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.
Casella S, Giannetto C, Fazio F, Giudice E, Piccione G.The aim of the present study was to assess the effect of different storage conditions on prothrombin time (PT), activated partial thromboplastin time (aPTT), and fibrinogen concentration in clinical samples from healthy horses. A total of 100 healthy horses of varying breeds and gender, ranging in age from 4 to 18 years, with a mean body weight of 480 +/- 70 kg, were used. Blood was collected by jugular venipuncture, and a hemochrome-cytometric examination was conducted on all samples. All blood samples were centrifuged and divided into 4 different aliquots to assess clotting parameters by mea...
Vonaparti A, Lyris E, Panderi I, Koupparis M, Georgakopoulos C.In equine sport, salicylic acid is prohibited with a threshold level of 750 microg mL(-1) in urine; hence, doping control laboratories have to establish quantitative and qualitative methods for its determination. A simple and rapid liquid chromatographic/mass spectrometric method was developed and validated for the quantification and identification of salicylic acid. Urine samples after 900-fold dilution and addition of the internal standard (4-methylsalicylic acid) were directly injected to the liquid chromatography/quadrupole time-of-flight mass spectrometry system. Electrospray ionization i...
Endo A, Futagawa-Endo Y, Dicks LM.Lactobacillus equi, Lactobacillus hayakitensis, Lactobacillus johnsonii, and Weissella confusa/cibaria were the dominant species in 12 South African horses. The Bifidobacterium-group was detected in the feces of only one of the 12 horses. Sequencing of the nested-PCR amplicon identified the Bifidobacterium-group as Parascardovia denticolens. Cell numbers of L. equi, L. hayakitensis, and W. confusa/cibaria were consistent in all samples. P. denticolens, Bifidodobacterium pseudolongum, and a phylogenetic relative of Alloscardovia omnicolens were rarely detected. L. equigenerosi, a dominant speci...
Musilova P, Kubickova S, Horin P, Vodicka R, Rubes J.Cross-species chromosome painting has been applied to most of the species making up the numerically small family Equidae. However, comparative mapping data were still lacking in Asiatic asses kulan (Equus hemionus kulan) and kiang (E. kiang). The set of horse arm-specific probes generated by laser microdissection was hybridized onto kulan (E. hemionus kulan) and kiang (E. kiang) chromosomes in order to establish a genome-wide chromosomal correspondence between these Asiatic asses and the horse. Moreover, region-specific probes were generated to determine fusion configuration and orientation of...
Chen JW, Uboh CE, Soma LR, Li X, Guan F, You Y, Liu Y.Proper identification of racehorses competing in an official race and maintenance of defensible chain of custody are important in doping control regulations. The purpose of this study was to develop a reliable multiplex PCR method for providing genetic evidence for matching donors to test samples by using short tandem repeat (STR) loci. Amplification of 21 STR loci from blood, urine or hair root was achieved in a single tube and STR length polymorphism was analyzed using fluorescent labeled capillary electrophoresis. This novel approach showed an allele confidence interval of 0.19-0.43 bp and ...
Bacigalupo MA, Meroni G, Secundo F, Scalera C, Quici S.Highly luminescent Eu(3+) and Tb(3+) complexes of 10-[4-(3-isothiocyanatopropoxy)benzoylmethyl]-1,4,7,10-tetraazacyclododecane-1,4,7 triacetic acid Eu(3+) is a subset of 1 and Tb(3+) is a subset of 1 were conjugated with a goat anti-rabbit IgG and a rabbit anti-mouse IgG, respectively, and applied as markers in a time resolved immunoassay for simultaneous quantitative determination of anabolic compounds clenbuterol (CL) and hydrocortisone (HC). The assay was performed in horse urine, using a monoclonal antibody specific to CL and a rabbit polyclonal antibody specific to the free HC. These lant...
Epstein KL, Brainard BM, Lopes MA, Barton MH, Moore JN.To develop a standardized technique for thrombelastography (TEG) analysis in healthy adult horses, with and without the ex vivo addition of tissue factor (TF) as an activator. To determine reference intervals for TEG parameters in the horse, and to determine if traditional coagulation tests correlate with TEG. Methods: Prospective, observational. Methods: Veterinary teaching hospital. Methods: Twenty-six healthy adult horses. Methods: None. Results: Thrombelastography with (TF-TEG) and without (TEG) the addition of TF performed by 4 operators. Coagulation profiles (prothrombin time, activated ...
Tompkins D, Hudgens E, Horohov D, Baldwin CL.This report describes the initial cloning and characterization of the equine interleukin-17 (IL-17) expressed gene sequence from mRNA obtained from equine intestinal tissue and interleukin-23 (IL-23) expressed gene sequence from mRNA obtained from equine peripheral blood mononuclear cells. Equine IL-17 has 462 nucleotides in the translated region, determined by homology with known human and mouse sequences, and shares 84% and 75% identity, respectively. For the deduced amino acid sequences, the identity with human and mouse is 76% and 70%. Equine IL-23 has 579 nucleotides in the translated reg...
Casasola A, Ramos-Cerrillo B, de Roodt AR, Carbajal Saucedo A, Chippaux JP, Alagón A, Stock RP.Venoms of snakes belonging to the same Genera tend to share biochemical, toxinological and antigenic characteristics. Accordingly, paraspecific neutralization of venom lethality by experimental antisera and commercial antivenoms has been reported. We studied the spectrum of neutralization of lethality of an experimental monovalent equine antiserum against the strongly neurotoxic African forest cobra (Naja melanoleuca) when tested against venoms of most species of African Naja, both neuro and cytotoxic as described by some authors. We report a comparison of the median lethal doses (LD50) of the...
Frampton AR, Uchida H, von Einem J, Goins WF, Grandi P, Cohen JB, Osterrieder N, Glorioso JC.To initiate infection, equine herpesvirus type 1 (EHV-1) attaches to heparan sulfate on cell surfaces and then interacts with a putative glycoprotein D receptor(s). After attachment, virus entry occurs either by direct fusion of the virus envelope with the plasma membrane or via endocytosis followed by fusion between the virus envelope and an endosomal membrane. Upon fusion, de-enveloped virus particles are deposited into the cytoplasm and travel to the nucleus for viral replication. In this report, we examined the mechanism of EHV-1 intracellular trafficking and investigated the ability of EH...
Lin M, Zhang C, Gibson K, Rikihisa Y.Neorickettsia risticii is an obligate intracellular bacterium of the trematodes and mammals. Horses develop Potomac horse fever (PHF) when they ingest aquatic insects containing encysted N. risticii-infected trematodes. The complete genome sequence of N. risticii Illinois consists of a single circular chromosome of 879 977 bp and encodes 38 RNA species and 898 proteins. Although N. risticii has limited ability to synthesize amino acids and lacks many metabolic pathways, it is capable of making major vitamins, cofactors and nucleotides. Comparison with its closely related human pathogen N. senn...
Schmitz A, Theurillat R, Lassahn PG, Mevissen M, Thormann W.CE with multiple isomer sulfated-CD as selector was used for the simultaneous analysis of the stereoisomers of ketamine, norketamine, 5,6-dehydronorketamine and hydroxylated metabolites of norketamine in liquid/liquid extracts of (i) in vitro incubations with ketamine or norketamine and equine liver microsomes and (ii) plasma and urine of ponies receiving a target-controlled infusion of ketamine under isoflurane anesthesia. Hydroxynorketamine metabolites with the hydroxy group at the cyclohexanone ring could be shown to be formed stereoselectively both in vitro and in vivo. Due to the lack of ...
White SD, Foley JE, Spiegel IB, Ihrke PJ.Equine sarcoidosis is a rare, multisystemic, noncaseating, granulomatous and lymphoplasmacytic disease of unknown etiology. A recent report described a horse with granulomatous skin disease displaying histologic, electron microscopic, and polymerase chain reaction (PCR) findings consistent with equine herpesvirus 2 (EHV-2). Objective: To investigate the presence of EHV-2 and equine herpesvirus 1 (EHV-1) in 8 horses with sarcoidosis. Methods: Eight horses with sarcoidosis, reported previously. Methods: Retrospective study. PCR assays of the tissues were performed to detect DNA associated with E...
Christmann U, Hite RD, Witonsky SG, Elvinger F, Werre SR, Thatcher CD, Tan RH, Buechner-Maxwell VA.Surfactant alterations are described in horses after exercise, anesthesia, and prolonged transport, in horses with recurrent airway obstruction, and in neonatal foals. The effect of horse age or bronchoalveolar lavage fluid (BALF) sample characteristics on surfactant is unknown. Objective: To evaluate surfactant phospholipid composition and function in healthy horses, and to investigate the influence of age and BALF sample characteristics on surfactant. Methods: Seventeen healthy horses 6-25 years of age maintained on pasture year-round. Methods: BALF was collected by standard procedures and w...
Casaite V, Bruzyte S, Bukauskas V, Setkus A, Morozova-Roche LA, Meskys R.Equine lysozyme (EL) is a calcium (Ca)-binding lysozyme and is an intermediary link between non-Ca-binding C-type lysozyme and alpha-lactalbumin. The feature of lysozymes to assemble into the fibrils has recently gained considerable attention for the investigation of the functional properties of these proteins. To study the structural and functional properties of EL, a synthetic gene was cloned and EL was overexpressed in Escherichia coli as a fused protein. The His-tagged recombinant EL was accumulated as inclusion bodies. Up to 50 mg/l of the recombinant EL could be achieved after purificati...
Rendle DI, Heller J, Hughes KJ, Innocent GT, Durham AE.Time delays between collection of blood samples and biochemical analysis of equine blood are unavoidably common in equine practice. The effect that delays may have on the accuracy of results of blood biochemical analyses is not well established. Objective: Delays in processing of blood of up to 72 h results in alterations in measured levels of common biochemical analytes that are of potential clinical relevance. Separation of serum prior to storage is protective against the effects of time delays. Methods: Samples of clotted blood, separated serum and oxalate fluoride plasma from 20 horses wer...
Goodrich LR, Choi VW, Carbone BA, McIlwraith CW, Samulski RJ.Cell transplantation for the treatment of joint disease is an important clinical tool. Genetic modification of cells before transplantation has shown enhanced healing. Ex vivo genetic modification of joint tissue cells with various adeno-associated virus (AAV) serotypes has not been investigated. The transduction efficiencies of self-complementary AAV serotypes (1-6 and 8) were determined in joint tissue containing chondrocytes and synoviocytes isolated from equine models. When comparing scAAV serotypes for efficient transduction ex vivo, in chondrocytes versus synoviocytes, serotypes 6 and 2,...
Morita H, Shimazu M, Shiono H, Toh H, Nakajima F, Akita H, Takagi M, Takami H, Murakami M, Masaoka T, Tanabe S, Hattori M.We previously isolated five strains of putative lactobacilli from the faeces of a thoroughbred horse (a 4-year-old male). Of the five strains, four were identified as members of existing Lactobacillus species; however, sequence analysis of the 16S rRNA gene revealed that the fifth isolate, DI70(T), showed approximately 97 % identity (1325/1366 bp) with the type strain of Lactobacillus delbrueckii. Therefore, we considered the possibility that DI70(T) represents a novel species of the genus Lactobacillus. Cells of strain DI70(T) were Gram-stain-positive, catalase-negative, non-spore-forming, no...
Echigoya Y, Sato T, Itou T, Endo H, Sakai T.The species-specific properties of LDH isozymes are essentially determined by M (muscle) and H (heart) subunit proteins encoded by the LDHA and LDHB genes, respectively. In the present study, we molecularly characterized the full-length equine lactate dehydrogenase A (eLDHA) and B (eLDHB) cDNAs. The eLDHA cDNA consisted of a 999-bp open reading frame (ORF), while the eLDHB and newly acquired bat LDHB consisted of a 1002-bp ORF, which is 3 bp shorter than the LDHB ORF of other registered mammals. The alignment of amino acid sequences showed that eLDHA acquired positively charged His 88 and 226,...
Bugno M, Słota E, Pieńkowska-Schelling A, Schelling C.Fluorescent in situ hybridisation (FISH) using a panel of molecular probes for all chromosome pairs obtained by chromosome microdissection of the domestic horse ( Equus caballus ) was used to diagnose karyotype abnormalities in 35 horses (32 mares, 2 stallions and 1 intersex), which were selected for the study due to infertility (23 horses), reduced fertility (10 horses) and developmental anomalies (2 horses). The use of the FISH technique with probes for each horse chromosome pair enabled the diagnosis of many different chromosome aberrations in this population. Among the horses analysed, 21 ...
Glazar AI, Mullen SF, Liu J, Benson JD, Critser JK, Squires EL, Graham JK.Stallion spermatozoa exhibit osmotic damage during the cryopreservation process. Recent studies have shown that the addition of cholesterol to spermatozoal membranes increases the cryosurvival of bull, ram and stallion spermatozoa, but the exact mechanism by which added cholesterol improves cryosurvival is not understood. The objectives of this study were to determine if adding cholesterol to stallion sperm membranes alters the osmotic tolerance limits and membrane permeability characteristics of the spermatozoa. In experiment one, stallion spermatozoa were treated with cholesterol-loaded cycl...
Cook RB, Zioupos P.The mechanical capacity and integrity of cancellous bone is crucial in osteoporosis, a condition which is set to become more prevalent with increasing lifespan and population sizes. The fracture toughness (FT) of cancellous bone has never been examined before and the conditions associated with the growth of a major crack through the lattice of cancellous bone, a cellular solid, may improve our understanding for structural integrity of this material. The aim of this study is to provide (i) basic data on cancellous bone FT and (ii) the experimental support for the hypothesis of Gibson, L.J., Ash...
Labeda DP, Price NP, Kroppenstedt RM, Donahue JM, Williams NM, Sells SF.Two actinomycete strains, NRRL B-24165(T) and NRRL B-24166(T), isolated from lesions on equine placentas in Kentucky, USA, were analysed using a polyphasic taxonomic approach. On the basis of phylogenetic analysis of 16S rRNA gene sequences, morphological observations and the presence of ll-diaminopimelic acid as the diagnostic diamino acid in whole-cell hydrolysates, the new isolates clearly belonged to the genus Streptomyces. Analyses of the phylogenetic positions of strains NRRL B-24165(T) and NRRL B-24166(T) based on 16S rRNA gene sequences of all recognized species of the genus Streptomyc...
Andoh K, Kai K, Matsumura T, Maeda K.Cell lines originating from horses are necessary for isolation and propagation of equine herpesviruses (EHV). Although we established an equine-derived cell line, FHK-Tcl3, propagation ceased after fewer than 40 passages. In this study, FHK-Tcl3 cell propagation continued beyond 40 passages, achieving over 100 passages. FHK-Tcl3 cells were then cloned by limiting dilution at the 100th passage. Cloned cells were termed FHK-Tcl3.1. FHK-Tcl3.1 cells grew well and were propagated every 3 to 4 days by splitting 1:5. In addition, EHV-1, -2 and -4 showed a clear cytopathic effect (CPE) in FHK-Tcl3.1 ...
Ortega Ferrusola C, González Fernández L, Macías García B, Salazar-Sandoval C, Morillo Rodríguez A, Rodríguez Martinez H, Tapia JA, Peña FJ.The ability of stallion spermatozoa to produce nitric oxide (NO) before (fresh) and after freezing and thawing (FT) was evaluated by means of flow cytometry after loading the sperm suspension with the probe, 4,5-diaminofluorescenin diacetate. The presence of NO synthase (NOS) was investigated by Western blotting using anti-NOS1, anti-NOS3, or anti-universal NOS antibodies (Abs). While NO was detected both in fresh and FT sperm suspensions, its production increased after cryopreservation only when egg yolk was removed from the extender. Anti-NOS1 Ab intensively labeled a single band with an app...
Morrell JM, Johannisson A, Dalin AM, Rodriguez-Martinez H.The objective of the current study was to optimize the volumes of Androcoll-E and sperm sample used in various sizes of centrifuge tube to scale up single-layer centrifugation (SLC) for routine use in the field. Although sperm suspensions of equivalent quality were produced using Androcoll-E in small and large tubes, the sperm yield was much lower in the latter (P<0.001). In contrast, in 200-mL tubes (XL), the yields were approximately 25% higher than those for the small tubes. An increased volume (4.5 mL) of extended ejaculate in small tubes (SLC-Inc) or 15 to 18 mL extended ejaculate on 1...
Kieken F, Pinel G, Antignac JP, Monteau F, Christelle Paris A, Popot MA, Bonnaire Y, Le Bizec B.Despite the worldwide existing regulation banning the use of the recombinant equine growth hormone (reGH) as growth promoter, it is suspected to be used in horseracing to improve performances. Various analytical methods previously developed to screen for its misuse have encountered some limitations in terms of detection timeframes, in particular during the first days following reGH administration. A novel strategy involving the characterization of global metabolomic fingerprints in urine samples of non-treated and reGH-treated horses by liquid chromatography-electrospray-high-resolution mass s...
Leprêtre PM, Metayer N, Giovagnoli G, Pagliei E, Barrey E.Measurements of minute ventilation (VE) and expired oxygen and carbon dioxide fractions (FeO2 and FeCO2) were measured at rest and during exercise in seven warmblood horses performing two consecutive standardised incremental treadmill exercise tests at submaximal speed, using the portable K4b2 telemetric unit and the laboratory Quark metabolic cart in random order. Oxygen consumption (VO2) and carbon dioxide production (VCO2) were estimated using the Haldane equation. There were no significant differences between the measurements made with the two devices. However, VE was overestimated when th...
Espino-Solis GP, Calderon-Amador J, Calderon-Aranda ES, Licea AF, Donis-Maturano L, Flores-Romo L, Possani LD.A three-dimensional model of the alphaX I-domain of the horse integrin CD11c from dendritic cells provided information for selecting two segments of the primary structure for peptide synthesis. Peptide 1 contains 20 amino acids and peptide 2 has 17 amino acid residues. The first spans from position Thr229 to Arg248 of an alpha-helix segment of the structure, whereas peptide 2 goes from Asp158 to Phe174 and corresponds to an exposed segment of the loop considered to be the metal ion-dependent adhesion site. Murine polyclonal antisera against both peptides were generated and assayed in periphera...
McKinney AR.Control of the use of performance-affecting substances in the horse is critical to the integrity of a wide range of equine sports, with major implications for both animal welfare and revenue streams. One class of medications enjoying particular public notoriety is the anabolic-androgenic steroid group, as highlighted by the recent 'Big Brown' affair and Congressional inquiries into the use of steroids in professional sports, including horse racing, in the USA. This review examines the latest developments pertaining to the analytical detection of these substances in equine biological samples an...
Desjardin C, Balliau T, Valot B, Zivy M, Wimel L, Guérin G, Cribiu E, Schibler L.Proteomic analyses of cartilage and, to a lesser extent, of bone have long been impaired because of technical challenges related to their structure and biochemical properties. We have developed a unified method based on phenol extraction, 2DE, silver staining, and subsequent LC-MS/MS. This method proved to be efficient to characterize the proteome of equine cartilage and bone samples collected in vivo. Since proteins from several cellular compartments could be recovered, our procedure is mainly suitable for in situ molecular physiology studies focused on the cellular content of chondrocytes, o...
Ahmed YA, Ali S, Ghallab A.Animal hair examination at a criminal scene may provide valuable information in forensic investigations. However, local reference databases for animal hair identification are rare. In the present study, we provide differential histological analysis of hair of some domestic animals in Upper Egypt. For this purpose, guard hair of large ruminants (buffalo, camel and cow), small ruminants (sheep and goat), equine (horse and donkey) and canine (dog and cat) were collected and comparative analysis was performed by light microscopy. Based on the hair cuticle scale pattern, type and diameter of the me...
Mathis D, Sass JO, Graubner C, Schoster A.Atypical myopathy (AM), an acquired multiple acyl-CoA dehydrogenase deficiency (MADD) in horses, induce changes in mitochondrial metabolism. Only few veterinary laboratories offer diagnostic testing for this disease. Inborn and acquired MADD exist in humans, therefore determination of organic acids (OA) in urine and acylcarnitines (AC) in blood by assays available in medical laboratories can serve as AM diagnostics. The evolution of OA and AC profiles in surviving horses is unreported. Methods: AC profiles using electrospray ionization tandem mass spectrometry (ESI-MS/MS) and OA in urine using...
Rapacz-Leonard A, Raś A, Całka J, Janowski TE.Fetal membrane retention can be a life-threatening condition and its incidence exceeds 50% in heavy draught mares. Although fetal membrane retention is commonly treated with repeated injections of oxytocin, based on the suggestion that it is caused mainly by secondary atony of the uterus, this treatment sometimes fails. This led us to ask if expression of oxytocin receptors differs in mares that retain fetal membranes due to secondary uterine atony. Objective: To determine whether expression of oxytocin receptors in equine placental tissues differs when heavy draught mares expel fetal membrane...
Kadivar A, Shams Esfandabadi N, Dehghani Nazhvani E, Shirazi A, Ahmadi E.Protamines substitute DNA-binding histones during late spermatogenesis in sperm nucleus. Stallion sperm contains all three variants of these arginine-rich and positively charged nuclear proteins (P1, P2 and P3). Two variants of protamine-2, that is P2 and P3, constitute approximately 15% of the entire protamine content. Also, the ratio of protamine-1 to protamine-2 varies among different mammalian species, and abnormal protamine ratios and protamine content are correlated with male infertility. In this study, changes in protamine mRNA abundance for all three protamines were investigated in sta...
Zhang L, Hu M, Chilton NB, Huby-Chilton F, Beveridge I, Gasser RB.The expansion segments or divergent (D) domains in the large subunit (LSU) of the ribosomal DNA have been suggested as genetic markers for taxonomic and/or phylogenetic studies of parasites. In the present study, we assessed the degree of sequence variation in the D3 domain and flanking core regions of the LSU for 21 species of equine strongyles (Strongylida: Strongylidae) and determined which positions in the secondary structure of the LSU were associated with the nucleotide alterations. No intraspecific sequence variation was detected in 17 species, for which multiple individual worms were a...
Zientara S, Sailleau C, Moulay S, Crucière C, el-Harrak M, Laegreid WW, Hamblin C.A coupled reverse transcriptase-polymerase chain reaction assay (RT-PCR) for the detection of African horse sickness virus (AHSV) dsRNA, has been developed using genome segment 7 as the target template for primers. RNA from isolates of all nine AHSV serotypes were readily detected. The potential inhibitory effects of either ethylene diamine tetra acetic acid (EDTA) or heparin on the RT-PCR were eliminated by washing blood samples before lysis of the red blood cells and storage. There was a close agreement in the sensitivity and the specificity of the RT-PCR and an indirect sandwich ELISA. Conf...
Heneine LG, Carvalho AD, Barbosa CF, Arávjo dos Santos MR.The objective of this study was the search for a suitable venom antigen to be used in an in vitro alternative immunoassay, to the standard antivenom neutralization assay using mice. Bothrops jararaca venom was fractionated in DEAE-Sephacel columns and the fractions were tested for a correlation between antibody capture enzyme linked immunosorbent assay (ELISA) absorbance values and the 'in vivo' antivenom potency. Individual antivenoms from 14 horses and 15 separate FUNED polyspecific Bothrops ampouled antivenoms (final product) were used. Fractions showing the higher correlations were further...
Ryan C, Giguère S, Hagen J, Hartnett C, Kalyuzhny AE.Peripheral blood mononuclear cells (PBMCs) were obtained from 6 foals <1 week of age, 6 foals between 3 and 4 months of age, and 10 adult horses. PBMCs were stimulated with concanavalin A (ConA) or calcium ionomycin-phorbol myristate acetate (CaI-PMA) and the frequency of interferon IFN-gamma and IL-4 secreting cells was measured using an equine-specific ELISPOT assay. The number of IFN-gamma secreting cells was significantly lower in both groups of foals than in adult horses regardless of the mitogen used for stimulation. The number of IFN-gamma secreting cells was significantly higher in ...
Budachetri K, Lin M, Yan Q, Chien RC, Hostnik LD, Haanen G, Leclère M, Waybright W, Baird JD, Arroyo LG, Rikihisa Y.Potomac horse fever (PHF) is an acute and potentially fatal enterotyphlocolitis of horses with clinical signs that include anorexia, fever, diarrhea, and laminitis. Its incidence is increasing despite a commercially available vaccine. PHF is caused by Neorickettsia risticii, and the recently rediscovered and classified . PHF diagnosis is currently accomplished using serology or nested PCR. However, both methods cannot distinguish the two species that cause PHF. Further, the current real-time PCR test fails to detect Thus, in this study, two species-specific real-time PCR assays based on a...
Racine T, Denizot M, Pannetier D, Nguyen L, Pasquier A, Raoul H, Saluzzo JF, Kobinger G, Veas F, Herbreteau CH.There is no vaccine or approved therapy against lethal Ebola virus (EBOV). We investigated a proven technology platform to produce polyclonal IgG fragments, F(ab')2, against EBOV. Horses immunized with nanoparticles harboring surface glycoprotein trimers of EBOV-Zaire/Makona produced anti-Ebola IgG polyclonal antibodies with high neutralization activity. Highly purified equine anti-Ebola F(ab')2 showed strong cross-neutralization of 2 Zaire EBOV strains (Gabon 2001 and Makona) and in vivo 3 or 5 daily F(ab')2 intraperitoneal injections provided 100% protection to BALB/c mice against lethal EBO...
Griffin RA, Baker M, Aitken RJ, Swegen A, Gibb Z.Stallions experience lower per-cycle conception rates compared to other livestock species, largely because they are selected for breeding based on athletic prowess and not reproductive fitness. Mares are seasonal breeders, and pregnancies cannot be detected until 10-14 days post cover via transrectal ultrasonography. This means the detection of stallion fertility fluctuations is delayed by at least 2 weeks, which within the short breeding season employed by the thoroughbred horse breeding industry, can prove quite costly. For these reasons, there is increased demand for robust laboratory assay...
Hu Z, Wu X, Ge J, Wang X.Type I interferons (IFNs) play important roles in the defense of host cells against viral infection by inducing the expression of a diverse range of antiviral factors. IFNs from different animals likely share similar features with human IFNs, and some of them have cross-species activities. Equine IFN-α was proved effective in both equine and human cells. However, the previous studies mostly focused on the inhibition of virus induced cytopathic effects. In this study, we used virus-specific assays to demonstrate the antiviral activities of equine IFN-α1 in both equine and human cells. Equine ...
Koskinen E, Lindeberg H, Kuntsi H, Katila T.Twenty-four Finnhorse mares were examined by rectal palpation and ultrasonography every 6 h during late oestrus to determine the time of ovulation. Milk and serum samples were collected every 6 h after the detected ovulation for progesterone analysis. The progesterone rises took place within 0-54 h and 0-60 h after ovulation, in milk and serum, respectively. Statistically significant differences (p less than 0.05) in progesterone levels were observed for the first time 12-18 h and 18-24 h after ovulation, in serum and milk, respectively, as compared to progesterone levels 0-6 h after ovulation...
Sahu SP, Alstad AD, Pedersen DD, Pearson JE.Immunoglobulin M (IgM) and G (IgG) capture enzyme-linked immunosorbent assays (ELISAs) were used as possible adjuncts to hemagglutination inhibition (HI) and virus neutralization (VN) tests to differentiate between reaction to recent exposure to eastern equine encephalomyelitis (EEE) virus and those due to prior vaccination. Serum samples were evaluated by the IgM-capture ELISA, and the results were compared with those of HI and VN tests. Of 381 serum samples, 51% (195 samples) were positive by HI test (> or = 1:40) and 54% (205 samples) were positive by VN test (> or = 1:10), but only 3...
Yadav SC, Kumar R, Kumar V, Jaideep , Kumar R, Gupta AK, Bera BC, Tatu U.Trypanosoma evansi is the most extensively distributed trypanosome responsible for disease called surra in livestock in many countries including frequent outbreaks in India. The prevalence of this disease is most commonly reported by standard parasitological detection methods (SPDM); however, antibody ELISA is being in practice by locally produced whole cell lysate (WCL) antigens in many countries. In the present investigation, we attempted to identify and purify immuno dominant, infection specific trypanosome antigens from T. evansi proteome using experimentally infected equine serum by immun...
Zinkl JG, Brown PD.Horse polymorphonuclear leukocytes (PMN) isolated from horse blood by sedimentation and isotonic lysis and having about 25% accompanying lymphocytes were as effective at chemotaxis as nearly pure PMN isolated by density gradient techniques. N-Formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP), used as a representative of the formylmethionyl peptides (produced by prokaryocytic organisms), was effective as a chemoattractant only at the high concentration of 10(-4) M. When serum was preincubated with FMLP at concentrations as low as 10(-8) M, the serum attracted horse PMN. This activity was not g...
Vaiman M, Chardon P, Cohen D.In the past few years it has been possible by combining enzymatic cleavage of genomic DNA and the Southern blot hybridization technique to explore the endonuclease recognition site polymorphism of the MHC. HLA class I and DR and DQ alpha and beta class II specific probes as well as human C4 and Bf class III probes were used. All these probes were shown to cross-hybridize with DNA from pigs, cattle, sheep and horses. Hybridization of human genomic DNA with a class I probe showed 15-25 bands per genome depending on the enzyme used. Distinct endonucleases generated clusters of restriction fragmen...
Shubitowski DM, Venta PJ, Douglass CL, Zhou RX, Ewart SL.The continued discovery of polymorphisms in the equine genome will be important for future studies using genomic screens and fine mapping for the identification of disease genes. Segments of 50 equine genes were examined for variability in 10 different horse breeds using a pool-and-sequence method. We identified 11 single nucleotide polymorphisms (SNPs) in 9380 bp of sequenced exon, and 25 SNPs, six microsatellites, and one insertion/deletion in 16961 bp of sequenced intron. Of all genes studied 52% contained at least one polymorphism, and polymorphisms were found at an overall rate of 1/613 b...
Tozaki T, Ohnuma A, Hamilton NA, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Kusano K, Nagata SI.Gene doping is prohibited for fair competition in human and horse sports. One style of gene doping is the administration of an exogeneous gene, called a transgene, to postnatal humans and horses. Although many transgene detection methods based on quantitative polymerase chain reaction (PCR), including real-time PCR and digital PCR, have been recently developed, it remains difficult to reliably detect low-copy transgenes. In this study, we developed and validated a nested digital PCR method to specifically detect low-copy transgenes. The nested digital PCR consists of (1) preamplification using...
Svendsen CK, Hjortkjaer RK, Hesselholt M.42 horses were examined. The physical signs with relation to circulatory insufficiency and the abdominal disease were registered following a two-phased examination procedure. Great prognostic value was found in the degree of circulatory insufficiency judged by pulse rate and character, filling of the jugular vein, skin temperature, colour of mucous membranes, capillary refill time, sweating, depression, skin turgor and degree of enophthalmus. In making a causal diagnosis the abdomen was examined for shape, tenderness, peristaltic sounds, gastric dilation by siphoning, abnormal rectal findings ...
Banse HE, Schultz N, McCue M, Geor R, McFarlane D.Accurate measurement of equine adrenocorticotropin (ACTH) is important for the diagnosis of equine pituitary pars intermedia dysfunction (PPID). Several radioimmunoassays (RIAs) and chemiluminescent immunoassays (CIAs) are used for measurement of ACTH concentration in horses; whether these methods yield similar results across a range of concentrations is not determined. We evaluated agreement between a commercial RIA and CIA. Archived plasma samples ( n = 633) were measured with both assays. Correlation between the 2 methods was moderate ( r = 0.49, p < 0.001). Bland-Altman analysis reveale...
Westcott DG, King DP, Drew TW, Nowotny N, Kindermann J, Hannant D, Belák S, Paton DJ.Routine detection of equine arteritis virus (EAV) can be achieved by virus isolation (VI) in cell culture, or by the amplification of viral genome by molecular methods. To simplify molecular diagnosis, a number of different Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) and RT-nested PCR (RT-nPCR) assays were compared, and a one-tube method was developed and optimised utilizing a fluorogenic probe (TaqMan). An artificial RNA template (Mimic) and associated probe were also constructed to provide in-tube validation of the RT-PCR system. To assess the utility of the RT-PCR TaqMan assay,...
Laughlin AM, Welsh TH, Love CC, Varner DD, Parrish AR, Forrest DW, Ing NH.In vitro culture systems are valuable tools for investigating reproductive mechanisms in the testis. Here, we report the use of the precision-cut in vitro system using equine testicular slices. Testes were collected from immature light breed stallions (n=3) and cut into slices (mean slice weight= 13.85 ± 0.20 mg; mean slice thickness=515.00 ± 2.33 μm) using the precision-cut tissue-slicing method. Four tissue slices were placed on a grid floating on medium in individual vials. After a 1-h preincubation, they were exposed to medium containing ovine luteinizing hormone (oLH) at concentrations...
Herrick WC, Kingsbury HB, Lou DY.This paper describes the result of an investigation of strains and strain rates which normally occur in the tendons of the equine foreleg and presents stress-strain curves and moduli for the tendons at these rates. It has previously been demonstrated that resistance to flexion of the joints of the distal part of the equine foreleg is provided by a passive system of tendons and ligaments. It is therefore possible, using a large displacement, high-rate testing machine, to duplicate in the laboratory the strain rates and forces which are normally produced in the tendons of the foreleg of the runn...
Buechner-Maxwell V, Zhang C, Robertson J, Jain NC, Antczak DF, Feldman BF, Murray MJ.Leukemia is a neoplastic disease of one or more of the cell types of the hemopoietic system and is rarely diagnosed in the horse. This report describes a case of subleukemic acute myelomonocytic leukemia in an 11-year-old gelding. Preliminary cytological diagnosis was supported by two types of laboratory investigations. Cytochemical characterization of blood and bone marrow neoplastic cells was consistent with a myelomonocytic origin. Neoplastic blast cells in peripheral blood were labeled by monoclonal antibodies specific for cell surface molecules of horse granulocytes, but they were not lab...
Khan SH, Islam A, Hassan MI, Sharma S, Singh TP, Ahmad F.A sequence alignment of horse cytochrome c (cyt c) with all known cyts c shows that Leu at position 94 is conserved, except in 14 species which have either Val or Ile at this position. It is also known that Leu94 of the mammalian cyt c plays an important role in folding and stability. The important question here is as to what will happen in terms of folding and stability if Leu94 of the mammalian cyt c is substituted by Val or Ile. To answer this question, we introduced natural substitutes of Leu94 by Val and Ile in horse cyt c. The purified L94V and L94I mutants under native condition (pH 6.0...
Pozor M, Conley AJ, Roser JF, Nolin M, Zambrano GL, Runyon SP, Kelleman AA, Macpherson ML.Recently, anti-Müllerian hormone (AMH) was validated as a reliable marker of testicular damage caused by various chemotherapy drugs in humans and in mice. In horses, the reference values of AMH concentrations in normal stallions, during different seasons of a year, have been recently reported. However, this hormone was not evaluated in subfertile or infertile stallions with testicular damage. Therefore, the objective of this study was to investigate the effects of experimentally induced testicular degeneration on the concentration of AMH in stallions. Severe but transient testicular degenerat...
Bailey SR, Elliott J.5-hydroxytryptamine (5-HT) is a potent vasoconstrictor of equine digital arteries and veins which may play a role in the ischaemic disease, laminitis. The present investigation compared the properties of 5-HT1B/1D receptors in arteries with those in veins using isolated rings of equine digital blood vessels. The 5-HT1B/1D receptor-selective agonists, anpirtoline and sumatriptan were 17.9 and 10 times more potent and produced 4.1 and 5.6 times greater maximum contractions, respectively, in veins when compared to arteries. Other agonists tested were of equal potency and produced the same maximum...
Green SA, Jenkins SJ, Clark PA.The biuret total protein method and a bromcresol green (BCG) albumin method were used on the Abbott ABA-100 chemistry analyzer to assay serum proteins in clinically normal cattle, sheep, ponies, pigs, and ducks. Total proteins were also read on a refractometer and mylar supported cellulose acetate electrophoresis was performed. Globulins and A/G ratios were calculated from the chemical method and the results compared with the electrophoretic method. Total protein, albumin and A/G ratios in the ponies, sheep and older cattle were in agreement between the two methods. The younger cattle and all ...
