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.
Bae JH, Ahn K, Nam GH, Lee CE, Park KD, Lee HK, Cho BW, Kim HS.The horse BMAL1 gene encodes the brain and muscle Arnt-like protein 1, which is a key regulator of circadian rhythmic systems in most organs and cells. The first exon of the horse-specific BMAL1 gene is produced by an exonization event of LINE3 (CR1) and SINE (MIR) was detected by bioinformatic analysis. Alternative variants generated by cassette exon event in various horse tissues were also detected by RT-PCR amplification and sequencing. The cDNA sequences of the horse transcripts (BMAL1a, BMAL1b) contain additional 21 bp and 71 bp fragments relative to horse BMAL1. Quantitative real-time RT...
Parker RA, Clegg PD, Taylor SE.To investigate the effects of commonly used antibiotics on cell viability and gene expression of equine bone marrow-derived mesenchymal stromal cells (MSC) in vitro. Methods: Bone marrow-derived MSC were cultured in media containing gentamicin, amikacin, penicillin, enrofloxacin or ceftiofur at concentrations of 50, 100, 200 and 500 µg/ml. The alamarBlue fluorescence assay was used to assess cell viability over 48 h. After 5 days the cells were released and lysed prior to RNA extraction and reverse transcription. RNA levels were assessed using spectrophotometry and quantitative PCR was used...
Epstein KL, Brainard BM.Accurate measurement of plasma fibrinogen concentrations is an important tool for assessment of horses with inflammatory diseases. Objective: To determine the precision and accuracy of a benchtop instrument using both fresh and frozen equine plasma by comparing the plasma fibrinogen concentration measured by a benchtop instrument to 2 separate laboratory standard methods (ACL 100 and STA Compact) for fibrinogen measurement. Methods: Accuracy and precision of the VSPro was evaluated using both human fibrinogen standards and samples from horses. Fifty frozen samples from horses with gastrointest...
Taylor SE, Clegg PD.Mesenchymal stromal cells (MSC) are derived from adult mesenchymal tissues and have the ability to undergo differentiation into bone, cartilage, and fat, and have therefore attracted great interest in regenerative medicine. Many isolation and culture methods have been described, making comparison between laboratories and quality-control protocols difficult. A uniform protocol to characterize equine MSC has recently been proposed, aiming to introduce consistency across the equine stem cell research field. This article reviews the published techniques for collection and propagation of equine MSC...
Taddei L, Benoit M, Sukta A, Peterson J, Gaensslen RE, Negrusz A.In order to protect the integrity of horse racing in Illinois, a complex testing of urine and blood specimens collected post-race from winning and special designation horses is continuously conducted. The initial screening by immunoassays was followed by the confirmation on presumptive positive samples. Instrumental screening was also conducted. Perimortem and postmortem specimens and special exhibits (syringes, needles, etc.) were also analyzed. The administration of alkalinizing agents was detected by measuring the total plasma carbon dioxide concentration. The laboratory analyzed specimens ...
Nemoto M, Yamanaka T, Bannai H, Tsujimura K, Kondo T, Matsumura T.Reverse transcription loop-mediated isothermal amplification (RT-LAMP) was applied to the detection of equine influenza virus (EIV). Because equine influenza is caused currently by EIV of the H3H8 subtype, the RT-LAMP primer set was designed to target the hemagglutinin gene of this subtype. The detection limit of the RT-LAMP assay was a virus dilution of 10(-5); which was 10(3) times more sensitive than the Espline Influenza A&B-N test and 10 times more sensitive than a reverse transcription polymerase chain reaction (RT-PCR) assay. The specificity of the RT-LAMP assay was examined by usin...
Menozzi A, Pozzoli C, Zullian C, Poli E, Serventi P, Bertini S.The effects of preferential µ (morphine), selective µ (fentanyl), selective κ (compound U69593) opioid receptor agonists, and nonselective (naloxone) and selective µ (naloxonazine) antagonists on equine small intestinal motility were evaluated in vitro. Samples of circular muscle from equine jejunum were placed in isolated organ baths and drug-induced modifications of both spontaneous and electrically evoked contractile activity were measured. None of the opioid agonists induced a significant change in spontaneous contractions. Fentanyl and U69593 reduced electrically induced contractions,...
Teixeira-Neto FJ, McDonell WN, Black WD, Harris W, Grovum L.This study investigated the effects of a muscarinic type 1 (M(1)), 2 (M(2)), and 3 (M(3)) antagonists (4-DAMP, pirenzepine, and methoctramine, respectively) on acetylcholine (Ach)-induced contractions of longitudinal jejunal muscle strips of horses. Strips were irrigated with Krebs-Henseleit solution gassed with 95% O(2) and 5% CO(2), and the developed tension in response to Ach was recorded before and after incubation with increasing concentrations of 4-DAMP (10(-8)-10(-6) M), pirenzepine (10(-6)-10(-4) M), and methoctramine (10(-5)-10(-3) M). When competitive antagonism was characterized, th...
Wagner B, Freer H, Rollins A, Erb HN, Lu Z, Gröhn Y.Lyme disease is a zoonotic, vector-borne disease and occurs in mammals including horses. The disease is induced by infection with spirochetes of the Borrelia burgdorferi sensu lato group. Infection of mammalian hosts requires transmission of spirochetes by infected ticks during tick bites. Lyme disease diagnosis is based on clinical signs, possible exposure to infected ticks, and antibody testing which is traditionally performed by ELISA and Western blotting (WB). This report describes the development and validation of a new fluorescent bead-based multiplex assay for the detection of antibodie...
Kirkland PD, Davis RJ, Wong D, Ryan D, Hart K, Corney B, Hewitson G, Cooper K, Biddle A, Eastwood S, Slattery S, Rayward D, Evers M, Wright T....Until August 2007, Australia was one of only three countries internationally recognised to be free of equine influenza (EI). This report documents the diagnosis of the first cases of EI in Australian horses and summarises the investigations that took place over the next 5 days. During that time, a multifocal outbreak was identified across eastern New South Wales and south-eastern Queensland. The use of an influenza type A pan-reactive real-time reverse transcription polymerase chain reaction allowed rapid confirmation of suspect cases of EI.
Bracken MK, Wøhlk CB, Petersen SL, Nielsen MK.Strongyle parasites are ubiquitous in grazing horses. Of these, the bloodworm Strongylus vulgaris is regarded as most pathogenic. Increasing levels of anthelmintic resistance in strongyle parasites has led to recommendations of decreased treatment intensities, and there is now a pronounced need for reliable tools for detection of parasite burdens in general and S. vulgaris in particular. The only method currently available for diagnosing S. vulgaris in practice is the larval culture, which is laborious and time-consuming, so veterinary practitioners most often pool samples from several horses ...
Silaghi C, Liebisch G, Pfister K.Equine Granulocytic Anaplasmosis (EGA) is caused by Anaplasma phagocytophilum, a tick-transmitted, obligate intracellular bacterium. In Europe, it is transmitted by Ixodes ricinus. A large number of genetic variants of A. phagocytophilum circulate in nature and have been found in ticks and different animals. Attempts have been made to assign certain genetic variants to certain host species or pathologies, but have not been successful so far. The purpose of this study was to investigate the causing agent A. phagocytophilum of 14 cases of EGA in naturally infected horses with molecular methods o...
Toth AM, Geisler C, Aumiller JJ, Jarvis DL.In an effort to produce processed, soluble Western equine encephalitis virus (WEEV) glycoproteins for subunit therapeutic vaccine studies, we isolated twelve recombinant baculoviruses designed to express four different WEEV glycoprotein constructs under the transcriptional control of three temporally distinct baculovirus promoters. The WEEV glycoprotein constructs encoded full-length E1, the E1 ectodomain, an E26KE1 polyprotein precursor, and an artificial, secretable E2E1 chimera. The three different promoters induced gene expression during the immediate early (ie1), late (p6.9), and very lat...
Liu Y, Uboh CE, Soma LR, Li X, Guan F, You Y, Chen JW.Multiple drug target analysis (MDTA) used in doping control is more efficient than single drug target analysis (SDTA). The number of drugs with the potential for abuse is so extensive that full coverage is not possible with SDTA. To address this problem, a liquid chromatography tandem mass spectrometric method was developed for simultaneous analysis of 302 drugs using a scheduled multiple reaction monitoring (s-MRM) algorithm. With a known retention time of an analyte, the s-MRM algorithm monitors each MRM transition only around its expected retention time. Analytes were recovered from plasma ...
Stahl J, Braun M, Siebert J, Kietzmann M.A water based combination of 0.1% octenidine dihydrochloride and 2% 2 - phenoxyethanol is registered in many European countries as an antiseptic solution (octenisept®) for topical treatment with high antimicrobial activity for human use, but octenidine based products have not been registered for veterinary use yet. The aim of the present study was to investigate whether octenidine dihydrochloride or 2 -phenoxyethanol, the two main components of this disinfectant, permeate through animal skin in vitro. Therefore, permeation studies were conducted using Franz-type diffusion cells. 2 ml of the t...
Miszczak F, Shuck KM, Lu Z, Go YY, Zhang J, Sells S, Vabret A, Pronost S, Fortier G, Timoney PJ, Balasuriya UB.This study showed that under specifically defined conditions with respect to nucleic acid extraction method and testing reagents, a previously described real-time reverse transcription-PCR (rRT-PCR) assay (T1 assay) provides sensitivity equal to or higher than that of virus isolation for the detection of equine arteritis virus in semen.
Mohammadi-Sangcheshmeh A, Held E, Ghanem N, Rings F, Salilew-Wondim D, Tesfaye D, Sieme H, Schellander K, Hoelker M.Efficiencies for in vitro production of equine embryos are still low due to highly variable developmental competences of equine immature oocytes. In contrast to the equine, in vitro developmental competence of immature oocytes has been predicted successfully by the activity of glucose-6-phosphate dehydrogenase (G6PDH) indicated by brilliant cresyl blue (BCB) dye in a range of different species. Therefore, the aim of the present study was to test the association between G6PDH activity in equine oocytes with: (1) cumulus morphology and oocyte properties in terms of diameter and volume; (2) matur...
Fintl C, Pearson GT, Mayhew IG, Hudson NP.The generation and maintenance of intestinal motility patterns involve the complex interactions of several components including the gastrointestinal pacemaker cells (interstitial cells of Cajal, ICC). Central to ICC function is the generation of rhythmic pacemaker currents, namely slow waves, which represent the rate limiting step for intestinal smooth muscle contractions. Currently, intracellular slow wave activity has not been demonstrated in the equine colon. Objective: To characterise the in vitro myoelectrical activity of the equine pelvic flexure using intracellular recording techniques....
Sutton DG, Preston T, Love S.Validation of a reliable, noninvasive clinical test for quantification of equine orocaecal transit time (OCTT) is required. This would facilitate an evidence-based approach to investigation and treatment of equine small intestinal disorders. Objective: 1) Comparison of the lactose (13) C-ureide breath test (LUBT) with the hydrogen breath test (H(2) BT) for OCTT measurement. 2) Identification of the characteristics of gastrointestinal microbial glycosylureide hydrolase activity in vitro. 3) Production of an optimised protocol for the LUBT for in vivo measurement of equine OCTT. Objective: Signi...
Ibrahim S, Steinbach F.Earlier studies investigating the cross-reactivity of antibodies submitted to the HLDA8 had used flow cytometry as a method of choice to screen mAbs for reactivity with equine leukocytes, including two-color flow-cytometry to characterize the lymphocyte population they detect. In addition, immuno-histochemistry (IHC) was used to detect distribution of positive cells in lymphoid tissue sections. In this study we performed immunoprecipitation (IP) to complement the previous results and add valuable information regarding the molecules detected by the cross-reacting antibodies. Surface molecules f...
Lu Z, Kasaragod DK, Matcher SJ.We present a phase fluctuation calibration method for polarization-sensitive swept-source optical coherence tomography (PS-SS-OCT) using continuous polarization modulation. The method uses a low-voltage broadband polarization modulator driven by a synchronized sinusoidal burst waveform rather than an asynchronous waveform, together with the removal of the global phases of the measured Jones matrices by the use of matrix normalization. This makes it possible to average the measured Jones matrices to remove the artifact due to the speckle noise of the signal in the sample without introducing aux...
Saint-Dizier M, Foulon-Gauze F, Lecompte F, Combarnous Y, Chopineau M.In order to investigate the role of the unique seventh N23-glycosylation site of the equine LH/CG receptor (eLHCGR) in the cAMP pathway activation, COS-7 cells were transiently transfected with either the wild-type or the mutant eLHCGR(N23Q) cDNA and challenged with porcine LH and eCG for cAMP production. We showed that the N23-glycosylation site of the eLHCGR is not required for the functional coupling of the receptor with the cAMP pathway and is not responsible for the limited potency of eCG relative to pLH to activate this receptor.
Orlando L, Ginolhac A, Raghavan M, Vilstrup J, Rasmussen M, Magnussen K, Steinmann KE, Kapranov P, Thompson JF, Zazula G, Froese D, Moltke I....Second-generation sequencing platforms have revolutionized the field of ancient DNA, opening access to complete genomes of past individuals and extinct species. However, these platforms are dependent on library construction and amplification steps that may result in sequences that do not reflect the original DNA template composition. This is particularly true for ancient DNA, where templates have undergone extensive damage post-mortem. Here, we report the results of the first "true single molecule sequencing" of ancient DNA. We generated 115.9 Mb and 76.9 Mb of DNA sequences from a permafrost-...
Morrell JM, Garcia BM, Pena FJ, Johannisson A.The purpose of this study was to determine if the quality of stored stallion semen doses could be enhanced by the scaled-up version of Single Layer Centrifugation using Androcoll-E-Large. Three semen doses from each of fifteen stallions were transported overnight to the Swedish University of Agricultural Sciences (SLU) for processing 24 h after semen collection. Sperm quality in the resulting SLC-selected samples was significantly improved compared to the uncentrifuged samples: mean progressive motility was increased by 8% on the day of processing (P < 0.001) and by 13% after 24 h cold storage...
Pereira GR, Lorenzo PL, Carneiro GF, Ball BA, Gonçalves PB, Pegoraro LM, Bilodeau-Goeseels S, Kastelic JP, Casey PJ, Liu IK.The objective of this study was to test the hypothesis that equine growth hormone (eGH), in combination with insulin growth factor-I (IGF-I), influences positively in vitro nuclear and cytoplasmic maturation of equine oocytes. Cumulus-oocyte complexes were recovered from follicles that were < 25 mm in diameter, characterized by morphology and were allocated randomly as follow: (a) control (no additives); (b) 400 ng/ml eGH; (c) 200 ng/ml IGF-I; (d) eGH + IGF-I; and (e) eGH + IGF-I + 400 ng/ml anti-IGF-I antibody. Oocytes were matured for 30 h at 38.5°C in air with 5% CO2 and then stained wi...
de Grauw JC, van de Lest CH, van Weeren PR.Articular tissues are capable of producing a range of eicosanoid mediators, each of which has individual biological effects and may be affected by anti-inflammatory treatment. We set out to develop and evaluate a high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) approach for the simultaneous analysis of multiple eicosanoid lipid mediators in equine synovial fluid (SF), and to illustrate its use for investigation of the in vivo effects of non-steroidal anti-inflammatory drug (NSAID) treatment. Methods: Synovial fluid samples were obtained from normal joints of 6 adult...
Kim SK, Kim S, Dai G, Zhang Y, Ahn BC, O'Callaghan DJ.The equine herpesvirus 1 (EHV-1) negative regulatory IR2 protein (IR2P), an early 1,165-amino acid (aa) truncated form of the 1487-aa immediate-early protein (IEP), lacks the trans-activation domain essential for IEP activation functions but retains domains for binding DNA, TFIIB, and TBP and the nuclear localization signal. IR2P mutants of the N-terminal region which lack either DNA-binding activity or TFIIB-binding activity were unable to down-regulate EHV-1 promoters. In EHV-1-infected cells expressing full-length IR2P, transcription and protein expression of viral regulatory IE, early EICP...
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...
Vidale P, Piras FM, Nergadze SG, Bertoni L, Verini-Supplizi A, Adelson D, Guérin G, Giulotto E.We mapped six genes (EIF4G3, HSP90, RBBP6, IL8, TERT, and TERC) on the chromosomes of Equus caballus, Equus asinus, Equus grevyi, and Equus burchelli by fluorescence in situ hybridization. Our results add six type I markers to the cytogenetic map of these species and provide new information on the comparative genomics of the genus Equus.
Nitta K, Tsuge H, Iwamoto H.Unfolding profiles of two calcium-binding lysozymes, equine milk lysozyme and pigeon egg-white lysozyme, were obtained by circular dichroism and proton NMR measurements. Equine lysozyme unfolds through a stable molten globule intermediate. The molten globule of equine lysozyme was characterized as more ordered than that of bovine alpha-lactalbumin. On the other hand, pigeon lysozyme unfolds by a two-state mechanism and the intermediate could not be observed in guanidine or thermal unfolding, the same as with conventional non-calcium-binding lysozymes. Thus, from the point of view of the unfold...
Kellon EM, Gustafson KM.Hyperinsulinemia associated with equine metabolic syndrome and pituitary pars intermedia dysfunction is a risk factor for laminitis. Research in other species has shown elevated body iron levels as both a predictor and consequence of insulin resistance. In humans, this is known as dysmetabolic hyperferritinemia. To explore the relationship between equine hyperinsulinemia and body iron levels. We reviewed case histories and laboratory results from an open access database maintained by the Equine Cushing's and Insulin Resistance Group Inc. (ECIR). We identified 33 horses with confirmed hyperinsu...
King SS, Campbell AG, Dille EA, Roser JF, Murphy LL, Jones KL.Dopamine (DA) agonist and antagonist treatments can affect ovarian reproductive events in the mare. To support our theory that DA produces these effects by acting directly on the ovary, we analyzed equine ovarian tissues for the presence of dopamine receptor-1 (D1r) and dopamine receptor-2 (D2r) mRNA by reverse transcription polymerase chain reaction (RT-PCR) and D1r and D2r proteins by Western blot and immunohistochemistry (IHC). RT-PCR was performed on RNA isolated from ovarian cortex, medulla, granulosa/theca or corpus luteum (CL) tissues and from pituitary (D2r control) and renal artery (D...
Underwood C, Southwood LL, Walton RM, Johnson AL.To determine: (1) changes in blood ammonia, bile acid (BA), bilirubin, triglyceride, and glucose concentrations and liver enzyme activities in perioperative colic patients and (2) the association between these laboratory findings and short-term survival. Methods: Prospective observational clinical study. Methods: Thirty-two adult horses undergoing exploratory celiotomy for colic. Methods: None. Results: Blood samples were collected preoperatively and at 24-36 and 72-84 hours postoperatively and analyzed for blood ammonia, BA, bilirubin, triglyceride, and glucose concentrations and sorbitol deh...
Chirino-Trejo JM, Prescott JF.The antigens extracted from strains belonging to seven capsular serotypes of Rhodococcus equi, as well as from two wild strains isolated from pneumonic foals, were examined. Whole-cell antigens and soluble products present in broth culture supernatants were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, electroblotted onto nitrocellulose, and stained with serum from hyperimmunized rabbits or foals. Foal sera used included sera from pneumonic animals with known titer to equi factors; from animals bled monthly on a farm with enzootic pneumonia, and from animals bled mont...
Brown MP, West LA, Merritt KA, Plaas AH.To determine effects of aging on sulfation of chondroitin sulfate (CS) in articular cartilage and synovial fluid from normal equine middle carpal joints, and to determine whether CS compositional analysis can be used to assess alterations in proteoglycan turnover in degenerative cartilage obtained from horses with carpal osteochondral fractures. Methods: Carpal articular cartilage and synovial fluid from 44 cadavers with normal joints and from 16 Thoroughbred racehorses during routine carpal arthroscopic surgery. Methods: After papain/chondroitinase digestion of cartilage, CS disaccharides (un...
Monzón CM, Mancebo OA, Roux JP.In a total of 165 blood samples from horses in the Province of Formosa (Argentina), the diagnosis for equine trypanosomiasis (T. evansi) was made using Giemsa-stained smears (GSS), wet blood films (WBF), Strout's concentration method (SCM), haematocrit centrifuge technique (HCT), buffy coat method (BCM) and mouse inoculation of blood (MBI). Trypanosoma evansi was demonstrated in 52 samples. Mouse inoculation gave a sensitivity of 88.2%; HCT 71.1%; BCM 63.4%; WBF 53.8%; SCM 46.1% and GSS 45.6%. No single method alone was totally effective. The haematocrit centrifuge technique, mouse inoculation...
Ohnuma K, Yokoo M, Ito K, Nambo Y, Miyake YI, Komatsu M, Takahashi J.Early pregnancy factor (EPF) is an immunosuppressive protein detected in the early pregnancy serum. We have already reported that we developed the rosette inhibition test for mare EPF and detected EPF in thoroughbreds. The aim of this study was to determine whether or not our method could be used clinically. Methods: The rosette inhibition test for equine EPF was carried out on serum from six nonpregnant and six pregnant Shetland ponies, a female and a male Chinese pony, and four nonpregnant and 13 pregnant thoroughbred mares. In the thoroughbreds sera were collected during the pregnancy perio...
Pellegrini A, Waiblinger S, Von Fellenberg R.Lysozyme from equine neutrophil granulocytes was isolated in a pure form by fast performance liquid chromatography, i.e. ion-exchange chromatography and reversed-phase chromatography. The lysozyme lysed Micrococcus luteus, Bacillus subtilis and Staphylococcus lentus and was also bactericidal against the Gram-negative bacteria Escherichia coli, Klebsiella pneumoniae, Bordetella bronchiseptica, and Serratia marcescens. Staphylococcus aureus and Staphylococcus epidermidis were not lysed. The lysozyme was only very slightly bactericidal for S. epidermidis and S. aureus. Equine neutrophil lysozyme ...
Van den Hoogen BM, van de Lest CH, van Weeren PR, Lafeber FP, Lopes-Cardozo M, van Golde LM, Barneveld A.The purpose of this study was to determine whether changes in the synovial fluid (SF) induced by in vivo loading can induce an alteration in the metabolic activity of chondrocytes in vitro. Therefore, SF was collected from ponies after a period of box rest and after they had exercise for a week. Normal, unloaded articular cartilage explants were cultured in 20% solutions of these SFs for 4 days and chondrocyte activity was determined by glycosaminoglycan (GAG) turnover. In explants cultured in post-exercise SF, GAG synthesis was enhanced and GAG release was diminished when compared to cultures...
Torfs SC, Maes AA, Delesalle CJ, Deprez P, Croubels SM.Serotonin is regularly measured in equine platelet-poor plasma in research settings. However, reported reference values vary between studies, partially because plasma serotonin concentrations are very low and a reliable and affordable detection method is lacking. A simple, rapid, and sensitive method for serotonin determination in equine platelet-poor plasma using liquid chromatography--tandem mass spectrometry (LC-MS/MS) was developed and validated. Results of a commercially available enzyme-linked immunosorbent assay (ELISA) were compared to the LC-MS/MS results, in order to validate a test ...
Choi YH, Love LB, Westhusin ME, Hinrichs K.Early development of embryos produced by transfer of equine nuclei to bovine cytoplasts is superior to that of intraspecies equine nuclear transfer embryos. This may be related to differences in chromatin remodeling or efficiency of activation between the two oocyte types. The pattern of donor nucleus remodeling was examined in equine-equine and equine-bovine reconstructed oocytes. Chromosome condensation occurred in equine cytoplasts by 2 h but was not seen in bovine cytoplasts until 4 h. We investigated the effect of activation of equine-equine reconstructed oocytes at <30 min or at 2 h a...
Arar S, Chan KH, Quinn BA, Waelchli RO, Hayes MA, Betteridge KJ, Monteiro MA.During the second and third weeks of pregnancy, the equine conceptus expands rapidly while it is enclosed within a glycan capsule. Around day 16 of gestation, the conceptus loses its mobility in the uterus by a process termed 'fixation', coinciding with various changes in the capsule. Here, we compared the structure of the carbohydrate moieties expressed by the capsule during pre- and post-fixation periods. The glycan structures were studied by chemical analyses in combination with mass spectrometry. Capsule material from conceptuses collected before fixation (days 13-16) was observed to carry...
Nanayakkara SN, Rahnama S, Harris PA, Anderson ST, de Laat MA, Bailey S, Sillence MN.Although it is well established that equine laminitis can be triggered by extreme hyperinsulinemia, the mechanism of insulin action is not known. High concentrations of insulin lead to separation of the weight-bearing apparatus from the hoof wall and are associated with an increased cycle of cell death and proliferation in the lamellae. Gene expression and immunohistochemistry studies have indicated that the lamellae are sparsely populated with insulin receptors, whereas IGF-1 receptors (IGF-1R) are abundant, suggesting that the action of insulin may be mediated by insulin binding to the IGF-1...
Baker CB, Adams MH, McDowell KJ.Horse conceptuses were collected on Days 13, 15, 20 and 25 after ovulation. Whole conceptuses (Days 13 and 15) or extra-embryonic membranes (Days 20 and 25) were homogenized and poly-adenylated RNA (poly A RNA) was isolated by binding to oligo (dT)-cellulose. Poly A RNA (1 microgram/well) was separated by size on a denaturing 1% agarose gel and blotted onto nitrocellulose filters (northern blotting). DNA probes were prepared from plasmids containing equine alpha 1, omega 1 and omega 2 interferons and human beta actin. The presence of messenger RNA (mRNA) was detected by specific hybridization ...
Tigre DM, Brandão CF, de Paula FL, Chinalia FA, Campos GS, Sardi SI.Equine infectious anemia is an important infectious disease that affects equids worldwide. Control of the disease is currently based on detection of anti-p26 EIAV by Agar Gel Immunodiffusion (AGID). In this work, 62 animals were examined by AGID and nested-PCR using primers for the gag gene. Fifty-three samples (85.5%) were positive by nested-PCR, whereas only 33 samples (53%) were positive for AGID. Fifteen amplicons obtained by nested-PCR were sequenced and the aligned results subjected to phylogenetic analysis. The analysis suggests that the Brazilian EIAV form a cluster with WSU5, EIAVUK a...
Gargouri Y, Pieroni G, Rivière C, Sugihara A, Sarda L, Verger R.We report further investigations on protein inhibition of pancreatic and microbial lipases carried out with the monolayer technique. When beta-lactoglobulin A, melittin, serum albumin, myoglobin, and a protein inhibiting lipase from soybean were preincubated with a dicaprin film at a surface pressure of 35 dynes/cm, no activity was detected with horse pancreatic or Rhizopus delemar lipases. By contrast, Rhizopus arrhizus and Geotrichum candidum lipase activities were not impaired under the same conditions. Experiments using mixed lipid-protein film transfer clearly show that the inhibition of ...
Sabeur K, Ball BA.To identify the generation of the superoxide anion by equine spermatozoa. Methods: Multiple ejaculates collected from 3 Thoroughbred stallions. Methods: Induced superoxide production by reduced nicotinamide adenine dinucleotides (NAD[P]H; ie, reduced nicotinamide adenine dinucleotide [NADH] and reduced nicotinamide adenine dinucleotide phosphate [NADPH]) was measured by use of a nitroblue tetrazolium (NBT) reduction assay on whole spermatozoa and a cytochrome c reduction assay on isolated membrane fractions of spermatozoa. Localization of superoxide generation was determined by use of NBT cyto...
Lu G, Fu C, Huang J, Xu H, Wu P, Ping X, Li S.Equine pegivirus (EPgV) is a newly discovered equine virus, which is taxonomically classified in the Pegivirus genus of the Flaviviridae family. Until now, only the complete genome sequence of the first reported EPgV strain, from the USA (strain name: C0035) is available on online databases. Considering this, horse serum samples were collected from horses in China and screened for EPgV RNA by RT-PCR. One EPgV strain, LW/2016, was obtained and its near-complete genome sequence was acquired by standard PCR. Further analysis of its nucleotide sequence indicates LW/2016 is genetically divergent fr...
Cuniberti B, Odore R, Barbero R, Cagnardi P, Badino P, Girardi C, Re G.Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenases (COX), and the inhibition of COX-2 rather than COX-1 can limit the onset of NSAID-related adverse effects. The pharmacodynamic properties of eltenac, naproxen, tepoxalin, SC-560 and NS 398 in healthy horses were investigated using an in vitro whole blood assay. To predict COX selectivity in clinical use, eltenac and naproxen were also studied ex vivo after intravenous administration. SC-560 acted as a selective COX-1 inhibitor, tepoxalin as a dual inhibitor with potent activity against COX-1, and NS 398 as a preferential CO...
Ishitani K, Listowsky I.Horse spleen ferritin was fractionated into its constituent isoferritins by isoelectric focusing. Separated isoferritins were stable and showed no tendency to redistribute when re-examined by analytical gel focusing. All of the isoferritins were immunologically indistinguishable when tested with antibodies raised against unfractionated horse spleen ferritin. The separated isoferritins also had similar conformations as determined by circular dichroism. Iron distribution studies, however, revealed a wide disparity among the isoferritins. The most acidic components had the lowest iron content but...
Read AJ, Arzey KE, Finlaison DS, Gu X, Davis RJ, Ritchie L, Kirkland PD.An outbreak of equine influenza (EI) occurred in Australia in 2007. During the laboratory support for this outbreak, real-time reverse transcriptase polymerase chain reaction (qRT-PCR) assays and a blocking enzyme linked immunosorbent assay (bELISA) were used as testing methods to detect infection with the virus. The qRT-PCR and bELISA tests had not been used for EI diagnosis before, so it was not known how soon after infection these tests would yield positive results, or for how long these results would remain positive. To answer these questions, nasal swabs and blood samples were collected d...
Pirrone A, Antonelli C, Mariella J, Castagnetti C.The aim of this study was to verify if changes in blood glucose, creatinine, urea, and fibrinogen concentrations evaluated at birth reflect gross placenta abnormalities, and are useful to identify foals that suffered from placental dysfunction. A total of 92 mares were included in the present study: 68 delivered healthy foals and they were included in group 1; 24 delivered sick foals and they were included in group 2. In group 2, foals' clinical diagnoses included perinatal asphyxia syndrome (PAS; n = 20) and prematurity and/or dysmaturity (n = 4). The proportion of sick foals was greater when...
Bonilla-Gutiérrez AF, Castillo-Franz C, López C, Álvarez ME, Giraldo CE, Carmona JU.The aim of this study was to evaluate the release of pro- and anti-inflammatory as well as anabolic mediators stimulated by a leukocyte-reduced platelet-rich gel supernatant (Lr-PRGS) and a leukocyte-reduced plasma supernatant (Lr-PL) at two concentrations (25 and 50%) on normal equine suspensory ligament explants (SLEs) and tendon explants (TEs). SLEs and TEs from six horses were independently incubated for 48 h with Lr-PRGS and Lr-PL at concentrations of 25 and 50%, respectively. Samples were collected from the incubated tissues at 1 h and 48 h, which were employed for ELISA determinat...
Sánchez-Calabuig MJ, Fernández-González R, Hamdi M, Smits K, López-Cardona AP, Serres C, Macías-García B, Gutiérrez-Adán A.Equine embryos exhibit an unusual pattern of glucose tolerance in vitro and are currently cultured in hyperglycaemic conditions. Objective: Our main objective was to analyse the effect of different glucose concentrations on in vitro-produced equine embryo development and quality. Methods: Experiments comparing in vitro and in vivo produced embryos. Methods: Oocytes (n = 641) were collected from post-mortem ovaries, matured in vitro and fertilised by intracytoplasmic sperm injection (ICSI). Embryo culture was divided from Day 0 to Day 4 and from Day 4 to Day 9 in three groups: 5-10 (5 and 10...
Huang L, Zhu W, Saunders CP, Macleod JN, Zhou M, Stromberg AJ, Bathke AC.Identification of biomarkers among thousands of genes arrayed for disease classification has been the subject of considerable research in recent years. These studies have focused on disease classification, comparing experimental groups of effected to normal patients. Related experiments can be done to identify tissue-restricted biomarkers, genes with a high level of expression in one tissue compared to other tissue types in the body. Results: In this study, cartilage was compared with ten other body tissues using a two color array experimental design. Thirty-seven probe sets were identified as...
Hyldahl Laursen S, Andersen PH, Kjelgaard-Hansen M, Wiinberg B.There is a paucity of information about the analytical performance of thromboelastography (TEG) in horses, specifically concerning components of variation among different analytical methods. Such data may be obtained by nested analysis of repetitive standardized sampling of healthy individuals. Objective: The objectives were (1) to assess the relative susceptibility to sources of preanalytical variation in a highly standardized setting, (2) to directly compare and evaluate the observed analytical variation, and (3) to assess the applicability of population-based reference intervals. Methods: B...
Anderson RS, Day NK, Good RA.Natural hemagglutinin activity against vertebrate erythrocytes is present in the hemolymph of the cockroach Blabarus craniifer. The hemagglutinin titer against rabbit erythrocytes is high, whereas sheep and horse red cells agglutinate weakly. Hemagglutinin activity was depressed by the complement inhibitor, cobra venom factor. Cockroach hemagglutinin is heat-labile; all activity is destroyed by heating at 56 C for 1 hr. A humoral factor similar to the complement component 3 proactivator is also present in cockroach hemolymph. The formation of the cobra venom factor-hemolymph "complex" is depen...
Abdulmawjood A, Lämmler CH.The 16S rRNA gene of 39 S. equi subsp. zooepidemicus strains and two S. equi subsp. equi strains was amplified by polymerase chain reaction and subsequently digested with the restriction enzyme Hinc II. A restriction profile with two fragments with sizes of 1250 bp and 200 bp could be observed for both S. equi subsp. equi strains and for 30 of the 39 S. equi subsp. zooepidemicus strains indicating a sequence variation within the V2 region of the 16S rRNA gene of the remaining nine S. equi subsp. zooepidemicus isolates. A segment of the 16S rRNA gene including the hypervariable V2 region of 11 ...
Junsiri W, Kamkong P, Chinkangsadarn T, Ouisuwan S, Taweethavonsawat P.Equine ocular setariasis is mainly caused by Setaria digitata, and the identification of this filarial nematode is based on morphology. However, morphological characterization alone is insufficient for the detection and differentiation of S. digitata from its congeners. In Thailand, the molecular detection of S. digitata is lacking and its genetic diversity is still unknown. This study aimed to phylogenetically characterize equine S. digitata from Thailand based on sequences derived from the mitochondrial cytochrome c oxidase subunit 1 (COI), the mitochondrial small subunit ribosomal DNA (12S ...
