB gene sequence data confirmed...
Meyers RA, Hermanson JW.The soleus muscle of horses is rather diminutive with respect to the overall size of adjacent synergist muscles in the hind limb of the horse. Whether or not such a muscle might be vestigial or may be providing some essential function has not been determined. We have studied the horse's soleus muscle using histochemical (ATPase), immunocytochemical (myosin isoform identification), and SDS-PAGE analysis to demonstrate that it is largely composed of 100% type I, presumed slow-twitch fibers. Only one soleus muscle studied (out of 13 adult horses) contained any type II muscle fibers. Given this co...
Jelocnik M, Nyari S, Anstey S, Playford N, Fraser TA, Mitchell K, Blishen A, Pollak NM, Carrick J, Chicken C, Jenkins C.C. psittaci has recently emerged as an equine abortigenic pathogen causing significant losses to the Australian Thoroughbred industry, while Equine herpesvirus-1 (EHV-1) is a well-recognized abortigenic agent. Diagnosis of these agents is based on molecular assays in diagnostic laboratories. In this study, we validated C. psittaci and newly developed EHV-1 Loop Mediated Isothermal Amplification (LAMP) assays performed in a real-time fluorometer (rtLAMP) against the reference diagnostic assays. We also evaluated isothermal amplification using commercially available colorimetric mix (cLAMP), and...
Raudsepp T, Otte K, Rozell B, Chowdhary BP.Three genomic subclones derived from a phage clone containing the equine IGF2 gene were used to FISH map the gene on horse (ECA) and donkey (EAS) metaphase chromosomes. The gene mapped on ECA 12q13 band and is the first locus mapped to this horse chromosome. In donkey the gene mapped very terminal on the long arm of one small submetacentric chromosome that shows almost identical DAPI-banding pattern with ECA12. This is the first locus mapped in donkey genome. Cross species chromosome painting of equine metaphase chromosomes with human Chromosome (Chr) 11-specific probe showed homoeology of thi...
Dumoulin M, Pille F, van den Abeele AM, Boyen F, Boussauw B, Oosterlinck M, Pasmans F, Gasthuys F, Martens A.Standard methods for culturing equine synovial fluid (SF) are often unrewarding. Evidence-based information on the relative efficiency of different systems used for optimisation of isolation of microorganisms from equine SF is lacking. Objective: To compare the results of different culture systems performed in parallel on SF samples from horses clinically diagnosed with synovial sepsis. Methods: Synovial fluid specimens were collected between February 2007 and October 2008 from all horses admitted to a referral hospital that were clinically diagnosed with synovial sepsis and from control horse...
Singh UK, Patel R.In vitro refolding of denatured protein and the influence of the alkyl chain on the refolding of a protein were tested using long chain imidazolium chloride salts, 1-methyl-3-octylimidazolium chloride [Cmim][Cl], and 1-decyl-3-methylimidazolium chloride [Cmim][Cl]. The horse heart cytochrome c (h-cyt c) was denatured by urea and guanidinium hydrochloride (GdnHCl), as well as by base-induced denaturation at pH 13, to provide a broad overview of the overall refolding behavior. The variation in the alkyl chain of the ionic liquids (ILs) showed a profound effect on the refolding of denatured h-cyt...
Hara S, Diesterbeck US, König S, Czerny CP.The present study analyzed equine λ-light chain genes (IGLV and IGLC) transcribed in the horse breeds Rhenish-German Coldblood (RGC) and Hanoverian Warmblood (HW). Primers were generated for the major expressed IGLV subgroup 8. The significant majority of the sequences represented IGLC6/7. In RGC, IGLC1 and IGLC5 were observed in significant higher frequencies than IGLC4. In HW, significant differences were obtained for the transcription of IGLC1 and IGLC5. IGLC4 was not determined in this breed. Five allotypic IGLC1 variants, four allotypic IGLC5 variants, and three allelic as well as two al...
Heras S, Smits K, De Schauwer C, Van Soom A.Global epigenetic reprogramming is considered to be essential during embryo development to establish totipotency. In the classic model first described in the mouse, the genome-wide DNA demethylation is asymmetric between the paternal and the maternal genome. The paternal genome undergoes ten-eleven translocation (TET)-mediated active DNA demethylation, which is completed before the end of the first cell cycle. Since TET enzymes oxidize 5-methylcytosine to 5-hydroxymethylcytosine, the latter is postulated to be an intermediate stage toward DNA demethylation. The maternal genome, on the other ha...
Dechant JE, Symm WA, Nieto JE.To compare agreement between a portable clinical analyzer and laboratory-based bench-top analyzer for analysis of pH, lactate, and glucose concentrations in synovial fluid. Methods: Prospective experimental study. Methods: Clinically normal horses (n=8); 6 horses euthanatized for reasons unrelated to the study; 11 horses that had synoviocentesis for reasons other than sepsis; 7 horses that had synoviocentesis for evaluation of sepsis; and 2 horses without recorded clinical data. Median age of horses was 8 years (range, 1 day to 24 years). Methods: Supernatant from each synovial fluid sample wa...
