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.
Diaz-Espiñeira M, Escolar E, Bellanato J, De La Fuente MA.Atomic spectrometry has been used in 20 samples of equine urinary sabulous deposits in order to detect minor elements accompanying the predominant element, calcium, which is present in the form of calcium carbonate (calcite and/or vaterite). The elements measured have been (besides calcium) magnesium, sodium, potassium, iron, copper and manganese. Phosphates, sulphates and silica are frequently present as minor constituents of equine urinary sabulous deposits and uroliths, but their detection can be difficult by infrared (IR) spectroscopy in the original samples due to overlapping with the ban...
Koupai-Abyazani MR, Esaw B, Laviolette B.Urine and serum samples collected from four standard-bred mares after and oral regimen administration of methocarbamol were extracted and analyzed. The method consisted of enzyme hydrolysis followed by a one-step liquid-liquid extraction, separation on a reversed-phase (RP-18) column, and detection using an ultraviolet (UV) detector. The confirmation was carried out using a liquid chromatography-atmospheric pressure ionization-mass spectrometry (LC-API-MS) system. Maximum methocarbamol concentrations of 1498, 1734, 1547, 2322 micrograms/mL in urine and 4.9, 1.7, and 3.6 micrograms/mL in serum ...
Reubel GH, Studdert MJ.We report the first nucleotide sequence data on equine adenovirus 2 (EAdV2) which corroborate on the molecular level that EAdV2 is distinct from equine adenovirus 1 (EAdV1). Based on sequence homology with Eadv1 the hexon gene of Eadv2 was identified. HindIII restriction fragments containing the hexon and eight other viral genes were cloned into the plasmid pUC19 and the nucleotide sequence of the hexon and the 23K proteinase genes completely determined. Amino acid (aa) comparison of sequence fragments with published adenovirus (AdV) proteins identified the genes for the E1B/19K, IVa2, DNA pol...
Sonea IM, Wilson DV, Bowker RM, Robinson NE.Tachykinins, of which substance P (SP) is the prototype, are neuropeptides which are widely distributed in the nervous systems. In the equine gut, SP is present in enteric nerves and is a powerful constrictor of enteric muscle; in other species, SP is also known to have potent vasodilatory and pro-inflammatory effects. The specific effects of SP are determined by the subtype of receptor present in the target tissue. There are 3 known subtypes of tachykinin receptors, distinguished by their relative affinities for SP and other tachykinins. The distribution of SP binding sites in the equine pelv...
Singh AK, Jiang Y, White T, Spassova D.The performances of a radioimmunoassay method, a chemiluminescent immunoassay method, and a chemiluminescent-enzyme immunoassay method were evaluated for the analysis of cortisol and total thyroxine in blood samples obtained from dogs, cats, horses, and humans (reference samples). The analysis of cortisol in human and animal samples exhibited good precision, linearity, and recovery. The 3 methods gave comparable values for the ACTH-induced increase and the dexamethasone-induced decrease in cortisol concentrations in animal samples. The recoveries of total thyroxine from human samples, analyzed...
Im Hof V, Gehr P, Gerber V, Lee MM, Schürch S.We measured the surface tension in the trachea of the non-anaesthetised horse from the spreading behaviour of fluid drops, using videotracheoscopy. To do this, we placed small oil drops onto the tracheal wall with a thin Teflon tubing inserted into a videocolonoscope used in humans. Either 5 ml of saline (control) or 5 ml of bovine lipid extract surfactant (BLES) at 4 mg/ml were administered. Tracheal surface tension was 31.9 +/- 0.54 mN/m (Mean +/- SEM, n = 30) in the control experiments and 24.5 +/- 0.51 mN/m (Mean +/- SEM, n = 21) in the entire trachea after the administration of BLES. Thes...
Bowles DE, Holden VR, Zhao Y, O'Callaghan DJ.To assess the role of the equine herpesvirus type 1 (EHV-1) ICP0 protein (EICP0) in gene regulation, a variety of molecular studies on the EICP0 gene and gene products of both the attenuated cell culture-adapted Kentucky A (KyA) strain and the Ab4p strain were conducted. These investigations revealed that (i) the ICP0 open reading frame (ORF) of the KyA virus strain is 1,257 bp in size and would encode a protein of 419 amino acids, and in comparison to the ICP0 gene (ORF63) of the Ab4p strain of 1,596 bp (E. A. Telford, M. S. Watson, K. McBride, and A. J. Davison, Virology 189:304-316, 1992), ...
Wang ZW, Nara M, Wang YX, Kotlikoff MI.The effects of sulfhydryl reduction/oxidation on the gating of large-conductance, Ca(2+)-activated K+ (maxi-K) channels were examined in excised patches from tracheal myocytes. Channel activity was modified by sulfhydryl redox agents applied to the cytosolic surface, but not the extracellular surface, of membrane patches. Sulfhydryl reducing agents dithiothreitol, beta-mercaptoethanol, and GSH augmented, whereas sulfhydryl oxidizing agents diamide, thimerosal, and 2,2'-dithiodipyridine inhibited, channel activity in a concentration-dependent manner. Channel stimulation by reduction and inhibit...
Baracca A, Bucchi L, Ghelli A, Lenaz G.A method to prepare coupled submitochondrial particles from horse platelets is described. The method allowed us to study the protonophoric activities of both complex I and complex V following the fluorescence quenching of the monoamine 9-amino-6-chloro-2 methoxyacridine (ACMA), a probe highly sensitive to the generation of a transmembrane delta pH. We carried out a kinetic analysis of each enzyme complex studying the proton translocation and the electron transfer activities of complex I as well as the proton translocation and the ATP hydrolytic activities of complex V. A micromethod to prepare...
Nagatomo H, Tokita Y, Shimizu T.Although it is known that commercialized bovine serum is sometimes contaminated with mycoplasmas, it is not clear whether mycoplasmas can survive in horse serum. In this study, as a preliminary examination of the survival of mycoplasmas inoculated in horse sera, the survivability of 8 strains of 7 mycoplasmas was tested. The results obtained reveal that two strains of M. bovis and M. gallisepticum were found to survive in non-heated and inactivated sera for 94 to 330 days at 30 or 37 degrees C. Three strains of M. bovirhinis, M. gateae and A. laidlawii lived for 7 to 330 days depending upon th...
van Nieuwstadt RA, Henricks PA, Hajer R, van der Meer van Roomen WA, Breukink HJ, Nijkamp FP.This study was undertaken to assess the importance of muscarinic receptor subtypes in equine airway disease. Smooth muscle strips from the mid-cervical portion of the trachea of horses were placed in tissue baths and isometric contractile force was measured. Active force was measured in response to metacholine and the selective muscarinic receptor agonists McN-A-343 (M1-selective) and pilocarpine (M2-selective) in cumulative concentrations (10(-9)M through 10(-3)M), with and without preincubation with three or four concentrations of the selective muscarinic receptor antagonists pirenzepine (M1...
Recio P, López PG, Fernández JL, Garcia-Sacristán A.1. The presence and types of alpha and beta-adrenoceptors in the corpus cavernosum of the horse were studied in vitro by using selected ligands of adrenoceptors and isometric tension recording. 2. Noradrenaline and phenylephrine induced concentration-dependent contractions in corpus cavernosum preparations. B-HT 920 had no effect. 3. Phentolamine and prazosin produced a shift to the right of the dose-response curve of noradrenaline, while the alpha(2)-antagonist, rauwolscine had no effect on the response to noradrenaline. Phenylephrine-evoked contractions of corporal strips were significantly ...
Lepage OM, Eicher R, Uebelhart B, Tschudi P.To evaluate applicability of a human osteocalcin (OC) immunoradiometric assay (IRMA) for use with equine serum and compare it with a bovine radioimmunoassay (RIA) previously proven valid for such samples, and to describe the effect of type and breed of horses on serum OC concentration. Methods: 100 healthy horses of either sex, classified as type I or II (draught or warmblood, respectively). Each type was represented by 2 breed groups, each comprising 25 horses. Methods: Blood samples were collected in the morning, and the serum was separated. Osteocalcin was measured, using commercially avail...
Lind C, Chen J, Byrjalsen I.We developed a rapid, competitive enzyme immunoassay (EIA) for measuring 25-hydroxyvitamin D3 [25(OH)D3] in serum. The EIA was based upon 25(OH)D3-3-hemisuccinate covalently coupled to secondary amino groups grafted onto the polystyrene surface of microtiter wells. Optimal coupling conditions were established, and we found that inclusion of 40 mumol/L chloramine T, an agent not previously described for use in coupling to these plates, resulted in both more reproducible coupling as well as more than a twofold increase in the coupling efficiency. Before EIA, 25(OH)D3 was extracted from the serum...
Richard F, Martinat N, Remy JJ, Salesse R, Combarnous Y.Among all mammalian FSH receptors (FSH-R; including donkey (dk) FSH-R), only horse (hs) FSH-R does not bind hsLH/chorionic gonadotrophin (CG). In order to delineate the structural origin of hsFSH-R specificity precisely, we have cloned dkFSH-R cDNA from donkey testis mRNA by RT-PCR. Transiently expressed dkFSH-R endowed COS-7 cells with both hsLH/CG- and FSH-binding activity, as well as FSH-induced cAMP production. The deduced dkFSH-R amino acid sequence shares 96% identity with the hsFSH-R: notably, in the hormone-binding domain, the specificity of hsFSH-R may be ascribed to only four diverge...
Morozova-Roche LA, Arico-Muendel CC, Haynie DT, Emelyanenko VI, Van Dael H, Dobson CM.Native state 1H NMR resonance assignments for 125 of the 129 residues of equine lysozyme have enabled measurement of the hydrogen exchange kinetics for over 60 backbone amide and three tryptophan indole hydrogen atoms in the native state. Native holo equine lysozyme hydrogen exchange protection factors are as large as 10(6), the most protected residues being located in elements of secondary structure. High exchange protection in the domain interface correlates with the binding of Ca2+ in this region. Equine lysozyme differs from most non-Ca2+ binding lysozymes in forming a highly populated par...
Shao H, Robek MD, Threadgill DS, Mankowski LS, Cameron CE, Fuller FJ, Payne SL.The macrophage tropic lentivirus, equine infectious anemia virus (EIAV), encodes a dUTPase in the pol gene that is required for efficient replication in macrophages. Two naturally occurring variants of the enzyme were expressed as recombinant proteins in Escherichia coli; metal chelate affinity chromatography was used to purify histidine-tagged recombinant enzymes to greater than 80% homogeneity in a single chromatographic step. Biochemical and enzymatic analyses of these preparations suggest that this method yields dUTPase that is suitable for detailed mutational analysis. Specific activities...
Kundu A, Cramer SM.The resolving power of displacement chromatography using low-molecular-weight displacers was investigated using a model mixture containing bovine and horse heart cytochrome c. The linear and nonlinear adsorption behavior of these two proteins was examined in cation-exchange chromatography and shown to be quite similar. Furthermore, an analysis of the dynamic affinity of these proteins indicated extremely similar affinities under displacement conditions. Despite the extreme similarities in the adsorption behavior, displacement chromatography using a protected amino acid displacer resulted in ex...
Popot MA, Houghton E, Ginn A, Jones M, Teale P, Samuels T, Lassourd V, Dunnett N, Cowan DA, Bonnaire Y, Toutain PL.The purpose of the present report was to estimate the population parameters of cortisol concentrations in urine, an endogenous hormone used as a 'doping' agent and for which an international threshold (1.0 micrograms/ml) has been proposed. Two data bases (French and UK) corresponding to 112 and 142 samples, respectively were considered. Urine was collected under specific post competition conditions. Cortisol concentrations were obtained by validated methods (HPLC for the French samples, and GC-MS for UK samples). No difference was observed between the 2 data sets and statistical analyses were ...
Gravance CG, Champion Z, Liu IK, Casey PJ.The evaluation of seminal characteristics is important in the clinical detection of stallion subfertility. Conventional semen evaluation includes subjective determination of sperm concentration, motility, and gross morphology. Due to the subjectivity and variability of the manual morphology assessment, computer automated sperm morphology analyses has been developed. Computer automated sperm morphology analysis was applied in the current study to determine if the morphometric measurements of sperm heads from collected and dismount samples of the same ejaculate were similar. If the post-ejaculat...
Sailleau C, Moulay S, Cruciere C, Laegreid WW, Zientara S.A reverse transcription-polymerase chain reaction (RT-PCR) assay followed by dot-blot hybridisation was used to detect African horse sickness virus (AHSV); the primers employed amplified the S7 gene that encodes the VP7 protein. The RT-PCR assay was compared with virus isolation for detecting AHSV in blood samples form horses experimentally infected with AHSV-4 and AHSV-9. The influence of sample storage and transportation and the effects of two anticoagulants (EDTA and heparin) were also studied. RT-PCR results were obtained within 48 hours as opposed to a minimum of 15 days for virus isolati...
Sarkar P, McIntosh JM, Leavitt R, Gouthro H.Nimesulide is a nonsteroidal anti-inflammatory drug recently detected in equine blood and urine samples taken at the race track. The detection of the drug in a blood sample led to the identification of an unknown thin-layer chromatographic (TLC) spot in track urine samples as a metabolite of nimesulide. Characterization of the unknown TLC spot and comparison with the synthesized compound shows that the unknown TLC spot is a previously unreported equine metabolite of nimesulide. The metabolite was identified as resulting from the reduction of the nitro group on nimesulide to an amino group. Thi...
Moore T, Wilcke J, Chilcoat C, Eyre P, Crisman M.Equine neutrophils (PMN) play a critical role in inflammatory processes in horses. The objective of this study was to characterize equine PMN function ex vivo following stimulation with calcium ionophore A23187 (A23187) and phorbol myristate acetate (PMA). These stimulants trigger different branches of the PMN activation process that occurs in vivo. Equine PMN were isolated from the whole blood of six clinically normal geldings using a one-step discontinuous Percoll gradient technique. Neutrophil aggregation, degranulation, and superoxide anion production were evaluated in assay systems which ...
Birch-Machin I, Rowan A, Pick J, Mumford J, Binns M.The nucleotide sequence of the nonstructural protein NS1 of the influenza virus A/equine 2/Suffolk/89 was determined and found to be 97% identical to that of A/equine 2/Miami/63. A similar level of identity was shown for the deduced NS1 amino acid sequence. The NS1 gene was expressed, in its entirety and in part, as fusion proteins with glutathione S-transferase using the pGEX-3X expression vector. Antibodies to NS1 protein were detected in serum samples from ponies experimentally infected with influenza virus, but not in animals vaccinated with whole inactivated virus or in unprimed control a...
Kumar S, Malhotra DV, Dhar S.The present study aimed to develop Dot-ELISA, complement fixation test (CFT) and capillary tube agglutination test (CAT) for serodiagnosis of Babesia equi infection and to compare their sensitivity with each other. For this study, sequential serum samples were collected from four donkeys experimentally infected with B. equi up to 90 days post infection (P.I.). B. equi antigen was prepared from the blood of a donkey showing more than 80% parasitaemia. Dot-ELISA, CF and CA tests were standardized as per the standard method. While performing CFT, it was observed that CFT standardized for the donk...
Venugopalan CS, Moore RM, Holmes EP, Sedrish SA.To determine the in vitro contractile responses of equine colonic arteries to angiotensin II, histamine, serotonin, norepinephrine, prostaglandin F2 alpha, vasopressin, and a thromboxane-B2-analogue. Methods: The tension generated in colonic arterial rings placed in organ baths with oxygenated Tyrode's solution at 37 degrees C after exposure to the previously mentioned chemical agents was measured using force-transducers interfaced with a polygraph. Methods: Large colon arterial rings collected from eight horses. Methods: The rings were allowed to equilibrate for 45 minutes after applying 2 g ...
Smith KC.Abortion or neonatal disease may follow infection with several alpha, beta and gamma-herpesviruses. The alpha-herpesvirus, equid herpesvirus-1 (EHV-1), causes single or epizootic abortions or neonatal deaths in equids, and the closely related virus EHV-4 causes sporadic equine abortions. In cattle, the alpha-herpesviruses, bovine herpesvirus-1 (infectious bovine rhinotracheitis virus) and bovine herpesvirus-5 (bovine encephalitis virus), and a gamma-herpesvirus, bovine herpesvirus-4, have all been implicated as causes of abortion. In pigs, suid herpesvirus-1 (SHV-1: pseudorabies virus), an alp...
Prieto D, Hernández M, Rivera L, García-Sacristán A, Simonsen U.The distribution of neuropeptide Y (NPY)-immunoreactive (IR) nerves, as well as the functional effects of NPY and the Y1- and Y2-receptor agonists, [Leu31,Pro34]NPY and NPY(13-36), respectively, have been investigated in vitro in both visceral and arterial smooth muscle of the horse intravesical ureter. NPY-IR nerve fibres were widely distributed along the entire length of the ureter, although the intravesical part was the most richly innervated region, and the only one where NPY-IR ganglion cells were found. NPY (10(-7) M) did not affect either basal tone or spontaneous rhythmic contractions ...
Gu X, Meleka-Boules M, Chen CL, Ceska DM, Tiffany DM.A capillary electrophoresis (CE) and a solid-phase extraction method was developed for the determination of flunixin in equine urine and serum. The suitable CE run conditions were described. The factors affecting flunixin recovery rates were investigated and optimum solid-phase extraction conditions for flunixin in equine urine and serum were established. Limits of detection and quantitation were 3.4 and 5.6 ng/ml for serum and 16.9 and 33.1 ng/ml for urine, respectively. The recoveries exceeded 96% for urine and 79% for serum. Urine samples from race horses and urine and serum samples from a ...
Hobbs KJ, Le Sueur ANV, Hallowell K, Martin E, Sheats MK, Ueda Y.An 8-year-old American Quarter Horse gelding was treated with extracorporeal hemoperfusion (HP) therapy for treatment of Clostridioides difficile (C. difficile) colitis-induced systemic inflammatory response syndrome (SIRS). The gelding developed C. difficile associated peracute colitis and severe SIRS as evidenced by a positive fecal C. difficile PCR and tachypnea, tachycardia, fever, neutropenia, altered mucous membrane color, and hyperlactatemia. Concurrent acute kidney injury in the horse limited the use of routine anti-inflammatory and anti-lipopolysaccharide treatments, including flunixi...
Becerro-Rey L, Martín-Cano FE, Fabres Robaina Sancler-Silva Y, Gil MC, Ortega-Ferrusola C, Aparicio IM, Gaitskell-Phillips G, da Silva-Álvarez E....Conservation of equine semen in the liquid state is a central procedure in horse breeding and constitutes the basis of associated reproductive technologies. The intense mitochondrial activity of the stallion spermatozoa increases oxidative stress along the storage period, leading to sperm demise within 24-48 h of storage, particularly when maintained at room temperature. Recently, the relationship between metabolism and oxidative stress has been revealed. The study aimed to extend the period of conservation of equine semen, at room temperature through modification of the metabolites present i...
Li X, Du M, Liu Y, Wang M, Shen Y, Xing J, Zhang L, Zhao Y, Bou G, Bai D, Dugarjaviin M, Xia W.During follicular development, changes in the composition of the follicular fluid are synchronized with the development of oocytes. Our aim was to screen the key factors affecting oocyte maturation and optimize the in vitro culture protocol by understanding the changes of proteins and metabolites in follicular fluid. Follicles are divided into three groups according to their diameter (small follicle fluid (SFF): 10 mm < d < 20 mm; medium follicle fluid (MFF): 20 mm < d < 30 mm; large follicle fluid (LFF): 30 mm < d). Proteins and metabolites from the follicular fluid w...
Serteyn D, Storms N, Mouithys-Mickalad A, Sandersen C, Niesten A, Duysens J, Graide H, Ceusters J, Franck T.Laminitis in horses is a crippling condition marked by the deterioration of the dermal-epidermal interface, leading to intense lameness and discomfort, often necessitating euthanasia. This study aimed to establish an in vitro model of laminitis using a continuous keratinocyte cell line exposed to anoxia-reoxygenation and an activated neutrophil supernatant. A significant decrease in the keratinocytes' metabolism was noted during the reoxygenation period, indicative of cellular stress. Adding muscle-derived mesenchymal stem/stromal cells during the reoxygenation demonstrated a protective effect...
Share ER, Mastellar SL, Suagee-Bedore JK, Eastridge ML.The measurement of fecal cortisol/corticosterone metabolites (FCMs) is often used to quantify the stress response. The sampling method is relatively non-invasive, reduces concern for elevation of cortisol from the sampling method, and has been shown to measure cortisol more consistently without the daily diurnal rhythm observed in blood. Commercial ELISA (enzyme-linked immunoassay) kits offer benefits over previously validated immunoassay methods but lack validation. The objective of this study was to evaluate a commercial ELISA kit (Arbor Assays DetectX Cortisol ELISA kit, K003-H1, Ann Arbor,...
Steuer A, Fritzler J, Boggan S, Daniel I, Cowles B, Penn C, Goldstein R, Lin D.Current methods for obtaining fecal egg counts in horses are often inaccurate and variable depending on the analyst's skill and experience. Automated digital scanning of fecal sample slides integrated with analysis by an artificial intelligence (AI) algorithm is a viable, emerging alternative that can mitigate operator variation compared to conventional methods in companion animal fecal parasite diagnostics. Vetscan Imagyst is a novel fecal parasite detection system that uploads the scanned image to the cloud where proprietary software analyzes captured images for diagnostic recognition by a d...
Martin-Pelaez S, Fuente A, Takahashi K, Perez IT, Orozco J, Okada CTC, Neto CR, Meyers S, Dini P.Intracytoplasmic sperm injection (ICSI) is the current clinical practice for the in vitro production of equine embryos. The use of conventional fertilization methods such as in vitro fertilization (IVF), has historically been associated with poor success in horses. However, recent improvements have led to better outcomes with IVF, though only when using fresh semen, which limits its use in clinical practice. IVF remains in its infancy in equine reproduction, and several unknowns remain about the technique. One significant gap in knowledge concerns the morphokinetics of IVF embryos and how they...
Samper JC, Hernandez Aviles JC, Ramirez-Agamez LF, Love CC, Gonzalez-Marin C, Fleury P, Dini P, De La Fuente A, Foss R, Campos FL, Ross PJ.The application of sex-sorted semen in horses has historically lagged the bovine industry due to differences in the reproductive physiology and grater variability in equine semen quality. Recent advancements, including SuperGen™ sorters and Ultraplus™ semen processing methods developed by STgenetics® (Navasota, Texas), have significantly improved the efficiency of the sex-sorting process and reduced the sperm damage previously reported. To facilitate the broader use sex-sorted semen in the equine industry, it is essential to address challenges such as shipping cooled semen to a central so...
Baghdadi HBA, Abdelsalam M, Attia MM.Strongylus vulgaris, a devastating parasitic nematode in equids, causes life-threatening verminous aneurysms that are challenging to diagnose early. This study pioneered integrating nanotechnology into an indirect enzyme-linked immunosorbent assay (i-ELISA) system to enhance the sensitivity and specificity for detecting S. vulgaris larval antigens in equine serum samples, with PCR confirmation of the species. A conventional i-ELISA and an innovative nano-based ELISA were developed using excretory-secretory antigens from adult S. vulgaris worms. The nano-ELISA incorporated gold nanoparticles (1...
Crisà A, Cardinali I, Giontella A, Silvestrelli M, Lancioni H, Buttazzoni L.Lipizzan is a famous horse breed dating back to 1580 when the original stud of Lipica was established by the Hasburg Archduke Charles II. Currently, the Italian State Stud of Lipizzan Horses (ASCAL) is a conservation nucleus managed through strict mating rules where mitochondrial DNA sequences are used to verify the correct assignment of mares to a historical pedigree maternal lineage. Here, we analyzed the D-loop sequences of Lipizzan horses from the ASCAL in Monterotondo (Rome, Italy) in order to confirm their pedigree assignment to known female founder families. The concurrent investigation...
Barot D, Usimaki A, Linardi RL, Arensberg CM, Ortved KF.Quantify the concentration of α2-macroglobulin (A2M), immunomodulatory cytokines, and TGF-β1 factors in 4 commercially available autologous blood-based products including conditioned A2M (CA2M; Alpha2EQ; Astaria Global), autologous protein solution (APS; Pro-Stride; Zoetis), platelet-rich plasma (PRP; Restigen; Zoetis), and autologous conditioned plasma (ACP; Arthrex ACP). We hypothesized that CA2M would have higher concentrations of A2M and lower concentrations of cytokines and growth factors compared to APS, PRP, and ACP. Unassigned: Blood was obtained from 6 healthy, adult horses and proc...
Kublicka A, Lorek D, Mikołajczyk-Martinez A, Chodaczek G, Chwirot A, Bażanów B, Matczuk AK.The process of viral entry into host cells is crucial for the establishment of infection and the determination of viral pathogenicity. A comprehensive understanding of entry pathways is fundamental for the development of novel therapeutic strategies. Standard techniques for investigating viral entry include confocal microscopy and flow cytometry, both of which provide complementary qualitative and quantitative data. Imaging flow cytometry, which integrates the advantages of both methodologies, offers significant potential in virological studies. In this investigation, we employed imaging flow ...
Byambaragchaa M, Park SH, Park MH, Kang MH, Min KS.Equine chorionic gonadotropin (eCG) hormone, comprising highly glycosylated α- and β-subunits, elicits responses similar to follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in non-equid species. This study aimed to establish a mass production of recombinant eCG (rec-eCG) using CHO DG44 cells. Single-chain rec-eCG β/α was expressed in CHO DG44 cells. FSH- and LH-like activities were evaluated in CHO-K1 and HEK 293 cells expressing the equine LH/CG receptor (eLH/CGR), rat LH/CGR (rLH/CGR), and rFSHR. pERK1/2 activation and β-arrestin 2 recruitment were assessed in PathHunter ...
Nielsen MK, Bartholdy ID, Kristensen KS, Borrye JC, Meilvang KSS, Rendtorff CEK, Hjortflod MD, Fuglbjerg V, Karlsson M, Petersen HH, Toft K....Equine strongylids are ubiquitous and can cause severe health issues. Anthelmintic resistance is widely common in cyathostomin parasites, and recent studies have documented increasing incidence of resistance to the macrocyclic lactone drug class. Several European countries have implemented prescription-only restrictions of anthelmintic usage by law to reduce anthelmintic treatment intensity and decrease the selection pressure for drug resistance. However, long term outcomes of this approach have not been thoroughly evaluated. The aim was to determine ivermectin efficacy in strongylids by means...
Szacawa E, Kozieł N, Brzezińska S, Augustynowicz-Kopeć E, Weiner M, Szulowski K, Krajewska-Wędzina M. is one of the most dangerous pathogens of both animals and humans. Bovine tuberculosis (BTB) is a disease caused by mycobacteria belonging to the complex (MTBC), which spreads mainly among domestic cattle but also to mammals other than cattle. The transmission of MTBC between different species requires research and epidemiological investigations to control its spread. When multiple species are a reservoir of infection, it poses a significant public health and veterinary concern. In this study, the diagnosis of alpaca, cattle, horses, dogs, a sheep and a cat from one farm suspected of bovine ...
Espinosa-López EM, Ortiz-Guisado B, Diez de Castro E, Durham A, Aguilera-Tejero E, Gómez-Baena G.Equine Metabolic Syndrome (EMS) is a multifactorial endocrine disorder characterized by obesity, insulin dysregulation (ID), and an increase in the risk of laminitis, a painful condition that can lead to euthanasia in severe cases. Diagnosing EMS is challenging and often relies on clinical history including obesity, difficulty in losing weight, and recurring episodes of laminitis. The gold standard for laboratory support of an EMS diagnosis is the identification of ID, with basal insulin being the simplest and most accessible method, especially in a field setting. However, various factors such...
Doherty O, Conway R, McGreevy P.Pressures applied to horses via nosebands are of growing concern. The current study applied noseband pressure to the head of a dead horse. Pressure sensors were placed on the left nasal bone to record pressures as the noseband was progressively tightened. Tightness increased as predicated by holes in the strap of the noseband (as supplied) through eight steps from two fingers' space, assessed using the standard International Society for Equitation Science Taper Gauge through to zero space. Sensors were also placed at the midline frontal plane and intra-orally at the level of the second premola...
Maniego J, Harding C, Habershon-Butcher J, Hincks P, Stewart G, Proudman C, Ryder E.Due to the ease of collection, transport and storage, the use of dried blood spots (DBS) offers an attractive alternative matrix for detection of the abuse of gene therapy, otherwise known as gene doping. This study evaluated the recovery, extraction efficiency and resulting detection capability of DNA from DBS by evaluating different target types, DNA extraction kits, the number of punches and blood tube preservatives. The long-term storage stability of low-copy-number transgene targets in DBS was not assessed in this study but would be noteworthy to investigate further. DNA was quantified us...
Bell J, Radial SL, Cuming RS, Trope G, Hughes KJ.Diarrhea in foals can be associated with disruption of the intestinal microbiota (dysbiosis). Effective management of intestinal dysbiosis in foals has not been demonstrated. Objective: Fecal microbiota transplantation (FMT) in foals with diarrhea influences the intestinal microbiota and improves clinical and clinicopathological outcomes. Methods: Twenty-five foals <6 months of age with diarrhea and systemic inflammatory response syndrome at 3 veterinary hospitals. Methods: A prospective randomized placebo-controlled cohort study. Foals in the FMT group (n = 19) or control group (n =...
Gabryś J, Pietras N, Kowal-Mierzwa W, Karnas E, Andronowska A, Nowak A, Kochan J, Bugno-Poniewierska M.The efficacy of in vitro embryo production (IVEP) in equines is relatively limited compared to other species due to the lack of a reliable superovulation technique, limited availability of cumulus oocyte complexes (COCs), low in vitro oocyte maturation (IVM) and fertilization rates. Extracellular vesicles (EVs), which are nanoparticles involved in intercellular signaling in the ovarian environment, have shown potential as supplements to improve oocyte development during IVM. This study tested the hypothesis that EVs from small (< 20 mm) ovarian follicles could enhance fertilization rates in m...
Scheike AS, Plomp S, Fugazzola MC, Meurot C, Berenbaum F, van Weeren PR, Tryfonidou MA, von Hegedus JH.This study investigates the anti-inflammatory properties of liraglutide, a glucagon-like peptide 1 receptor agonists, in equine in vitro models and in an in vivo acute synovitis model in Shetland ponies. The anti-inflammatory effect of liraglutide was assessed by measuring concentrations of inflammatory biomarker C-C Motif Chemokine Ligand 2 (CCL2) in culture media of equine whole blood, peripheral blood mononuclear cells (PBMCs), chondrocytes, and synoviocytes, with or without lipopolysaccharide (LPS) or interleukin-1β. In the in vivo experiment, acute synovitis was bilaterally induced with ...
Li D, Wang X, Yao J, Chen S.Adipose tissue-derived extracellular vesicles (ATEVs) have garnered attention for their roles in intercellular communication and regulation. This study presents the first investigation of equine adipose tissue, with ATEV being extracted concurrently during the isolation of adipose-derived mesenchymal stem cells (ADSCs). Through CCK-8 cell proliferation assays and scratch migration assays, a significant promotional effect of ATEV on ADSCs was observed, which not only accelerated the proliferation rate of the stem cells but also enhanced their migratory capacity. The application of Trypan Blue e...
Hébert L, Froger D, Madeline A, Lecouturier F, Lemans C, Zientara S.Dourine is a sexually transmitted parasitic disease affecting equids. Its causative agent is referred to as and the prescribed serodiagnosis method is the complement fixation test (CFT). In the context of our European Reference Laboratory mandate for equine diseases (excluding African horse sickness), we organised dourine CFT inter-laboratory proficiency tests (ILPTs) in 2015, 2018 and 2022 to evaluate the performance of the European Union network of National Reference Laboratories (NRLs) for dourine. ILPT panels were composed of horse sera with or without antibodies against spp. originating...
Sedgwick RL, ElBohy O, Daly JM.In this review, we explore how pseudotyped viruses (PVs) are being applied to the study of viruses affecting both humans and horses. For the purposes of this review, we define PVs as non-replicative viruses with the core of one virus and the surface protein(s) of another and encapsulating a reporter gene such as luciferase. These 'reporter' PVs enable receptor-mediated entry into host cells to be quantified, and thus can be applied to study the initial stages of viral replication. They can also be used to test antiviral activity of compounds and measure envelope protein-specific antibodies in ...
Ivanova Z, Petrova V, Grigorova N, Vachkova E.Obtaining accurate and reliable gene expression results in real-time RT-PCR (qRT-PCR) data analysis requires appropriate normalization by carefully selected reference genes, either a single or a combination of multiple housekeeping genes (HKGs). The optimal reference gene/s for normalization should demonstrate stable expression across varying conditions to diminish potential influences on the results. Despite the extensive database available, research data are lacking regarding the most appropriate HKGs for qRT-PCR data analysis in rabbit and horse adipose-derived stem cells (ASCs). Therefore,...
Maniego J, Harding C, Habershon-Butcher J, Hincks P, Ryder E.Gene doping, which includes the non-therapeutic use of genes or genetic elements that have the capacity to enhance athletic performance, is prohibited in horseracing and equestrian sports. To provide a comprehensive assessment of matrix and detection techniques, a custom adeno-associated virus serotype 8 vector was designed to include PCR binding sites for multiple target genes and assay types. The vector was injected via an intramuscular route into two Thoroughbred horses and matrices collected at defined timepoints. DNA was analysed using 3 detection methods: qPCR, digital PCR, and NGS. Over...
Switonski M, Szczerbal I, Nowacka-Woszuk J.Identification of chromosomal abnormalities is an important issue in animal breeding and veterinary medicine. Routine cytogenetic diagnosis of domestic animals began in the 1960s with the aim of identifying carriers of centric fusion between chromosome 1 and 29 in cattle. In the 1970s, chromosome banding techniques were introduced, and in the 1980s, the first cytogenomic techniques, based on the development of locus- and chromosome-specific probes, were used. Since the beginning of the twenty-first century, molecular techniques (such as polymorphism of microsatellite markers, droplet digital P...
Villalba R, Tena-Tomás C, Ruano MJ, Valero-Lorenzo M, López-Herranz A, Cano-Gómez C, Agüero M.The African horse sickness virus (AHSV) belongs to the Genus Orbivirus, family Sedoreoviridae, and nine serotypes of the virus have been described to date. The AHSV genome is composed of ten linear segments of double-stranded (ds) RNA, numbered in decreasing size order (Seg-1 to Seg-10). Genome segment 2 (Seg-2) encodes outer-capsid protein VP2, the most variable AHSV protein and the primary target for neutralizing antibodies. Consequently, Seg-2 determines the identity of the virus serotype. An African horse sickness (AHS) outbreak in an AHS-free status country requires identifying the seroty...
Cochet M, Piumi F, Gorna K, Berry N, Gonzalez G, Danckaert A, Aulner N, Blanchet O, Zientara S, Donadeu FX, Munier-Lehmann H, Richardson J....Outbreaks of West Nile virus (WNV) occur periodically, affecting both human and equine populations. There are no vaccines for humans, and those commercialised for horses do not have sufficient coverage. Specific antiviral treatments do not exist. Many drug discovery studies have been conducted, but since rodent or primate cell lines are normally used, results cannot always be transposed to horses. There is thus a need to develop relevant equine cellular models. Here, we used induced pluripotent stem cells to develop a new in vitro model of WNV-infected equine brain cells suitable for microplat...
