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.
Yarnell K, Purcell RS, Walker SL.Adrenal activity can be assessed in the equine species by analysis of feces for corticosterone metabolites. During a potentially aversive situation, corticotrophin releasing hormone (CRH) is released from the hypothalamus in the brain. This stimulates the release of adrenocorticotrophic hormone (ACTH) from the pituitary gland, which in turn stimulates release of glucocorticoids from the adrenal gland. In horses the glucocorticoid corticosterone is responsible for several adaptations needed to support equine flight behaviour and subsequent removal from the aversive situation. Corticosterone met...
Andrews ES, Turell MJ.Emerging and re-emerging arboviruses continue to be a threat to global public health, and viral surveillance of mosquito populations is critical for mosquito control operations. Due to the tropical climate of many of the affected areas, it may be difficult to maintain a cold chain as the samples travel from collection sites to laboratories for testing. We determined how suboptimal holding temperatures affected the ability to detect viruses in pools of mosquitoes. Adult female Aedes albopictus and Ae. taeniorhynchus individuals were inoculated with chikungunya virus or Venezuelan equine encepha...
Bassarak B, Moser I, Menge C.A modified Baltz's in vitro cultivation system for the propagation of Trypanosoma equiperdum strain OVI was established to develop a replacement for the conventional production procedure of dourine diagnostic antigen in rats. To increase trypanosome yields we designed an optimized culture medium by addition of supplemental compounds. Trypanosomes were adapted to this medium by two succeeding cultivation steps which led to a substantial proliferation rate and an increased cell density tolerance, respectively. As a result, adapted parasites could be propagated to maximum cell densities of >2Ã...
Peglar MT, Nerad TA, Anderson OR.A new species of lobosean amoeba, Stenamoeba polymorpha n. sp., was isolated from the diarrheic stool of a domesticated horse in Great Falls Virginia, U.S. It shares characteristics with the five other described Stenamoeba species. However, electron microscopy revealed S. polymorpha has a substantially thickened cell surface lamina. Under light microscopy, the amoebae had a dynamic polymorphic appearance because hyaloplasm readily formed and resorbed subpseudopodia from any peripheral region of the cell. While in locomotion, the amoebae produced subpseudopodia that led and alternated the dire...
Peña FJ, Ortega Ferrusola C, MartÃn Muñoz P.Flow cytometry is currently recognized as a robust tool for the evaluation of sperm quality and function. However, within equine reproduction, this technique has not reached the sophistication of other areas of biology and medicine. In recent years, more sophisticated flow cytometers have been introduced in andrology laboratories, and the number of tests that can be potentially used in the evaluation of sperm physiology has increased accordingly. In this review, recent advances in the evaluation of stallion spermatozoa will be discussed. These new techniques in flow cytometry are able to simul...
Habibi G, Esmaeilnia K, Hablolvarid MH, Afshari A, Zamen M, Bozorgi S.Equine piroplasmosis (EP) is the cause of persistent tick-borne infection with no symptoms, but the most important problem of EP is due to the persistent carrier state. Carrier animals to Babesia (Theileria) equi (Laveran 1901) and B. caballi (Nuttall, 1910) infestation could be identified by extremely sensitive PCR-based method. The purpose of this study was to identify the causative agents of equine piroplasmosis based on molecular and microscopic assays in equids from Kurdistan Province, Iran. Methods: Thirty one horse and mule blood samples were used with history of living in Kurdistan Pro...
Kumar J, Chaudhury A, Yadav SC.Trypanosomosis (Surra) is an economically important disease caused by Trypanosoma evansi which is an extracellular parasite present in the plasma, tissues and other body fluids of a wide range of hosts including domesticated animals. Currently, serological reports are based on detection of antibodies by ELISA using whole cell lysate (WCL) antigen, which has a limitation of persistence of anti-trypanosomal antibodies after successful treatment of the disease. Moreover, it has some ethical issues also like requirement of mice for in vivo maintenance of parasite for preparing the antigen. Therefo...
Byrnes JF, Bush PJ.Forensic anthropologists are more often turning to nondestructive methods to assist with skeletal analyses, specifically for trace elemental analyses. Portable XRF (pXRF) instruments are versatile and are able to be used in diverse settings or for specimens of a shape and size that cannot be accommodated by laboratory-based instruments. Use of XRF requires knowledge of analysis parameters such as X-ray penetration and exit depth. Analysis depth was determined by examining pure elements through known thicknesses of equine bone slices. Correlation between the element's X-ray emission energy and ...
Wood PL, Scoggin K, Ball BA, Troedsson MH, Squires EL.Using a nontargeted lipidomics analysis of equine sperm and seminal plasma, we were able to characterize a diverse array of individual lipids including ethanolamine and choline ether lipids and seminolipids essential to membrane raft function. We also detected, for the first time in sperm, the presence of (O-acyl)-ω-hydroxy-fatty acids (OAHFA) with up to 52 carbon chain lengths, which were localized to the head and not the tail of sperm. The only previous identification of OAHFAs has been in meibomian glands and their sebaceous secretions. The identities of these lipid amphiphiles were valida...
Takesue M, Osaka Y, Muranaka M, Katayama Y, Ikadai H.In the present study, the serum and cerebrospinal fluid of horses diagnosed with Setaria digitata cerebrospinal filariasis were analyzed by western blot. The results revealed S. digitata protein bands measuring 65, 34, 22, and 18 kDa in molecular weight. In particular, the 18 kDa band is a possible candidate for clinical immunodiagnosis on the basis of western blot findings.
Jarvis JA, Franke MA, Davis AD.An examination using the routine rabies direct fluorescent antibody test was performed on rabies or Eastern equine encephalitis positive mammalian brain tissue to assess inactivation of the virus. Neither virus was inactivated with acetone fixation nor the routine test, thus laboratory employees should treat all samples as rabies and when appropriate Eastern equine encephalitis positive throughout the whole procedure.
Barrier Battut I, Kempfer A, Becker J, Lebailly L, Camugli S, Chevrier L.Several laboratories routinely use flow cytometry to evaluate stallion semen quality. However, objective and practical tools for the on-field interpretation of data concerning fertilizing potential are scarce. A panel of nine tests, evaluating a large number of compartments or functions of the spermatozoa: motility, morphology, viability, mitochondrial activity, oxidation level, acrosome integrity, DNA integrity, "organization" of the plasma membrane, and hypoosmotic resistance, was applied to a population of 43 stallions, 33 of which showing widely differing fertilities (19%-84% pregnancy rat...
Lee B, Kang HY, Lee DO, Ahn C, Jeung EB.Claudins, which are known as transmembrane proteins play an essential role in tight junctions (TJs) to form physical barriers and regulate paracellular transportation. To understand equine diseases, it is helpful to measure the tissue-specific expression of TJs in horses. Major equine diseases such as colic and West Nile cause damage to TJs. In this study, the expression level and distribution of claudin-1, -2, -4, and -5 in eight tissues were assessed by Western blotting and immunohistochemistry methods. Claudin-1 was primarily identified in the lung, duodenum, and uterus, claudin-2 was evenl...
Lee KH, Park C, Bang H, Ahn M, Moon C, Kim S, Shin T.The olfactory mucosae of the horse were examined by using histology and lectin histochemistry to characterize the carbohydrate sugar residues therein. Histological findings revealed that olfactory epithelium (OE) consisted of both olfactory marker protein (OMP)- and protein gene product (PGP) 9.5-positive receptor cells, supporting cells and basal cells with intervening secretory ducts from Bowman's glands. Mucus histochemistry showed that Bowman's gland acini contain periodic acid-Schiff (PAS) reagent-positive neutral mucins and alcian blue pH 2.5-positive mucosubstances. Lectin histochemistr...
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...
Dunbar LK, Mielnicki KA, Dembek KA, Toribio RE, Burns TA.Several tests have been evaluated in horses for quantifying insulin dysregulation to support a diagnosis of equine metabolic syndrome. Comparing the performance of these tests in the same horses will provide clarification of their accuracy in the diagnosis of equine insulin dysregulation. Objective: The aim of this study was to evaluate the agreement between basal serum insulin concentrations (BIC), the oral sugar test (OST), the combined glucose-insulin test (CGIT), and the frequently sampled insulin-modified intravenous glucose tolerance test (FSIGTT). Methods: Twelve healthy, light-breed ho...
Erol E, Jackson C, Horohov D, Locke S, Smith J, Carter C.Determination of fetal serum amyloid A (SAA) concentrations in aborted fetuses can provide valuable information regarding the infectious and/or inflammatory process of abortion in horses. To investigate the relationship between fetal SAA levels and the infectious/inflammatory disease process in feto-placental tissues, a SAA ELISA was used to test heart serum samples of 89 equine fetuses that were necropsied and diagnosed in the following groups: a multiorgan disease process diagnosed with an identified microorganism (14 cases, group 1); only placentitis diagnosed with an identified microorgan...
Kwok WH, Choi TL, So PK, Yao ZP, Wan TS.Xenon can activate the hypoxia-inducible factors (HIFs). As such, it has been allegedly used in human sports for increasing erythropoiesis. Krypton, another noble gas with reported narcosis effect, can also be expected to be a potential and less expensive erythropoiesis stimulating agent. This has raised concern about the misuse of noble gases as doping agents in equine sports. The aim of the present study is to establish a method for the simultaneous detection of xenon and krypton in equine plasma for the purpose of doping control. Xenon- or krypton-fortified equine plasma samples were prepar...
Espina M, Jülke H, Brehm W, Ribitsch I, Winter K, Delling U.Background. Mesenchymal stromal cells (MSCs) are increasingly used for clinical applications in equine patients. For MSC isolation and expansion, a laboratory step is mandatory, after which the cells are sent back to the attending veterinarian. Preserving the biological properties of MSCs during this transport is paramount. The goal of the study was to compare transport-related parameters (transport container, media, temperature, time, cell concentration) that potentially influence characteristics of culture expanded equine MSCs. Methods. The study was arranged in three parts comparing (I) fiv...
Wang Y, Mao Q, Chang H, Wu Y, Pan S, Li Y, Zhang Y.Integrins can function as receptors for foot-and-mouth disease virus (FMDV) in epithelium. Horses are believed to be insusceptible to this disease, but the mechanism of resistance remains unclear. To detect whether FMDV can use integrin to attach to equine epithelial, we compared the utilities of αvβ3 and αvβ6 between bovine and equine kidney epithelial cells (KECs). Equine KECs showed almost equal efficiency to those of bovine. Further, the integrin αv, β3, and β6 subunits from bovine and equine were cloned and vectors were transfected into SW480 cells and COS-1 cells alone or together...
Chindera K, Mahato M, Sharma AK, Horsley H, Kloc-Muniak K, Kamaruzzaman NF, Kumar S, McFarlane A, Stach J, Bentin T, Good L.To combat infection and antimicrobial resistance, it is helpful to elucidate drug mechanism(s) of action. Here we examined how the widely used antimicrobial polyhexamethylene biguanide (PHMB) kills bacteria selectively over host cells. Contrary to the accepted model of microbial membrane disruption by PHMB, we observed cell entry into a range of bacterial species, and treated bacteria displayed cell division arrest and chromosome condensation, suggesting DNA binding as an alternative antimicrobial mechanism. A DNA-level mechanism was confirmed by observations that PHMB formed nanoparticles whe...
McElhone RL, Meakin GE, French JC, Alexander T, Morgan RM.A study was designed to investigate the effects of external variables, including blood type, flooring surface, footwear tread depth and blood dryness, on the appearance of blood-based footwear marks, with particular reference to simulating a specific casework scenario. Results showed that footwear marks left in human blood tended to be of greater quality than those in equine blood, highlighting a potential issue in applying data generated with equine blood to human bloodstains in casework. Footwear tread effects were also dependent on blood type, but the type of flooring surface did not affect...
Hue ES, Fortier CI, Laurent AM, Quesnelle YF, Fortier GD, Legrand LJ, Pronost SL.The protocol describes a quantitative RT-PCR method for the detection and quantification of EHV-2 in equine respiratory fluids according to the NF U47-600 norm. After the development and first validation step, two distinct characterization steps were performed according to the AFNOR norm: (a) characterization of the qRT-PCR assay alone and (b) characterization of the whole analytical method. The validation of the whole analytical method included the portrayal of all steps between the extraction of nucleic acids and the final PCR analysis. Validation of the whole method is very important for vi...
Duan H, Gao JF, Su X, Fu X, Yue DM, Gao Y, Qiu JH, Wang CR.Triodontophorus spp. parasitizes the large intestine of equine, causing strongylid diseases. The present study assessed genetic variation in five gene regions within and between Triodontophorus brevicauda and Triodontophorus nipponicus from Heilongjiang Province and the Inner Mongolia Autonomous region. The five gene markers were three mitochondrial (mt) genes, cytochrome c oxidase subunit I (cox1), NADH dehydrogenase subunit 5 (nad5), cytochrome b (cytb); and two ribosomal RNA genes, the internal transcribed spacer 1 (ITS1) and the internal transcribed spacer 2 (ITS2). Partial (p) sequences o...
Crawley JA, Chapman SN, Lummaa V, Lynsdale CL.Parasite infection status, intensity and resistance have traditionally been quantified via flotation techniques, but the need for immediate analysis following defecation imposes limitations and has led to the use of several faecal storage techniques. However, their effect on nematode egg counts has not been systematically validated and is often generalised across taxa despite evidence of differences between species. Here, we take the domestic horse Equus ferus caballus as a model to examine the impact of commonly used storage techniques on egg recovery: 1) high and low concentrations of ethano...
Van de Water E, Oosterlinck M, Duchateau L, Pille F.The purpose of this study was to determine whether the scil Vet abc Plus(+) (SCIL Animal Care Company, Altorf, France), an impedance hematology analyzer, can accurately quantify and differentiate nucleated blood cells (NBCs) in equine synovial fluid. Synovial fluid samples (n=242) in different stages of experimentally induced inflammation were analyzed with and without hyaluronidase pretreatment and compared to manual hemocytometer counts and smear reviews. No significant effect of hyaluronidase pretreatment was observed. Total nucleated cell counts of the scil Vet abc Plus(+) were significant...
Hansson A, Knych H, Stanley S, Thevis M, Bondesson U, Hedeland M.Selective androgen receptor modulators (SARMs) are prohibited in sports due to their performance enhancing ability. It is important to investigate the metabolism to determine appropriate targets for doping control. This is the first study where the equine metabolites of SARMs S1, S4 (Andarine) and S22 (Ostarine) have been studied in plasma. Methods: Each SARM was administered to three horses as an intravenous bolus dose and plasma samples were collected. The samples were pretreated with protein precipitation using cold acetonitrile before separation by liquid chromatography. The mass spectrome...
de Carvalho Filho MB, Ramos RM, Fonseca AA, de Lima Orzil L, Sales ML, de Assis Santana VL, de Souza MM, Dos Reis Machado E, Filho PR, Leite RC....The allergic test of mallein is one of the most frequently used tests, together with the Complement Fixation Test (CFT), for the diagnosis of glanders in endemic areas. Mallein, a purified protein derivative (PPD), is produced similarly to PPD tuberculin and the end product is a primarily proteic antigen, which is only poorly purified. The immuno-allergic activity of mallein is believed to be due to a high molecular weight group of proteins present in the antigen. To improve the quality of the antigen, in terms of sensitivity and specificity, a new method of mallein production was developed, i...
Radin MJ, Eubank MC, Weiser MG.Anemia was induced in three groups of horses by moderate or severe acute hemorrhage, or by acetyl phenylhydrazine-induced hemolysis (Groups I, II, and III, respectively). Serial hemograms were done on a multichannel automated blood cell counter with histogram capability. Changes in hematocrit, mean cell volume, erythrocyte number, red cell distribution width (RDW), and standard deviation of erythrocyte volume were examined over time. Significant increases in mean cell volume were first detectable by days 17, 20, and 14 and reached maximum by days 43, 41, and 29, in Groups I, II, and III, respe...
Townson DH, Pierson RA, Ginther OJ.Plasma progesterone concentrations in mares were determined in two experiments during the time that the luteal glands were detectable by transrectal ultrasonography. In both experiments, corpora lutea were classified into two types of morphologies based on their ultrasonic appearance: centrally nonechogenic luteal glands (fluid-filled) and uniformly echogenic luteal glands (non-fluid-filled). In Experiment 1, daily blood samples were taken from horse mares during August through October and May through July. There were no significant effects of season or luteal morphology on progesterone concen...
Bjöersdorff A, Bagert B, Massung RF, Gusa A, Eliasson I.We report the isolation and partial genetic characterization of two equine strains of granulocytic Ehrlichia of the genogroup Ehrlichia phagocytophila. Frozen whole-blood samples from two Swedish horses with laboratory-verified granulocytic ehrlichiosis were inoculated into HL-60 cell cultures. Granulocytic Ehrlichia was isolated and propagated from both horses. DNA extracts from the respective strains were amplified by PCR using primers directed towards the 16S rRNA gene, the groESL heat shock operon gene, and the ank gene. The amplified gene fragments were sequenced and compared to known seq...
Strauch S, Wichert B, Greef JM, Hillegeist D, Zeyner A, Liesegang A.The aim of this study was to improve an in vitro system in order to gather optimized information on the digestion of different forages in the horse's upper gastrointestinal tract. Therefore, foregut digestion of several forages was simulated in vitro (Part 1). The effect of different pH values on in vitro fructan degradation of two selected grasses (Part 2) was tested subsequently. Part 1: We hypothesized that our system produces representative results simulating digestive processes in the upper alimentary tract, but neglects microbial fermentation. In vitro digestion of six forages (grass ...
Desta B, Maldonado G, Reid H, Puschner B, Maxwell J, Agasan A, Humphreys L, Holt T.Just prior to an international polo event, 21 horses from one team exhibited clinical signs of central nervous system disturbance, hyperexcitability, sweating, ataxia, tachycardia, dyspnea, pyrexia, and rapid death. The suspected cause of this peracute onset of illness and death included intentional contamination of feed or iatrogenic administration of performance-enhancing drugs resulting in a severe adverse reaction. Six horses were submitted to the Bronson Animal Disease Diagnostic Laboratory for necropsy and toxicological examination. The clinical signs and sudden death, the similarity to ...
Aziz DM, Ahlswede L, Enbergs H.The assay of MTT reduction depends on the ability of metabolically active cells to reduce the tetrazolium salt (3[4,5-dimethylthiazol-2-y1]-2,5-diphenyltetrazolium bromide) to formazan. This study was conducted to examine and validate of a simple and less costly MTT test in determining equine sperm viability and compare the efficiency of this test with a flow cytometer. Fresh ejaculates from 11 stallions (warm blood) were included in this study. Semen was diluted to 100 million cells/ml in a Hepes 0.1% BSA. The rates of MTT reduction were measured in microtiter plates after incubation for 1 an...
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 ...
Annandale EJ, Valberg SJ, Mickelson JR, Seaquist ER.Equine polysaccharide storage myopathy (PSSM) is an inherited disorder characterized by the accumulation of glycogen and abnormal polysaccharide in muscle with normal glyco(geno)lytic enzyme activities. The purpose of this study was to evaluate in vivo insulin sensitivity and glucose excursion in PSSM using a euglycemic hyperinsulinemic clamp. In addition, the content of muscle glucose transporters (GLUT1 and GLUT4) and the insulin receptor was determined in muscle biopsies using Western blot analysis. The glycogen content was 1.8-fold higher, and isolated polysaccharide analyzed by iodine abs...
Pirosanto Y, Laseca N, Valera M, Molina A, Moreno-Millán M, Bugno-Poniewierska M, Ross P, Azor P, Demyda-Peyrás S.Chromosomal abnormalities are a common cause of infertility in horses. However, they are difficult to detect using automated methods. Here, we propose a simple methodology based on single nucleotide polymorphism (SNP)-array data that allows us to detect the main chromosomal abnormalities in horses in a single procedure. As proof of concept, we were able to detect chromosomal abnormalities in 33 out of 268 individuals, including monosomies, chimerisms, and male and female sex-reversions, by analyzing the raw signal intensity produced by an SNP array-based genotyping platform. We also demonstrat...
Ribeiro BI, Love LB, Choi YH, Hinrichs K.Use of assisted reproduction to obtain foals from valuable mares post-mortem typically necessitates holding of ovaries during shipment to a laboratory. The present study evaluated whether holding ovaries briefly at a warm ( approximately 30 degrees C) temperature improves meiotic and developmental competence of oocytes, as determined after maturation in vitro and intracytoplasmic sperm injection. Ovaries were packaged in pairs in insulated containers, and held either at 24 or 25-35 degrees C for 4h, followed by cooling. Ovaries in both treatments were held for either a short (mean, 7-7.4h) or ...
Brito LF, Greene LM, Kelleman A, Knobbe M, Turner R.The objective of this study was to determine the effects of method and clinician on stallion sperm morphology evaluation. Five clinicians evaluated 60 semen samples using wet-mount preparations with phase-contrast, eosin/nigrosin-stained semen smears, and Papanicolaou-stained semen smears. There were significant differences among methods for all sperm morphology categories and most intra-class correlation coefficients were only fair to moderate. The use of wet-mount preparations facilitated detection of acrosome defects, nuclear vacuoles, and cytoplasmic droplets when compared to stained smear...
Edwards LA, Donnelly CG, Reed SM, Valberg S, Chigerwe M, Johnson AL, Finno CJ.Currently, there is little information regarding the concentrations of phosphorylated neurofilament heavy protein (pNfH) in the serum and cerebrospinal fluid (CSF) of horses with neurodegenerative diseases. Specifically, pNfH concentrations have not yet been evaluated in horses with equine neuroaxonal dystrophy/equine degenerative myeloencephalopathy (eNAD/EDM). Objective: To determine pNfH concentrations using a commercial enzyme-linked immunosorbent assay (ELISA) in serum and CSF from control horses and horses with eNAD/EDM, cervical vertebral compressive myelopathy (CVCM) and Shivers. Metho...
Choi KS, Ko YJ, Nah JJ, Kim YJ, Kang SY, Yoon KJ, Joo YS.A rapid immunoassay for detecting and quantifying West Nile virus (WNV)-neutralizing antibodies in sera was developed as an alternative to the plaque reduction neutralization test (PRNT), the gold standard test for WNV. The assay is a competitive, enzyme-linked immunosorbent assay using neutralizing monoclonal antibody 5E8 (NT-ELISA). A cutoff percent inhibition (PI) value of 35% (mean PI plus 3 standard deviations), with a specificity of 99%, was established based on analysis of 246 serum samples from horses free of WNV. The NT-ELISA detected neutralizing antibodies in all sera collected 7 or...
O'Shea CM, Werre SR, Dahlgren LA.(1) To compare the performance of 4 platelet counting technologies in equine platelet concentrates and (2) to evaluate the ability of the Magellan platelet rich plasma (PRP) system to concentrate equine platelets. Methods: Experimental study to assess method agreement. Methods: Adult mixed breed horses (n = 32). Methods: Acid citrate dextrose-A anti-coagulated whole blood was collected and PRP produced using the Magellan system according to the manufacturer's instructions. Platelets were quantified using 4 counting methods: optical scatter (Advia 2120), impedance (CellDyn 3700), hand counting,...
Laroucau K, Saqib M, Martin B, Deshayes T, Bertin C, Wernery U, Joseph S, Singha H, Tripathi BN, Beck C.Burkholderia mallei is the etiologic agent of glanders, an infectious disease of solipeds, with renewed scientific interest due to its increasing incidence in different parts of the world. More rapid, sensitive and specific assays are required by laboratories for confirmatory testing of this disease. A microsphere-based immunoassay consisting of beads coated with B. mallei recombinant proteins (BimA, GroEL, Hcp1, and TssB) has been developed for the serological diagnosis of glanders. The proteins' performance was compared with the OIE reference complement fixation test (CFT) and an indirect en...
Ravaioli V, Raffini E, Tamburini M, Galletti G, Frasnelli M.Endometritis is a major cause of infertility and subfertility in the mare. Early diagnosis and identification of the pathogens involved in infectious endometritis are crucial to initiate correct treatments in time, in order to optimize fertility and reduce the risk of bacterial resistance development. In this retrospective study (from 2014 to 2018), 394 samples (uterine swabs and lavages) obtained from mares before breeding, regardless of clinical history of endometritis were analyzed. Our bacteriological procedure included the subculturing from the enrichment in Brain Heart Infusion Broth of ...
Vina-Rodriguez A, Eiden M, Keller M, Hinrichs W, Groschup MH. (VEEV) is an from the family that causes epizootic outbreaks in equids and humans in Central and South America. So far, most studies use conventional reverse transcriptase PCR assays for the detection of the different VEEV subtypes. Here we describe the development of a TaqMan quantitative real-time reverse transcriptase PCR assay for the specific detection and quantitation of all VEEV subtypes which uses in parallel a universal equine encephalitis virus control RNA carrying target sequences of the three equine encephalitis viruses. The control RNA was used to generate standard curves for t...
Gillespie J, Kalica A, Conner M, Schiff E, Barr M, Holmes D, Frey M.From 105 field cases of diarrhea in neonatal or young foals, rotavirus was detected by electron microscopy (EM) and/or by enzyme-linked immunosorbent assay (ELISA) in the feces of 65 foals on 16 different premises. ELISA was performed with Rotazyme test kits developed by Abbot and Company for the detection of rotaviruses. Twenty-four field isolates from the feces of diarrheic foals with equine rotavirus infection as ascertained by EM were placed in MA-104 cell cultures after pretreatment of the viral suspension with 10 micrograms ml-1 of trypsin and incorporation of 0.5 micrograms ml-1 or 1 mi...
Paillot R, Kydd JH, MacRae S, Minke JM, Hannant D, Daly JM.Equine influenza virus (EIV) is a leading cause of respiratory disease in horses. Equine influenza infection induces a long-term immunity to re-infection. Recent strategies of vaccination aim to mimic this immunity by stimulating both antibody and cellular immune responses. Cell-mediated immunity (CMI) to influenza is well defined in man, but little has been done to characterise the responses in the horse. Additionally, the development of reliable assays for the measurement of equine CMI has lagged behind serological methods and vaccine development. In this study, two methods of measuring EIV-...
Desantis S, Accogli G, Albrizio M, Rossi R, Cremonesi F, Lange Consiglio A.Equine amniotic mesenchymal cells (eAMCs) are involved in many mechanisms in tissue regenerative processes. Their secreted vesicles are important effectors in a wide array of biological processes, and contribute to in vivo healing of equine tendon lesions and endometrial inflammation. Glycoconjugates are involved in cellular recognition and in the efficient uptake of extracellular vesicles (EVs) by recipient cells. In this study, we evaluated the surface glycosylation pattern of eAMCs and their EVs from the eAMCs released in conditioned medium. We used a microarray procedure in which eAMCs and...
Tennent-Brown BS, Wilkins PA, Lindborg S, Russell G, Boston RC.Blood lactate concentration [LAC] is considered a useful indicator of disease severity in horses. Agreement of point-of-care (POC) lactate monitors with laboratory standards has not been established for clinically abnormal horses. Objective: It was hypothesized that results from a POC lactate monitor would be in agreement with a laboratory-based measurement of [LAC]. Methods: The study included adult horses presented for emergency evaluation. Methods: A prospective observational study was performed. [LAC] was measured with whole blood (AWB) and plasma (APL) by means of a POC monitor (Accutrend...
Katila T.The article reviews methods used for in vitro evaluation of sperm, with particular emphasis on frozen-thawed stallion sperm. The techniques, limitations of the methods and correlations with fertility results are discussed. Very few studies have tried to find correlation between fertility of frozen stallion semen and laboratory tests. It is difficult and expensive to inseminate an adequate number of mares to achieve statistically significant differences. Significant, but low correlations have been demonstrated between the foaling rate and subjective motility of sperm incubated for 2 h and 4 h a...
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...
Tamzali Y, Guelfi JF, Braun JP.Plasma fibrinogen is widely used in horse practice as an unspecific positive marker of inflammatory diseases; it is also lowered in disseminated intravascular coagulation. Three fibrinogen measurement methods--Millar's heat-denaturation in a microhaematocrit tube, automated reader for heat-denaturation, and chronometric measurement of clot formation after addition of excess thrombin-were compared by means of Passing-Bablock's regression and Bland-Altman difference plots, in blood plasma of 30 clinically healthy and 57 diseased horses. Correlations between the three techniques were excellent (r...
Kittelberger R, McFadden AM, Hannah MJ, Jenner J, Bueno R, Wait J, Kirkland PD, Delbridge G, Heine HG, Selleck PW, Pearce TW, Pigott CJ, O'Keefe JS.New Zealand is free from equine influenza and has never experienced an incursion in its horse population. As part of New Zealand's preparedness to an incursion of an exotic animal disease, it was considered necessary to select the most accurate test for equine influenza (EI) from the array of those available. Four readily available blocking/competitive enzyme-linked immunosorbent assays (ELISA), originally developed and marketed for the detection of antibodies against the avian influenza virus, were evaluated using serum samples from New Zealand non-infected, non-vaccinated horses (n=365), and...
Theurillat R, Knobloch M, Schmitz A, Lassahn PG, Mevissen M, Thormann W.CE with multiple isomer sulfated beta-CD as the chiral selector was assessed for the simultaneous analysis of the enantiomers of ketamine and metabolites in extracts of equine plasma and urine. Different lots of the commercial chiral selector provided significant changes in enantiomeric ketamine separability, a fact that can be related to the manufacturing variability. A mixture of two lots was found to provide high-resolution separations and interference-free detection of the enantiomers of ketamine, norketamine, dehydronorketamine, and an incompletely identified hydroxylated metabolite of no...
Braune S, Walter M, Schulze F, Lendlein A, Jung F.For in vitro studies assessing the interaction of platelets with implant materials, common and standardized protocols for the preparation of platelet rich plasma (PRP) are lacking, which may lead to non-matching results due to the diversity of applied protocols. Particularly, the aging of platelets during prolonged preparation and storage times is discussed to lead to an underestimation of the material thrombogenicity. Here, we study the influence of whole blood- and PRP-storage times on changes in platelet morphology and function. Blood from apparently healthy subjects was collected according...
Forrer R, Gautschi K, Lutz H.It was the goal of this paper to establish total selenium reference values for Switzerland in different animal species and in humans. To this purpose, a flameless atomic absorption method with deuterium background compensation utilizing a graphite furnace atomization system with a pyrolytic platform inside and palladium solution as matrix modifier was developed for the measurement of selenium in serum. The method was characterized by rapid performability, small sample requirement, acceptable detection limit (0.04 mumol/L) and precision and a linear range of measurement up to 4 mumol/L. The met...
Papa FO, FelÃcio GB, Melo-Oña CM, Alvarenga MA, De Vita B, Trinque C, Puoli-Filho JN, Dell'Aqua JA.The objective of this study was to determine whether replacing the egg yolk with soybean lecithin in the Botu-Crio® cryodiluent would maintain the fertility of cryopreserved stallion sperm. Two experiments were performed to evaluate cell freezability. In experiment 1, sperm from 15 stallions were frozen in Botu-Crio® (BC) or Botu-Crio® which contained 45g/L soybean lecithin (BCLS45) in place of the egg yolk. In experiment 2, we compared different concentrations of soybean lecithin: 0, 10.0, 12.5, 15.0, 17.5 and 20.0g/L (BC, BCLS10, BCLS12.5, BCLS17.5 and BCLS20, respectively). In experiment...
