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.
Ajeebsanu MM, Koshy SA, Karakka Kal AK, Subhahar MB, Karatt TK, Philip M.Suzetrigine, a recently approved Na1.8 sodium channel blocker, shows strong potential in the treatment of neurological, psychiatric, and pain-related conditions. Its peripheral selectivity enables effective pain management while avoiding central nervous system complications and addiction risks linked to opioid use. Following FDA approval in January 2025, concerns have emerged regarding its possible misuse for performance enhancement in sports, highlighting the need for reliable detection tools in doping control. Methods: An analytical procedure was designed and validated to detect suzetrigine ...
Payette F, Kulp JC, Graves A, Janes J, Morrow J, Orr K, Reed S, Ruby R, Stefanovski D, Johnson AL.To measure and compare CSF cytokine concentrations among horses with equine neuroaxonal dystrophy (eNAD)/equine degenerative myeloencephalopathy (EDM), horses with cervical vertebral stenotic myelopathy (CVSM), and control horses and to evaluate for associations with clinical parameters. Unassigned: Banked equine CSF samples from horses with neurologic disease that underwent a complete neurologic examination and postmortem diagnosis confirmation of CVSM or eNAD/EDM or from control horses were included. Cytokines were measured with an equine-specific cytokine/chemokine magnetic bead multiplex p...
Massardi E, Gaudenzi G, Carra S, Oldani M, Rybinska I, Persani L, Vitale G.Cushing's disease (CD) is a rare neuroendocrine disorder caused by ACTH-secreting pituitary adenomas, presenting significant diagnostic and therapeutic challenges. Given the evolutionary conservation of the hypothalamic-pituitary-adrenal axis, this review explores the translational value of spontaneous CD forms in dogs, horses, cats, small mammals, and rats, as well as of experimental models in mice, rats, and zebrafish. Dogs are the most studied, showing strong molecular and clinical similarities with human CD, making them valuable for preclinical drug and diagnostic research. While equine an...
Fehin B, Scott CJ, Arango-Sabogal JC, de Mestre AM, Mouncey R.To date, relationships between pre-covering endometrial swab cytology and bacteriology and fertility outcomes in Thoroughbred broodmares in the United Kingdom are unknown and could aid clinical decision making. Objective: To investigate associations between cytology and bacteriology findings from the last endometrial swab taken in the breeding season (15 February to 15 July) and live-foal rates (predicted mean probability of producing a live foal) in UK Thoroughbred broodmares. Methods: Retrospective cohort study. Methods: Endometrial cytology and bacteriology findings were extracted from labo...
Silva GLS, Curcio UA, Boufleur JA, Borges DGL, Nakatani MTM, Freitas MG, Borges FA, Faria FJC, Silva RHP, Franco GL.Anthelmintic resistance has led to the use of organic extracts as alternative methods of parasite control. Objective: The study aimed to assess the effects of Acacia mearnsii extract (tannin) on the control of cyathostomins in horses, both in vitro and in vivo. Methods: Thirty Pantaneiro horses naturally infected with cyathostomins were sourced from two distinct farms, designated as Farms A and B. At the start of the study, all third-stage larvae were cyathostomins, and an egg hatchability test (EHT) was performed using fecal samples from horses on both farms. Two randomized 28-day field studi...
Valderrama-Martinez C, Packham A, Smith W, Mendoza-Flores JE, Zheng S, Chigerwe M, Plancarte M, Aleman M.Long-term freezing storage can alter the stability of proteins, thereby compromising accurate determination of indirect fluorescent antibody test (IFAT) titers that support the diagnosis of equine protozoal myeloencephalitis. Objective: Assess the effect of long-term storage at -80°C on IFAT against S. neurona and N. hughesi in equine serum and cerebrospinal fluid (CSF). Methods: Paired serum and CSF (n = 46), and serum only (n = 25) samples. Methods: Prospective study of samples stored 6-12, 13-18, and 19-24 months. Comparing antibody titers across time points, McNemar and Wilcoxon...
Åžahinkesen Ä°, Bilge-DaÄŸalp S.Equine herpesvirus 1 (EHV-1) and Equine herpesvirus 4 (EHV-4) are major viral pathogens of horses that contribute to respiratory disease, abortion, and neurological disorders, leading to economic losses in the equine industry. Accurate serological diagnosis is critical for disease surveillance and control. This study aimed to develop and validate an in-house enzyme-linked immunosorbent assay (ELISA) for detecting antibodies against EHV-1 and EHV-4 in horse sera. Serum samples previously confirmed by virus neutralization test (VNT) and a commercial ELISA were used to optimize the assay. Cut-off...
Shimizu Y, Sugai-Bannai M, Tanabe H, Saito K, Ito H, Tokushige H, Kamiya K, Hirano-Kodaira M, Yamada M, Leung GN.Somapacitan is the second generation of recombinant human growth hormone (rhGH) medication that retains the pharmacological effects of rhGH but exhibits a longer duration of action due to its reversible albumin-binding in the body. In general, the use of all recombinant growth hormone (rGH) analogues is banned by the human and animal sports regulatory authorities due to their anabolic and lipolytic effects. However, little is known about the elimination kinetics and biological effects of the newly introduced long-acting rhGH, somapacitan, in horses. This paper describes the administration stud...
Kovarikova S, Blahova J, Steffenova V, Vaskova N, Jahn P.Currently, there is relatively little published information on the urine protein-to-creatinine (UPC) ratio in horses, a parameter commonly used in dogs and cats. The aim of this study was to determine the reference range of UPC in urine samples of apparently healthy horses and to assess the influence of age and sex. Urinalysis and the determination of urinary protein and creatinine concentrations and ratios were performed. A total of 118 urine samples from apparently healthy horses were included in the study: 10 from foals younger than 6 months and 108 from horses older than 1 year. The median...
Liu F, Zhang J, Kong L, Wu R, Jiang Q, Lu Y, Li X.As an interspecies hybrid inheriting genetic material from horse and donkey lineages, mules provide a unique model for studying allele-specific regulatory dynamics. Here, we isolated adult fibroblasts (AFs) and placental fibroblasts (PFs) from mule tissues and reprogrammed them into induced pluripotent stem cells (iPSCs). Intriguingly, placental fibroblast-derived iPSCs (mpiPSCs) exhibited reduced reprogramming efficiency compared to adult fibroblast-derived iPSCs (maiPSCs). Through allele-specific expression (ASE) analysis, we systematically dissected transcriptional biases in parental cell t...
Castelain D, Bokma J, Pas ML, Verbanck S, Paepe D, Pardon B, Boyen F.Accurate diagnosis of bloodstream infections is crucial for survival and antimicrobial de-escalation in veterinary medicine. Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry enables faster identification of bacteria in BSIs. This study aimed to compare diagnostic accuracy of two Sepsityper methods (Rapid Sepsityper and Sepsityper Extraction) with conventional culture method for bacterial identification in clinical samples. Mean time-to-positivity and frequency of bacteria in BSIs were also determined. This diagnostic test study used bloodstream infection samples fro...
Van Buren SL, Petersen JL, Brown CT, Finno CJ.With advancing genomic technologies, single-nucleotide polymorphism (SNP) arrays and whole genome sequencing (WGS) have become essential tools in equine genetic research. In this study, we assessed the concordance in SNP calls and trait-mapping efficacy by comparing data of 21 horses both genotyped on the Equine 670 K SNP array and sequenced at either ~12× or ~30× depth. Our analysis revealed that higher sequencing depths were significantly associated with fewer discordant calls between platforms. Additionally, we investigated the most frequent no-call and discordant positions and identifi...
Carmona JU, López C, Jurado-Grisales C.Platelet-rich plasma (PRP) is widely utilized as a routine treatment for chronic musculoskeletal conditions in horses, such as osteoarthritis, tendinopathies, and desmopathies. This effectiveness stems from the high concentration of growth factors and anti-inflammatory cytokines that are released upon activation of this orthobiologic agent, whether activated endogenously or exogenously. Despite its growing popularity, there is a notable absence of instructional videos that demonstrate the techniques for obtaining PRP in horses. This study examines a double centrifugation tube method for obtain...
Crosby CE, O'Connor A, Munsterman AS.Colic is a significant source of morbidity and mortality in horses, and no single biomarker can distinguish surgical from medical colic or predict mortality. Alactic base excess (ABE) has shown an association with mortality and renal insufficiency in humans but has not been evaluated in veterinary species. The purpose of this study was to determine the value of ABE as a diagnostic tool for horses treated for gastrointestinal disease. Unassigned: This retrospective case series evaluated horses admitted for acute gastrointestinal disease over a 5-year period (2019-2024). Signalment, physical exa...
Aboelmaaty AM, El-Seadawy IE, Kotp MS, Mohamed AA, El-Debaky HA, El-Badry DA, Anwer AM, Tsvetkov T, Daneva T, Elgabry MA.Alpha-tocopherol is one of the non-enzymatic lipophilic antioxidants. Butylated hydroxytoluene (BHT) is a synthetic analog with similar modes of action in protecting the cryopreserved sperms. Objective: This study hypothesized that a certain concentration of any antioxidant is suitable for improving the post-thaw semen quality of stallions. Methods: To determine the optimum BHT concentration, a synthetic antioxidant similar to vitamin E in potency and scavenging oxidative stress power in concentrations of 0.0, 0.25, 0.50, 1.0, 2.0, and 4.0 mM/ml were added to semen extender. The post-thaw sper...
Graham AE, Colgate VA, Floyd EF.Geographical specific data is required to guide empirical antimicrobial selection in equine neonates. Objective: Evaluate antibiograms and survival in foals from a United Kingdom (UK)-based hospital to guide antimicrobial selection. Methods: Blood and synovial fluid samples from 208 foals ≤ 30 days old admitted to Rossdales Equine Hospital from 2018 to 2023. Methods: Retrospective cohort study. Antimicrobial susceptibility was assessed by disc diffusion method. Bacterial culture and susceptibility and foal survival were recorded. The effects of the presence of positive culture or multi...
Faircloth J, Fielding L, Epstein K, Crabtree N, Mochal C, Magdesian G, Major D.To describe the clinical presentation of horses with synchronous diaphragmatic flutter (SDF) and identify clinical and laboratory variables associated with survival. Methods: Retrospective study from January 2004 to September 2022. Methods: Five large animal referral hospitals in the United States. Methods: Thirty-seven horses of various breeds and ages presenting with clinical signs of SDF, excluding animals participating in an endurance competition. All horses were >1Â year of age. Methods: None. Results: Variables recorded and analyzed included signalment, clinical and laboratory data, trea...
Wasiński B, Złotnicka J, Kubajka M, Olejarczyk M, Szulowski K.Contagious equine metritis (CEM) is a cosmopolitan infectious, venereal disease of equids caused by the bacterium . Its frequently asymptomatic course leads to its prevalence sometimes being underestimated, and knowledge of the spread of infections with its causative agent is insufficient. The aim of this study is to summarise and present data on the incidence of infections in horses in Poland. Unassigned: In the years 2018-2023, routine laboratory tests of horses for CEM were carried out. Between 52 and 99 horses were examined annually. Swabs from the external parts of the urogenital system ...
Sohn Y, An SJ, Forbes E, Yoon J, Kim BS, Kim JG, Ryu SH.To model lactate recovery kinetics in relation to sex, age, running speed, and racing experience, and to estimate blood lactate clearance time under passive recovery conditions that reflect current management practices in Jeju horses, a less well-characterized pony-sized indigenous breed. Unassigned: From October through December 2022, BLC was measured at baseline, 10 minutes after 800-m barrier trials (time [T]-1), and 50 minutes (T2) after 800-m barrier trials in 40 race-trained Jeju horses (2 to 4 years old). Lactate clearance was modeled using nonlinear regression with an exponential decay...
Shaffer SK, Medjaouri O, Swenson B, Eliason T, Nicolella DP.The ability to quantify equine kinematics is essential for clinical evaluation, research, and performance feedback. However, current methods are challenging to implement. This study presents a motion capture methodology for horses, where three-dimensional, full-body kinematics are calculated without instrumentation on the animal, offering a more scalable and labor-efficient approach when compared with traditional techniques. Kinematic trajectories are calculated from multi-camera video data. First, a neural network identifies skeletal landmarks (markers) in each camera view and the 3D location...
Zimmerman KL, Diehl KA.To determine if serum α2-macroglobulin (A2M) concentration varies based on donor species-canine or equine-and signalment. Methods: Serum A2M concentration ([A2M]) was measured in healthy dogs (n = 30) and horses (n = 31) using species-specific ELISAs. Results: Canine and equine [A2M] median (IQR; range) were 98.70 ng/mL (92.79 ng/mL; 34.33-696.18 ng/mL) and 557 000 ng/mL (437 900 ng/mL; 62 600-3 042 900 ng/mL), respectively. Equine [A2M] was significantly higher than canine [A2M]. Depending on the statistical analysis performed, sex had either no statistical (pâ€...
Pinn-Woodcock TL, Llanos-Soto SG, Ivanek R, Goodrich E, Frye E, Wells A, Mullen K, Arbittier E, Hughes L, Berryhill E, Brosnahan P, Buslinger L....To determine whether seasonal variation in equine plasma ACTH concentrations is affected by latitude between 29.7°N and 45.0°N in the US. Unassigned: A cohort study was performed between June 2019 and May 2020 in which plasma ACTH was measured monthly by chemiluminescence immunoassay. The study involved 201 apparently healthy horses from 9 locations within the US classified into northern, central, and southern latitude cohorts. Data analysis was performed with 3 major objectives: (1) identify outlier ACTH concentrations and horses; (2) estimate ACTH upper reference limits to define expected ...
Rey-Conejo R, Toribio RE, Möller S, Müller E, Fores-Jackson P.Insulin dysregulation is a key component of equine metabolic syndrome and is commonly assessed using basal insulin concentrations. Different analyzers may yield variable insulin results, limiting comparability. Objective: To assess the analytical performance of the ADVIA Centaur XPT chemiluminescent immunoassay (CLIA) for equine insulin measurement and to establish adapted decision thresholds based on comparison with a previously used CLIA method. Methods: Precision, linearity, and dilution recovery were assessed for the ADVIA Centaur XPT. A total of 89 equine serum samples submitted for basal...
Belhout C, Wang F, Rossano A, Collaud A, Fernandez JE, Marchionatti E, Keller JE, Overesch G, Kaessmeyer S, Schwendener S, Perreten V.Nine Gram-positive, non-motile, facultatively anaerobic cocci, designated EM39Eáµ€, JEK85, 18KM676, 21M1142, 18KM445, 18KM444, 18KM245, 18KM583 and 21KM1573, were isolated from diverse animal material, including horse and pig skin, bovine mastitis milk and feline urine from a urinary tract infection. Phylogenomic analysis based on amino-acid alignment obtained from translated whole-genome sequences; digital DNA-DNA hybridization (dDDH); 16S rRNA, , , and gene comparison; and MALDI-TOF MS spectral profiles placed the strains within the clade. They were closely related to CCM 7100áµ€, CCM 7...
Gupta S, Sethi K, Vohra S, Gupta S, Rani R, Kumar S, Kumar R.We report on the in vitro and in vivo success of Imidocarb dipropionate (IDP), an aromatic diamidine compound, as an effective drug against Surra-causing organism, Trypanosoma evansi. IDP is a clinically acclaimed drug for the treatment of babesiosis and anaplasmosis. We have also investigated cytotoxic effect of IDP on equine Peripheral mononuclear cells (PBMCs) and Vero cell line. The data generated indicate that the IC of IDP (2.73 µM) was significantly lower than its cytotoxic concentration (32.66 and 80.70 µM against PBMCs and Vero cell line, respectively). We have recorded signific...
Celik F, Karabulut B, Uslug M, Cankaya E, Eroksuz Y, Simsek S. is a parasitic cestode primarily maintained within an equine-canine life cycle, with horses, donkeys, mules, and other ungulates serving as intermediate hosts. Although has historically been considered non-zoonotic, recent molecular studies suggest that this assumption may need to be reevaluated. This study aimed to investigate the presence and molecular characterization of in equids from Türkiye. A retrospective analysis of 52 equine necropsies performed between 2020 and 2025 identified hydatid cysts in one Arabian horse and two donkeys. Gross and histopathological examination confirmed t...
Migliorisi A, Hassel DM, Moore AR, Blair BW, Wilkins PA.To characterize coagulopathies before and after antivenom administration in rattlesnake-envenomated equids. Unassigned: Client-owned envenomated equids treated with antivenom at Colorado State University (2023 to 2024). Rattlesnake bite severity score (RBSS), platelet count, prothrombin time, activated thromboplastin time, D-dimers, antithrombin, and viscoelastic testing variables were assessed before and after antivenom administration. Paired comparisons of hemostatic variables between admission and approximately 24 hours, and correlation analysis with RBSS, were performed. Length of hospital...
Frazer KA, Tao H, Osoegawa K, de Jong PJ, Chen X, Doherty MF, Cox DR.Cross-species DNA sequence comparison is a fundamental method for identifying biologically important elements, because functional sequences are evolutionarily conserved, wheres nonfunctional sequences drift. A recent genome-wide comparison of human and mouse DNA discovered over 200,000 conserved noncoding sequences with unknown function. Multispecies DNA comparison has been proposed as a method to prioritize these conserved noncoding sequences for functional analysis based on the hypothesis that elements present in many species are more likely to be functional than elements present in limited ...
Gravance CG, Garner DL, Baumber J, Ball BA.The fluorescent carbocyanine dye, JC-1, labels mitochondria with high membrane potential orange and mitochondria with low membrane potential green. Evaluation of mitochondrial membrane potential with JC-1 has been used in a variety of cell types, including bull spermatozoa; however, JC-1 staining has not yet been reported for equine spermatozoa. The aim of this study was to apply JC-1 staining and assessment by flow cytometry or a fluorescence microplate reader for evaluation of mitochondrial function of equine spermatozoa. Six ejaculates from four stallions were collected and centrifuged thro...
Arnhold S, Elashry MI, Klymiuk MC, Geburek F.Adipose tissue-derived mesenchymal stem cells (ASCs) offer a promising cell source for therapeutic applications in musculoskeletal disorders. The appropriate selection of ASCs from various fat depots for cell-based therapy is challenging. The present study aims to compare stemness and multipotency of ASCs derived from retroperitoneal (RP), subcutaneous (SC), and lipoma (LP) fat to assess their usefulness for clinical application. Equine ASCs from the three fat tissue sources were isolated and characterized. The cell viability, proliferation, and self-renewal were evaluated using MTT, sulforhod...
Rizk MA, El-Sayed SA, Terkawi MA, Youssef MA, El Said el Sel S, Elsayed G, El-Khodery S, El-Ashker M, Elsify A, Omar M, Salama A, Yokoyama N....A rapid and accurate assay for evaluating antibabesial drugs on a large scale is required for the discovery of novel chemotherapeutic agents against Babesia parasites. In the current study, we evaluated the usefulness of a fluorescence-based assay for determining the efficacies of antibabesial compounds against bovine and equine hemoparasites in in vitro cultures. Three different hematocrits (HCTs; 2.5%, 5%, and 10%) were used without daily replacement of the medium. The results of a high-throughput screening assay revealed that the best HCT was 2.5% for bovine Babesia parasites and 5% for equ...
Paria N, Raudsepp T, Pearks Wilkerson AJ, O'Brien PC, Ferguson-Smith MA, Love CC, Arnold C, Rakestraw P, Murphy WJ, Chowdhary BP.Studies of the Y chromosome in primates, rodents and carnivores provide compelling evidence that the male specific region of Y (MSY) contains functional genes, many of which have specialized roles in spermatogenesis and male-fertility. Little similarity, however, has been found between the gene content and sequence of MSY in different species. This hinders the discovery of species-specific male fertility genes and limits our understanding about MSY evolution in mammals. Here, a detailed MSY gene catalogue was developed for the horse--an odd-toed ungulate. Using direct cDNA selection from horse...
Roth SP, Glauche SM, Plenge A, Erbe I, Heller S, Burk J.Decellularization of tendon tissue plays a pivotal role in current tissue engineering approaches for in vitro research as well as for translation of graft-based tendon restoration into clinics. Automation of essential decellularization steps like freeze-thawing is crucial for the development of more standardized decellularization protocols and commercial graft production under good manufacturing practice (GMP) conditions in the future. In this study, a liquid nitrogen-based controlled rate freezer was utilized for automation of repeated freeze-thawing for decellularization of equine superficia...
Smit H, Gaastra W, Kamerling JP, Vliegenthart JF, de Graaf FK.The erythrocyte receptor for Escherichia coli K99 fimbrial adhesin was isolated from equine erythrocytes and characterized as Neu5Gc-alpha(2----3)-Galp-beta(1----4)-GLcp-beta(1----1)-Ceramide. This glycolipid acted as the receptor for K99 by four different experimental approaches: inhibition of equine erythrocyte hemagglutination by preincubation of K99-positive bacteria or purified K99 fimbriae with the isolated glycolipid; inhibition of attachment of K99-positive bacteria to porcine intestinal epithelial cells in the presence of the isolated glycolipid; induction of binding of K99-positive b...
Beck C, Lowenski S, Durand B, Bahuon C, Zientara S, Lecollinet S.West Nile Fever is a zoonotic disease caused by a mosquito-borne flavivirus, WNV. By its clinical sensitivity to the disease, the horse is a useful sentinel of infection. Because of the virus' low-level, short-term viraemia in horses, the primary tools used to diagnose WNV are serological tests. Inter-laboratory proficiency tests (ILPTs) were held in 2010 and 2013 to evaluate WNV serological diagnostic tools suited for the European network of National Reference Laboratories (NRLs) for equine diseases. These ILPTs were designed to evaluate the laboratories' and methods' performances in detectin...
Verheij HM, Volwerk JJ, Jansen EH, Puyk WC, Dijkstra BW, Drenth J, de Haas GH.It is known that His-48 is part of the active center in pancreatic phospholipase. To further elucidate the role of histidine-48 in the active center of pancreatic phospholipase A2, we have modified the enzyme with a number of bromo ketones and methyl benzenesulfonates. Rapid methylation occurred with methyl p-nitrobenzenesulfonate. Methylated phospholipase shows total loss of enzymatic activity whereas binding of substrate and the cofactor Ca2+ remains intact. Amino acid analysis of methylated equine phospholipase showed the loss of the single molecule of histidine and the formation of one mol...
Poissant J, Gavriliuc S, Bellaw J, Redman EM, Avramenko RW, Robinson D, Workentine ML, Shury TK, Jenkins EJ, McLoughlin PD, Nielsen MK, Gilleard JS.Horses are ubiquitously infected by a diversity of gastro-intestinal parasitic helminths. Of particular importance are nematodes of the family Strongylidae, which can significantly impact horse health and performance. However, knowledge about equine strongyles remains limited due to our inability to identify most species non-invasively using traditional morphological techniques. We developed a new internal transcribed spacer 2 (ITS2) DNA metabarcoding 'nemabiome' assay to characterise mixed strongyle infections in horses and assessed its performance by applying it to pools of infective larvae ...
Scepek S, Coorssen JR, Lindau M.Using the patch-clamp technique, we studied the role of protein phosphorylation and dephosphorylation on the exocytotic fusion of secretory granules with the plasma membrane in horse eosinophils. Phorbol 12-myristate 13-acetate (PMA) had no effect on the amplitude and dynamics of degranulation, indicating that the formation of fusion pores is insensitive to activation of protein kinase C (PKC). Fusion pore expansion, however, was accelerated approximately 2-fold by PMA, and this effect was abolished by staurosporine. Elevating intracellular Ca2+ to 1.5 microM also resulted in a 2-fold accelera...
Dobbe JC, van der Meer Y, Spaan WJ, Snijder EJ.The recent development of arterivirus full-length cDNA clones makes possible the construction of chimeric arteriviruses for fundamental and applied studies. Using an equine arteritis virus (EAV) infectious cDNA clone, we have engineered chimeras in which the ectodomains of the two major envelope proteins, the glycoprotein GP(5) and the membrane protein M, were replaced by sequences from envelope proteins of related and unrelated RNA viruses. Using immunofluorescence microscopy, we monitored the transport of the hybrid GP(5) and M proteins to the Golgi complex, which depends on their heterodime...
Nikiforov TT, Rendle RB, Goelet P, Rogers YH, Kotewicz ML, Anderson S, Trainor GL, Knapp MR.A new method for typing single nucleotide polymorphisms in DNA is described. In this method, specific fragments of genomic DNA containing the polymorphic site(s) are first amplified by the polymerase chain reaction (PCR) using one regular and one phosphorothioate-modified primer. The double-stranded PCR product is rendered single-stranded by treatment with the enzyme T7 gene 6 exonuclease, and captured onto individual wells of a 96 well polystyrene plate by hybridization to an immobilized oligonucleotide primer. This primer is designed to hybridize to the single-stranded target DNA immediately...
Di Gennaro A, Lorusso A, Casaccia C, Conte A, Monaco F, Savini G.A serum neutralization assay (SN) was compared with the official plaque reduction neutralization test for the quantitation of West Nile virus antibodies. A total of 1,348 samples from equid sera and 38 from human sera were tested by these two methods. Statistically significant differences were not observed, thus supporting the use of SN for routine purposes.
Nally JE, Artiushin S, Timoney JF.Leptospira interrogans is a mammalian pathogen which must adapt to a range of new environmental conditions including temperature change when it infects new hosts. In vitro studies of organisms cultured at 30 degrees C and shifted to 37 degrees C for 5 to 7 days have confirmed that synthesis of several proteins involved in equine infection is regulated in response to temperature change (J. E. Nally, J. F. Timoney, and B. Stevenson, Infect. Immun. 69:400-404, 2001). In order to specifically identify antigenic proteins upregulated at 37 degrees C, groups of three ponies were immunized with organi...
Alhassan A, Thekisoe OM, Yokoyama N, Inoue N, Motloang MY, Mbati PA, Yin H, Katayama Y, Anzai T, Sugimoto C, Igarashi I.Loop-mediated isothermal amplification (LAMP) is a novel nucleic acid method whereby DNA is amplified with high specificity, efficiency, and rapidity under isothermal conditions using a set of four specifically designed primers and a DNA polymerase with strand displacement activity. In this study, we used LAMP primer sets designed from EMA-1 and Bc 48 genes for detection of Theileria equi and Babesia caballi infections, respectively. These primer sets specifically amplified DNA of the respective parasites. Both primer sets amplified T. equi and B. caballi up to 10(-6) dilution of 10-fold seria...
Takahashi T, Yawata M, Raudsepp T, Lear TL, Chowdhary BP, Antczak DF, Kasahara M.In rodents, the Ly49 family encodes natural killer (NK) receptors interacting with classical MHC class I molecules, whereas the corresponding receptors in primates are members of the killer cell immunoglobulin-like receptor (KIR) family. Recent evidence indicates that the cattle, domestic cat, dog, and pig have a single LY49 and multiple KIR genes, suggesting that predominant NK receptors in most non-rodent mammals might be KIR. Here, we show that the horse has at least six LY49 genes, five with an immunoreceptor tyrosine-based inhibition motif (ITIM) and one with arginine in the transmembrane...
Slusarewicz P, Pagano S, Mills C, Popa G, Chow KM, Mendenhall M, Rodgers DW, Nielsen MK.Intestinal parasites are a concern in veterinary medicine worldwide and for human health in the developing world. Infections are identified by microscopic visualisation of parasite eggs in faeces, which is time-consuming, requires technical expertise and is impractical for use on-site. For these reasons, recommendations for parasite surveillance are not widely adopted and parasite control is based on administration of rote prophylactic treatments with anthelmintic drugs. This approach is known to promote anthelmintic resistance, so there is a pronounced need for a convenient egg counting assay...
Zhen H, Han T, Fennell DE, Mainelis G.We report here that stress experienced by bacteria due to aerosolization and air sampling can result in severe membrane impairment, leading to the release of DNA as free molecules. Escherichia coli and Bacillus atrophaeus bacteria were aerosolized and then either collected directly into liquid or collected using other collection media and then transferred into liquid. The amount of DNA released was quantified as the cell membrane damage index (ID), i.e., the number of 16S rRNA gene copies in the supernatant liquid relative to the total number in the bioaerosol sample. During aerosolization by ...
Kim CM, Blanco LB, Alhassan A, Iseki H, Yokoyama N, Xuan X, Igarashi I.We developed a TaqMan real-time polymerase chain reaction (PCR) assay for the quantitative detection of Theileria equi from the in vitro-cultured parasite and field blood samples collected from horses living in Ghana and Brazil. The detection limit for the assay was determined to be 1.5 parasites/microl per sample, and the quantitative capacity was demonstrated using the in vitro-cultured parasite. For field applications, the real-time PCR assay was compared to a previously established nested PCR assay used as the gold standard for the real-time PCR assay. Of 65 field blood samples, 46 samples...
Richardson DW, Dodge GR.To determine the effects of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) on expression and regulation of several matrix-related genes by equine articular chondrocytes. Methods: Articular cartilage harvested from grossly normal joints of 8 foals, 6 yearling horses, and 8 adult horses. Methods: Chondrocytes maintained in suspension cultures were treated with various doses of human recombinant IL-1beta or TNF-alpha. Northern blots of total RNA from untreated and treated chondrocytes were probed with equine complementary DNA (cDNA) probes for cartilage matrix-related ge...
McPartlin LA, Suarez SS, Czaya CA, Hinrichs K, Bedford-Guaus SJ.Capacitation is a complex and not well-understood process that encompasses all the molecular changes sperm must undergo to successfully fertilize an oocyte. In vitro fertilization has remained elusive in the horse, as evidenced by low in vitro fertilization (IVF) rates (0%-33%); moreover, only two foals have ever been produced using IVF. Incubation of stallion sperm in modified Whittens supplemented with bovine serum albumin and sodium bicarbonate yielded significant rates of time-dependent protein tyrosine phosphorylation and induced acrosomal exocytosis, consistent with capacitation. The obj...
Rudolph J, O'Callaghan DJ, Osterrieder N.The genome of equine herpesvirus type 1 (EHV-1) strain RacL11, a highly virulent isolate obtained from an aborted foal, and that of the modified live vaccine strain KyA, were cloned as bacterial artificial chromosomes (BAC) in Eseherichia coli. Mini F plasmid sequences were inserted into the viral genomes by homologous recombination instead of the gene 71 (EUS4) open reading frame after co-transfection of viral DNA and recombinant plasmid pdelta71-pHA2 into RK13 cells. After isolation of recombinant viruses by three rounds of plaque purification, viral DNA was isolated from RK13 cells infected...
Fornstedt B, Rosengren E, Carlsson A.The 5-S-cysteinyl derivatives of dopamine, dopa (3,4-dihydroxyphenylalanine) and dopac (3,4-dihydroxyphenylacetic acid) were synthesized and used as reference compounds in high performance liquid chromatography analyses of extracts from various brain regions of eight mammalian species. All three metabolites were detected in the brains of all the species studied. The regional distribution of the metabolites was similar to that of dopamine; the metabolite concentrations ranged from less than 0.1 percent to more than 1 percent of the dopamine level, the highest ratios generally being found in sub...
Haber M, Cao Z, Panjwani N, Bedenice D, Li WW, Provost PJ.The physiologic mechanisms involving growth factors, including PDGF-BB, EGF, and TGF-beta 1, as potent mediators of fibroblasts and epithelial cells in corneal wound healing remain unknown. The goal of this study was to determine culture methods for equine epithelial cells and keratocytes and to investigate how exogenous growth factors influence proliferation of both cell types. Methods: Cell cultures were established from healthy corneas harvested from horses immediately following euthanasia and maintained using standard tissue culture protocols. To determine the effects of PDGF-BB, EGF, TGF-...
Nielsen MK, Peterson DS, Monrad J, Thamsborg SM, Olsen SN, Kaplan RM.Strongylus vulgaris is an important strongyle nematode with high pathogenic potential infecting horses world-wide. Several decades of intensive anthelmintic use has virtually eliminated clinical disease caused by S. vulgaris, but has also caused high levels of anthelmintic resistance in equine small strongyle (cyathostomin) nematodes. Recommendations aimed at limiting the development of anthelmintic resistance by reducing treatment intensity raises a simultaneous demand for reliable and accurate diagnostic tools for detecting important parasitic pathogens. Presently, the only means available t...
Kolodziejek J, Dürrwald R, Herzog S, Ehrensperger F, Lussy H, Nowotny N.The aim of this study was to gain more detailed insights into the genetic evolution and variability of Borna disease virus (BDV). Phylogenetic analyses were performed on field viruses originating from naturally infected animals, the BDV vaccine strain 'Dessau', four widely used laboratory strains and the novel BDV subtype No/98. Four regions of the BDV genome were analysed: the complete p40, p10 and p24 genes and the 5'-untranslated region of the X/P transcript. BDV isolates from the same geographical area exhibited a clearly higher degree of identity to each other than to BDV isolates from ot...
Zhao Y, Holden VR, Smith RH, O'Callaghan DJ.The UL3 protein of equine herpesvirus 1 (EHV-1) KyA strain is a homolog of the ICP27 alpha regulatory protein of herpes simplex virus type 1 (HSV-1) and the ORF 4 protein of varicella-zoster virus. To characterize the regulatory function of the UL3 gene product, a UL3 gene expression vector (pSVUL3) and a vector expressing a truncated version of the UL3 gene (pSVUL3P) were generated. These effector plasmids, in combination with an EHV-1 immediate-early (IE) gene expression vector (pSVIE) and chimeric EHV-1 promoter-chloramphenicol acetyltransferase (CAT) reporter constructs, were used in trans...
