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.
Faulk C.Sequencing a genome by students has now become practical as we demonstrated with our recent publication of the Przewalski's horse (Equus ferus przewalskii) genome. In this review, I describe my experience teaching genome assembly in the classroom. In my course, students sequenced, assembled, and published a high-quality genome for Przewalski's horse using Oxford Nanopore long-read sequencing with only $4000 of materials. Along with the genome, we assembled the mitochondrial genome, sequence variants, predicted gene annotations, and DNA methylation levels. Our genome statistics far exceeded the...
Littiere TO, Costa GB, Sales NAA, Carvalho JRG, Rodrigues IDM, Ramos GV, Ferraz GC.In humans, there is an ongoing academic discussion about measuring plasma lactate concentration ([La]) during an incremental exercise test (IET) to establish thresholds that can predict the maximal lactate steady state (MLSS). Training studies on horses have utilized the onset of [La] (OBLA), also known as V, the velocity at which the [La] of 4 mM is reached. This study assesses the effectiveness of four methods for estimating the MLSS in teaching horses. Ten teaching horses underwent a single IET to determine four exercise intensity thresholds for predicting MLSS and performed several constan...
Alim S, Cheppali SK, Pawar SS, Swamy MJ.Fibronectin type-II (FnII) proteins are major constituents in the seminal plasma of many mammals and play a crucial role in sperm capacitation. Additionally, the seminal FnII proteins from bull and horse exhibit chaperone-like activity (CLA), by acting as small heat shock proteins (shsps). The present work demonstrates that the major FnII protein of donkey seminal plasma, DSP-1 exhibits CLA with broad specificity and protects various client proteins such as alcohol dehydrogenase, lactate dehydrogenase and enolase against thermal and oxidative stress. Binding of phosphorylcholine (PrC) - the he...
Byambaragchaa M, Park SH, Park MH, Kang MH, Min KS.Equine chorionic gonadotropin (eCG) hormone, comprising highly glycosylated α- and β-subunits, elicits responses similar to follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in non-equid species. This study aimed to establish a mass production of recombinant eCG (rec-eCG) using CHO DG44 cells. Single-chain rec-eCG β/α was expressed in CHO DG44 cells. FSH- and LH-like activities were evaluated in CHO-K1 and HEK 293 cells expressing the equine LH/CG receptor (eLH/CGR), rat LH/CGR (rLH/CGR), and rFSHR. pERK1/2 activation and β-arrestin 2 recruitment were assessed in PathHunter ...
Simões BP, Cerri FM, Takahira RK, Borges AS, Oliveira-Filho JP, Amorim RM.Chlorfenapyr is a pesticide that interferes with mitochondrial function, leading to cell death and mortality. Although poisoning is primarily documented in humans and dogs, it can result in severe clinical signs, including sweating, respiratory distress, and neurological dysfunction. This report describes the clinical and laboratory findings of a horse with suspected accidental intoxication with chlorfenapyr. A 4-year-old Quarter Horse mare was referred to the veterinary hospital with signs of excessive sweating, hyperthermia, tachypnea, muscle fasciculation, and fearful facial expression. The...
Johnston TJ, Stewart AJ, Dryburgh EL, Bertin FR.Pituitary pars intermedia dysfunction (PPID) is diagnosed by measuring the ACTH concentrations. Due to the reported instability of ACTH, it is recommended to transfer centrifuged plasma into cryovials; however, in practice, cryovials are infrequently used, and serum (red-top) tubes are used instead. This study investigated whether this procedure affects ACTH concentrations and the diagnosis of PPID. Unassigned: This was a cohort study. Blood was collected into EDTA tubes from 9 horses with PPID and 7 controls. After centrifugation, plasma was either aliquoted into a cryovial or into a serum tu...
Facile V, Magliocca M, Dini FM, Imposimato I, Mariella J, Freccero F, Urbani L, Rinnovati R, Sel E, Gallina L, Castagnetti C, Galuppi R, Battilani M....Equine piroplasmosis is a tick-borne disease caused by and species. Despite its presence in Europe, no laboratory testing is required for animal movement, even though some countries remain free of this disease. Differentiating between species and genotypes is crucial to determine the most effective treatment, as dosage, active compounds, and duration vary. However, diagnosis is often challenging due to genetic variability and the limited sensitivity of molecular methods. The aims of this study were to compare the performances of different molecular diagnostic tests to identify the most effec...
Scheike AS, Plomp S, Fugazzola MC, Meurot C, Berenbaum F, van Weeren PR, Tryfonidou MA, von Hegedus JH.This study investigates the anti-inflammatory properties of liraglutide, a glucagon-like peptide 1 receptor agonists, in equine in vitro models and in an in vivo acute synovitis model in Shetland ponies. The anti-inflammatory effect of liraglutide was assessed by measuring concentrations of inflammatory biomarker C-C Motif Chemokine Ligand 2 (CCL2) in culture media of equine whole blood, peripheral blood mononuclear cells (PBMCs), chondrocytes, and synoviocytes, with or without lipopolysaccharide (LPS) or interleukin-1β. In the in vivo experiment, acute synovitis was bilaterally induced with ...
Barot D, Usimaki A, Linardi RL, Arensberg CM, Ortved KF.Quantify the concentration of α2-macroglobulin (A2M), immunomodulatory cytokines, and TGF-β1 factors in 4 commercially available autologous blood-based products including conditioned A2M (CA2M; Alpha2EQ; Astaria Global), autologous protein solution (APS; Pro-Stride; Zoetis), platelet-rich plasma (PRP; Restigen; Zoetis), and autologous conditioned plasma (ACP; Arthrex ACP). We hypothesized that CA2M would have higher concentrations of A2M and lower concentrations of cytokines and growth factors compared to APS, PRP, and ACP. Unassigned: Blood was obtained from 6 healthy, adult horses and proc...
Barbazanges P, Couroucé A, Le Digarcher G, Cardwell JM, Schmitt E, Toquet MP, Lemonnier LC, Richard EA.Fungi are ubiquitous in horses' environment. Their contribution to the pathophysiology of severe asthma (SA) is acknowledged, while controversies remain for mild-moderate asthma (MA). Objective: We hypothesised that fungi are a risk factor for asthma. Our objective was to compare different combinations of analytical methods (cytology, culture) and sampling sites (tracheal wash (TW), bronchoalveolar lavage fluid (BALF)) in relation to clinical status (control, MA, SA). Methods: Prospective cross-sectional study. Methods: The study population included asymptomatic racing horses in the field and ...
Finno CJ.In the field of equine muscle disorders, many conditions have a genetic basis. Therefore, genetic testing is an important part of the diagnostic evaluation. Validated genetic tests are currently available for 5 equine muscle disorders: hyperkalemic periodic paralysis, malignant hyperthermia, glycogen branching enzyme disease, type 1 polysaccharide storage myopathy, and myosin heavy chain myopathy. These diseases should be tested for in the appropriate breeds with clinical signs of disease or as part of breeding management. Genetic testing in veterinary medicine is not regulated, and therefore,...
Pusterla N, Barnum S, Lawton K, Craig B, James K.Equid alphaherpesvirus 4 (EqAHV4; Orthoherpesviridae, Varicellovirus equidalpha4; equine rhinopneumonitis virus) has seldom been associated with complications such as abortion and myeloencephalopathy, given the low tendency of this virus to induce viremia. We investigated the frequency of EqAHV4 viremia in horses with fever and respiratory signs. Case selection included all equids with EqAHV4 quantitative real-time PCR (qPCR)-positive nasal secretions (defined as EqAHV4 qPCR-positive cases) submitted to a diagnostic laboratory. Controls consisted of each case submitted before and after each Eq...
Halo M, Kirchner R, Tirpák F, Slanina T, Tokárová K, Kováčik A, Miškeje M, Greń A, Formicki G, Halo M, Madeddu R, Massányi P.The abundance of chemical elements in the blood of horses can indicate the physiological balance, health of animal as well as can be taken as an indicator of environmental pollution. The aim of this work was to analyse haematological, biochemical parameters, TOS, FRAP, SOD, Gpx, TAS and their correlations with concentrations of essential and risk elements in blood of horses stabled in two different locations: The National Stud Farm Topoľčianky (n = 11; 11 stallions, consisting of the breeds 6 Lipizzan, 3 Slovak warmblood, 2 Holsteiner) and Experimental Centre at Institute of Animal Husba...
Flood JA, Collins NM, Russell CM, Cuming RS, Carrick JB, Cudmore LA.Sepsis is a leading cause of morbidity and mortality in neonatal foals. Administration of appropriate antimicrobials and early aggressive supportive treatment is central to the efficacious treatment of neonatal sepsis and has proven to positively influence outcomes. The primary aim of our study was to evaluate microorganism results and antimicrobial sensitivities of neonatal foals with a positive blood culture submitted to our intensive care unit (ICU) during 2005-2022. The secondary aim was to compare the results to those of a similar previous study performed at the same ICU during 1999-2004....
Kublicka A, Lorek D, Mikołajczyk-Martinez A, Chodaczek G, Chwirot A, Bażanów B, Matczuk AK.The process of viral entry into host cells is crucial for the establishment of infection and the determination of viral pathogenicity. A comprehensive understanding of entry pathways is fundamental for the development of novel therapeutic strategies. Standard techniques for investigating viral entry include confocal microscopy and flow cytometry, both of which provide complementary qualitative and quantitative data. Imaging flow cytometry, which integrates the advantages of both methodologies, offers significant potential in virological studies. In this investigation, we employed imaging flow ...
Fegraeus K, Riihimäki M, Nordlund J, Akula S, Wernersson S, Raine A.Single-cell RNA sequencing (scRNA-seq) is a valuable tool for investigating cellular heterogeneity in diseases such as equine asthma (EA). This study evaluates the HIVE™ scRNA-seq method, a pico-well-based technology, for processing bronchoalveolar lavage (BAL) cells from horses with EA. The HIVE method offers practical advantages, including compatibility with both field and clinical settings, as well as a gentle workflow suited for handling sensitive cells. Our results show that the major cell types in equine BAL were successfully identified; however, the proportions of T cells and macropha...
Wang J, Ren W, Li Z, Ma S, Li L, Wang R, Zeng Y, Meng J, Yao X.This study aims to analyze the whole-genome DNA methylation differences in Yili horses before and after racing, with the goal of identifying differentially methylated genes associated with racing performance and exploring the epigenetic mechanisms underlying exercise in horses. Blood samples were collected from the jugular veins of the top 3 Yili horses in a 5000 m race, which included 25 competitors, both prior to and within 5 min after the race. Genomic DNA was extracted, followed by sequencing using Whole-Genome Bisulfite Sequencing (WGBS) to assess DNA methylation levels, differentially me...
The AnalystJanuary 23, 2025
doi: 10.1039/d4an01501k
Göktaş EF, Kabil E, Söylemez Yeşilçimen E, Dirikolu L.Various technical methodologies are required to accurately detect substances of different chemical and pharmacological properties in biological samples, which are increasing in number and variety daily. Therefore, laboratories where many samples and different factors are analyzed simultaneously need methods with easy sample preparation, short analysis times and low analysis costs. In this study, the objective was to scan substances susceptible to chemical degradation, amenable to analysis without hydrolysis, and exhibiting short-term stability by employing a straightforward, expeditious, and c...
Whitlock F, van Dijk J, Hodgkinson JE, Grewar JD, Newton JR.Anthelmintic resistance (AR) threatens effective equine parasite control. Quarterly data summaries from faecal worm egg count testing (FWECT) performed by UK laboratories have appeared in Equine Quarterly Disease Surveillance Reports (EQDSR) since 2007, but have not previously been assessed. Objective: To assess strongyle FWECT methods and thresholds used by UK laboratories. To investigate factors associated with quarterly laboratory FWECT positivity rates between 2007 and 2023. Methods: Laboratory surveys and analysis of laboratory summary data. Methods: Laboratories were surveyed in Q3 2018 ...
Palomino Lago E, Ross AKC, McClellan A, Guest DJ.Bone fractures are a significant problem in Thoroughbred racehorses. The risk of fracture is influenced by both genetic and environmental factors. To determine the biological processes that are affected in genetically susceptible horses, we utilised polygenic risk scoring to establish induced pluripotent stem cells (iPSCs) from horses at high and low genetic risk. RNA-sequencing on iPSC-derived osteoblasts revealed 112 genes that were significantly differentially expressed. Forty-three of these genes have known roles in bone, 27 are not yet annotated in the equine genome and 42 currently have ...
Nemoto M, Kawanishi N, Kambayashi Y, Bannai H, Yamanaka T, Tsujimura K.Equine influenza virus (EIV) can be transmitted by inhalation of aerosolized droplets, direct contact, and contaminated fomites. However, to our knowledge, there are no reports of the recovery of EIV from the air surrounding infected horses. Here, we evaluated whether EIV can be recovered from the air in the stalls of experimentally infected horses by using an air sampler. Furthermore, we examined whether rapid molecular test kits with reaction times of less than 30 min can detect EIV from air samples for potential field application. Two horses kept in individual stalls were experimentally i...
Hidalgo M.Sperm vitrification is an alternative freezing method, which includes high cooling rates and non-permeable cryoprotectants agents. The first attempt in equids was using the spheres technique by directly dropping small volumes of the sperm into liquid nitrogen. Later, vitrification was developed using 0.25 mL straws with outer covers, which resulted in similar progressive motility when compared to conventional freezing in donkeys (44.3 ± 15.0 % vs. 44.7 ± 18.2 %) or even higher in horses (48.2 ± 2.3 % vs. 37.3 ± 2.2 %). Subsequently, the vitrification of larger volumes of sperm in 0.5 mL st...
Königová A, Babják M, Kuzmina TA, Burcáková Ľ, Syrota Y, Várady M.The study aimed to conduct a survey on the occurrence of benzimidazole (BZ) resistance in strongyles by in vitro egg hatch test (EHT) and larval development test (LDT) and to identify the effective indicators of early resistance detection on horse farms with associated risk factors analysis appraisal. In total, 203 horses from 8 farms underwent the fecal sampling of which 77 horses were selected for in vitro testing. Simultaneously, 18 horses were chosen to analyse the results of in vitro tests compared to the in vivo fecal egg count reduction test (FECRT). The EHT indicated the presence of re...
Fernandes T, Robin M, Freeman KP.Biological variation in laboratory results refers to physiological fluctuations that occur around a homeostatic setpoint (HSP) for various laboratory measurands. Assessment of biological variation includes determining individual variation (CV), group variation (CV), and analytical variation (CV). Reference change value (RCV) is an objective tool for an evidence-based approach to interpret data by assessing the significance of consecutive results in an individual for the diagnosis, prognosis, and monitoring of disease. The individualised reference intervals (iRI) are derived from the determinat...
Ekinci G, Deniz O, Tüfekci E, Timur MC, Onmaz AC, Günes V, Citil M, Keles I, Hoven RVD.Rhodococcus equi (R. equi) is a primary cause of pyogranulomatous pneumonia of foals between three weeks and five months of age. Early diagnosis of rhodococcal pneumonia has always been considered a preferable approach as it can lead to more successful treatment and better outcomes. Horse stud farms where the disease is common, neonatal foals are subjected to tests such as complete blood count and fibrinogen analysis at certain intervals. However, new biomarkers are needed in addition to blood count and fibrinogen measurement in this field for early diagnosis of diseases. Based on this need, i...
Parsley AL, Hollingshead NA, Gruber EJ.To minimise storage-related artefacts, EDTA-anticoagulated whole blood should be evaluated promptly or refrigerated, a challenge in ambulatory practice. The effects of suboptimal blood storage on equine leukocyte parameters are not well-documented. Objective: Document changes in leukocyte number, automated and manual differential counts, and morphology in equine blood stored at four temperatures for up to 5 days. Methods: In vitro experimental. Methods: EDTA-anticoagulated whole blood from 13 horses was stored in a refrigerator (4°C), cooler with icepacks (3°C-19°C), room temperature (22...
Baghdadi HBA, Abdelsalam M, Attia MM.Strongylus vulgaris, a devastating parasitic nematode in equids, causes life-threatening verminous aneurysms that are challenging to diagnose early. This study pioneered integrating nanotechnology into an indirect enzyme-linked immunosorbent assay (i-ELISA) system to enhance the sensitivity and specificity for detecting S. vulgaris larval antigens in equine serum samples, with PCR confirmation of the species. A conventional i-ELISA and an innovative nano-based ELISA were developed using excretory-secretory antigens from adult S. vulgaris worms. The nano-ELISA incorporated gold nanoparticles (1...
Valderrama-Martinez C, Packham A, Zheng S, Smith W, Plancarte M, Aleman M.Evaluating antibody titers for Sarcocystis neurona for the diagnosis of equine protozoal myeloencephalitis from serum samples is a common practice. However, ensuring timely and proper refrigeration is not always possible. Objective: To evaluate immunofluorescent antibody (IFA) titers for S. neurona from serum samples stored at room temperature and 4°C. Methods: Twenty-two serum samples. Methods: Prospective longitudinal study. Two serum aliquots of 1 mL each were stored at room temperature (20-23.3°C) and 4°C. The unrefrigerated aliquot was immediately tested for IFA titers. Both aliquots...
Broeckl CV, Hiereth S, Straubinger RK.Diagnosis of equine Lyme borreliosis (LB), an infection caused by members of the Borrelia burgdorferi sensu lato complex (Bbsl), is challenging due to the nonspecific clinical signs of the disease and due to the variety of non-standardized serological tests. Specific vaccine-induced antibodies against LB, providing an effective protection against the infection, complicate the issue further. The standard for the detection of specific antibodies against Bbsl is a two-tier test system based on an enzyme-linked immunosorbent assay (ELISA) or indirect fluorescent antibody test (IFA) for antibody sc...
Chen JW, Stefanovski D, Haughan J, Jiang Z, Soma LR, Robinson MA.Interleukin 1 beta (IL-1β) and IL-1 receptor antagonist (IL-1RA) are both upregulated following traumatic injury. As IL-1RA blocks inflammatory signaling by IL-1β, overexpression of IL-1β relative to IL-1RA may drive inflammatory diseases. As such, determination of the relationship between IL-1β to IL-1RA expression levels in horses may provide insight into disease states or serve as a therapeutic readout of response to medical interventions. As techniques to detect plasma concentrations of IL-1β and IL-1RA in horses lack sensitivity, we developed and validated novel enzyme-linked immunos...
Yuki N, Shimazaki T, Kushiro A, Watanabe K, Uchida K, Yuyama T, Morotomi M.Selective adhesion to only certain epithelia is particularly common among the bacterial members of the indigenous microflora of mammals. We have found that the stratified squamous epithelium of the nonsecreting area of horse stomach is colonized by gram-positive rods. The microscopic features of a dense layer of these bacteria on the epithelium were found to be similar to those reported in mice, rats, and swine. Adhering microorganisms were isolated and identified as Lactobacillus salivarius, L. crispatus, L. reuteri, and L. agilis by DNA-DNA hybridization and 16S rRNA gene sequencing techniqu...
Maury W.In vivo, equine infectious anemia virus (EIAV) replicates in tissues rich in macrophages, and it is widely believed that the tissue macrophage is the principal, if not sole, cell within the host that replicates virus. No viral replication has been detected in circulating peripheral blood monocytes. However, proviral DNA can be detected in these cells, and monocytes may serve as a reservoir for the virus. In this study, an in vitro model was developed to clarify the role of monocyte maturation in regulating EIAV expression. Freshly isolated, nonadherent equine peripheral blood monocytes were in...
Derse D, Dorn PL, Levy L, Stephens RM, Rice NR, Casey JW.The long terminal repeats (LTRs) of equine infectious anemia virus (EIAV) were examined with respect to their ability to function as transcriptional promoters in various cellular environments. Nucleotide sequence analyses of the LTRs derived from two unique proviral clones revealed the requisite consensus transcription and processing signals. One of the proviruses possessed a duplication of a 16-base-pair sequence in the CCAAT box region of the LTR which was absent in the other provirus. To assess its functional activity, each LTR was coupled to the bacterial chloramphenicol acetyltransferase ...
Mancuso VA, Hope TJ, Zhu L, Derse D, Phillips T, Parslow TG.By systematically dissecting the Rev proteins of feline immunodeficiency virus (FIV) and equine infectious anemia virus (EIAV), we have identified within each a short peptide that is functionally interchangeable with the effector domains found in Rev-like proteins from other retroviruses. The active sequences from FIV and EIAV differ in several respects from other known effectors and may represent a distinct class of effector domain.
Klaus C, Hörügel U, Hoffmann B, Beer M.During a routine examination of 130 horse sera from 13 herds in Thuringia one TBEV antibody positive serum - with a very high titre - could be detected. The horse had been bought from a holding in Bavaria, and was reported to have clinical signs that may have been caused by a TBEV infection. To identify the source of the suspected TBEV infection, ticks from the surroundings of the barn in Thuringia as well as horse sera and ticks from two herds in Bavaria were examined. In the holding in Bavaria, where the horse was kept before, two out of ten horse sera were found to be TBEV antibody positive...
Clutterbuck AL, Smith JR, Allaway D, Harris P, Liddell S, Mobasheri A.This study employed a targeted high-throughput proteomic approach to identify the major proteins present in the secretome of articular cartilage. Explants from equine metacarpophalangeal joints were incubated alone or with interleukin-1beta (IL-1β, 10ng/ml), with or without carprofen, a non-steroidal anti-inflammatory drug, for six days. After tryptic digestion of culture medium supernatants, resulting peptides were separated by HPLC and detected in a Bruker amaZon ion trap instrument. The five most abundant peptides in each MS scan were fragmented and the fragmentation patterns compared to m...
Kilmartin JV, Rossi-Bernardi L.1. Three modified horse haemoglobins have been prepared: (i) alpha(c) (2)beta(c) (2), in which both the alpha-amino groups of the alpha- and beta-chains have reacted with cyanate, (ii) alpha(c) (2)beta(2), in which the alpha-amino groups of the alpha-chains have reacted with cyanate, and (iii) alpha(2)beta(c) (2), in which the two alpha-amino groups of the beta-chain have reacted with cyanate. 2. The values of n (the Hill constant) for alpha(c) (2)beta(c) (2), alpha(2)beta(c) (2) and alpha(c) (2)beta(2) were (respectively) 2.5, 2.0 and 2.6, indicating the presence of co-operative interactions ...
van den Hoogen BM, van Weeren PR, Lopes-Cardozo M, van Golde LM, Barneveld A, van de Lest CH.The amount of uronic acid residues in samples containing glycosaminoglycans or pectin is an important parameter in the quantitative and structural analysis of these complex carbohydrates. This paper describes a method to determine the content of uronic acids in biological samples, using conventional polystyrene microtiter plates and microtiter plate-reading equipment with standard interference filters (i.e., 540 or 492 nm). This assay is a modification of a commonly used procedure, viz. hydrolysis of uronic acid containing carbohydrate polymers in 80% sulfuric acid containing tetraborate ions ...
Gustafson AL, Tallmadge RL, Ramlachan N, Miller D, Bird H, Antczak DF, Raudsepp T, Chowdhary BP, Skow LC.A physical map of ordered bacterial artificial chromosome (BAC) clones was constructed to determine the genetic organization of the horse major histocompatibility complex. Human, cattle, pig, mouse, and rat MHC gene sequences were compared to identify highly conserved regions which served as source templates for the design of overgo primers. Thirty-five overgo probes were designed from 24 genes and used for hybridization screening of the equine USDA CHORI 241 BAC library. Two hundred thirty-eight BAC clones were assembled into two contigs spanning the horse MHC region. The first contig contain...
Welch HM, Bridges CG, Lyon AM, Griffiths L, Edington N.The polymerase chain reaction (PCR) and co-cultivation were used to identify the lymphoreticular system as the site of latency of equid herpesvirus I (EHV-1). Primers for PCR were designed from aligned nucleotide sequences of the glycoprotein gB genes to amplify the same region of both the EHV-1 and EHV-4 genomes. Subsequent restriction digests using specific enzymes distinguished the amplified fragments of the EHV-1 genome from those of the EHV-4 genome. Ten weeks following an experimental infection of five ponies with EHV-1, latent virus was detected by PCR and recovered by co-cultivation, p...
Christensen M, Jacobsen S, Ichiyanagi T, Kjelgaard-Hansen M.Major acute phase proteins (APPs) have proven diagnostically useful in dogs, cats and horses with routine use facilitated by commercially available automated heterologous assays. An automated assay applicable across all three species would highly facilitate further dissemination of routine use, and the aim of this study was to validate an automated latex agglutination turbidimetric immunoassay based on monoclonal anti-human serum amyloid A (SAA) antibodies for measurement of canine, feline and equine SAA. Serum samples from 60 dogs, 40 cats and 40 horses were included. Intra- and inter-assay i...
Dorn PL, Derse D.Deletion analysis of the equine infectious anemia virus long terminal repeat revealed that sequences responsive to virus-specific transactivation are located within the region spanning the transcriptional start site (-31 to +22). In addition, an active exon of a trans-acting factor (tat) was identified downstream of pol and overlapping env (nucleotides 5264 to 5461). Activation by tat is accompanied by an increase in the steady-state levels of mRNA directed by the equine infectious anemia virus long terminal repeat.
Böer U, Lohrenz A, Klingenberg M, Pich A, Haverich A, Wilhelmi M.Decellularized equine carotid arteries (dEAC) may represent a reasonable alternative to alloplastic materials in vascular replacement therapy. Acellularity of the matrix is standardly evaluated by DNA quantification what however may not record sufficiently the degree of matrix immunogenicity. Thus, our aim was to analyze dEAC with a low DNA content for residual cellular proteins. A detergent-based decellularization protocol including endonuclease treatment resulted in dEAC with 0.6 ± 0.15 ng DNA/mg dry weight representing 0.33 ± 0.14% of native tissue DNA content. In contrast, when matrices ...
Wieringa R, de Vries AA, Rottier PJ.Equine arteritis virus (EAV) is an enveloped, positive-stranded RNA virus belonging to the family Arteriviridae of the order NIDOVIRALES: Six transmembrane proteins have been identified in EAV particles: the nonglycosylated membrane protein M and the glycoprotein GP(5) (previously named G(L)), which occur as disulfide-bonded heterodimers and are the major viral envelope proteins; the unglycosylated small envelope protein E; and the minor glycoproteins GP(2b) (formerly designated G(S)), GP(3), and GP(4). Analysis of the appearance of the GP(2b), GP(3), and GP(4) proteins in viral particles by g...
Bowles DE, Holden VR, Zhao Y, O'Callaghan DJ.To assess the role of the equine herpesvirus type 1 (EHV-1) ICP0 protein (EICP0) in gene regulation, a variety of molecular studies on the EICP0 gene and gene products of both the attenuated cell culture-adapted Kentucky A (KyA) strain and the Ab4p strain were conducted. These investigations revealed that (i) the ICP0 open reading frame (ORF) of the KyA virus strain is 1,257 bp in size and would encode a protein of 419 amino acids, and in comparison to the ICP0 gene (ORF63) of the Ab4p strain of 1,596 bp (E. A. Telford, M. S. Watson, K. McBride, and A. J. Davison, Virology 189:304-316, 1992), ...
Diallo IS, Hewitson G, Wright L, Rodwell BJ, Corney BG.Equid herpesvirus 1 (EHV1) is a major disease of equids worldwide causing considerable losses to the horse industry. A variety of techniques, including PCR have been used to diagnose EHV1. Some of these PCRs were used in combination with other techniques such as restriction enzyme analysis (REA) or hybridisation, making them cumbersome for routine diagnostic testing and increasing the chances of cross-contamination. Furthermore, they involve the use of suspected carcinogens such as ethidium bromide and ultraviolet light. In this paper, we describe a real-time PCR, which uses minor groove-bindi...
Jörg A, Henderson WR, Murphy RC, Klebanoff SJ.Horse eosinophils purified to greater than 98% generated slow reacting substance (SRS) when incubated with the calcium ionophore A23187. On a per cell basis, eosinophils generated four to five times the SRS produced by similarly treated horse neutrophils. Eosinophil SRS production was inhibited by 5,8,11,14-eicosatetraynoic acid and augmented by indomethacin and arachidonic acid, suggesting that it was a product(s) of the lipoxygenase pathway of arachidonic acid metabolism. Compounds with SRS activity were purified by high-pressure liquid chromatography (HPLC) and identified by ultraviolet spe...
Sabeur K, Ball BA.Reactive oxygen species (ROS) play an important role in normal sperm function, and spermatozoa possess specific mechanisms for ROS generation via an NAD(P)H-dependent oxidase. The aim of this study was to identify the presence of an NADPH oxidase 5 (NOX5) in equine testis and spermatozoa. The mRNA of NOX5 was expressed in equine testis as detected by northern blot probed with human NOX5 cDNA and by RT-PCR. Immunoblotting with affinity purified alpha-NOX5 revealed one major protein in equine testis and other tissues. Immunolocalization of NOX5 showed labeling over the rostral sperm head with so...
Wagner B, Miller DC, Lear TL, Antczak DF.This report contains the first map of the complete Ig H chain constant (IGHC) gene region of the horse (Equus caballus), represented by 34 overlapping clones from a new bacterial artificial chromosome library. The different bacterial artificial chromosome inserts containing IGHC genes were identified and arranged by hybridization using overgo probes specific for individual equine IGHC genes. The analysis of these IGHC clones identified two previously undetected IGHC genes of the horse. The newly found IGHG7 gene, which has a high homology to the equine IGHG4 gene, is located between the IGHG3 ...
Science (New York, N.Y.)October 30, 1981
Volume 214, Issue 4520 562-564 doi: 10.1126/science.6270790
Studdert MJ, Simpson T, Roizman B.Viruses classified by immunologic criteria as equine herpesvirus 1 cause respiratory disease and abortion in horses. Restriction endonuclease analyses of the DNA's of viruses from animals with respiratory disease and from aborted fetuses show that the patterns for respiratory viruses, while similar to each other, are entirely different from the patterns for fetal viruses. It is therefore proposed that the DNA restriction endonuclease patterns of fetal and respiratory viruses analyzed in this study be designated as prototypic of equine herpesvirus 1 and 4, respectively.
Yokoyama S, Radlwimmer FB.To elucidate the molecular mechanisms of red-green color vision in mammals, we have cloned and sequenced the red and green opsin cDNAs of cat (Felis catus), horse (Equus caballus), gray squirrel (Sciurus carolinensis), white-tailed deer (Odocoileus virginianus), and guinea pig (Cavia porcellus). These opsins were expressed in COS1 cells and reconstituted with 11-cis-retinal. The purified visual pigments of the cat, horse, squirrel, deer, and guinea pig have lambdamax values at 553, 545, 532, 531, and 516 nm, respectively, which are precise to within +/-1 nm. We also regenerated the "true" red ...
Svraka S, van der Veer B, Duizer E, Dekkers J, Koopmans M, Vennema H.Acute gastroenteritis is one of the most common diseases worldwide, with viruses, particularly noroviruses, being the leading cause in developed countries. In The Netherlands, systematic surveillance of gastroenteritis outbreaks of suspected viral etiology was established by the National Institute for Public Health and the Environment in 1994. Since 2002, the total number of outbreaks reported has been increasing, and with that comes the need for sensitive assays that can be performed quickly. In addition, the diagnostic demand changed so that now the proportion of samples from hospitals is hi...
Wagner B, Radbruch A, Rohwer J, Leibold W.In this study we describe the generation of monoclonal antibodies (mAbs), which recognize different epitopes of the equine IgE constant heavy chain. Equi-murine recombinant IgE (rIgE), composed of the murine V(H)186.2 heavy chain variable region, linked to the equine IgE constant heavy chain and expressed together with the murine lambda(1) chain in J558L cells was used to immunize BALB/C mice. A total of 17 different mAbs were obtained, which recognized the rIgE heavy chain constant region. None of the mAbs reacted with monoclonal equine isotypes IgM, IgG1 (IgGa), IgG3 (IgG(T)), IgG4 (IgGb) or...
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...
Garvican ER, Cree S, Bull L, Smith RK, Dudhia J.Autologous mesenchymal stem cell (MSC) injection into naturally-occurring equine tendon injuries has been shown to be safe and efficacious and protocols inform translation of the technique into humans. Efficient transfer of cells from the laboratory into tissue requires well-validated transport and implantation techniques. Methods: Cell viability in a range of media was determined over 72 hours and after injection through a 19G, 21G or 23G needle. Viability, proliferation and apoptosis were analysed using TrypanBlue, alamarBlue® and AnnexinV assays. Results: Cell viability was similar in all...
Glaser AL, de Vries AA, Dubovi EJ.Three murine monoclonal antibodies (MAbs) that neutralize equine arteritis virus (EAV) infectivity were identified and characterized. The antibodies, 93B, 74D(B) and 38F, recognized the major envelope glycoprotein (GL) encoded by open reading frame (ORF) 5 in immunoblots and by immunoprecipitation. All three MAbs were used to compare the Bucyrus isolate of EAV and MAb neutralization-resistant (NR) escape mutants with the vaccine virus and 19 independent field isolates of EAV by virus neutralization. The different abilities of the MAbs to neutralize virus isolates indicated that they recognize ...
Nielsen JM.The objective of this study was to compare results from endometrial culture swabs with results from culturing of endometrial biopsies. The culture results were related to cytological findings (polymorphonuclear; PMN-cells) and histological observations (PMN-cells). Biopsy and swab samples were smeared on the surface of a blood agar petri dish, and examined for growth of bacteria. Cytology samples were obtained from endometrial biopsies, stained and examined under microscopy for the presence of PMN-cells. Endometrial biopsies were examined for the presence of PMN-infiltration of the endometrial...
Hoshino Y, Gorziglia M, Valdesuso J, Askaa J, Glass RI, Kapikian AZ.An equinine rotavirus FI-14 strain, originally isolated from a diarrheic foal in New York state, was shown to belong to serotype 3 by neutralization assay. In addition, it was found to react with both subgroup I and subgroup II monoclonal antibodies by enzyme-linked immunosorbent assay (ELISA), thus representing the first rotavirus strain to exhibit both subgroup specificities. By using hybridoma technology, we successfully produced monoclonal antibodies directed against the major inner capsid protein VP6 (the sixth gene product) of FI-14 virus. Such monoclonal antibodies reacted specifically ...
