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.
Zibura AE, Cullen MA, Rutledge H, Lassalle L, Salmon JH, Gilger BC, Westermeyer HD.Determine optimal iontophoresis times for riboflavin delivery to the corneal stroma across different species and compare these to corneal injection. Methods: Ex vivo horse, dog, rabbit, and pig globes were treated with riboflavin administered with either iontophoresis for 2.5-20 minutes with or without corneal epithelium; or with purpose-designed precise corneal injection (PCI) application with intact epithelium. Immediately following riboflavin administration, samples were harvested, frozen, and sectioned. Riboflavin penetration was imaged using fluorescence microscopy. Results: Horse sample...
Chan GHM, Tang W, Curl P, Lin Y, Wan TSM, Ho ENM.Arsenic can be easily found in our surrounding environment. Because of its ubiquitous nature, horse urine and blood invariably contain low levels of arsenic. Nevertheless, inorganic arsenic, despite its general use as a tonic for horses, is an effective doping agent having a deleterious effect because of its ability to induce gastroenteritis. The misuse of arsenic in horseracing has been controlled by an international urinary threshold of total arsenic at 0.3 μg/mL. However, an equivalent threshold for total arsenic in plasma is yet to be established. In this study, an inductively coupled pla...
Kwok KY, Choi TLS, Kwok WH, Lau MY, Leung EMK, Leung GNW, Wong JKY, Wan TSM, Adrian FF, Prabhu A, Ho ENM.The use of bioactive peptides as a doping agent in both human and animal sports has become increasingly popular in recent years. As such, methods to control the misuse of bioactive peptides in equine sports have received attention. This paper describes a sensitive accurate mass method for the detection of 40 bioactive peptides and two non-peptide growth hormone secretagogues (< 2 kDa) at low pg/mL levels in horse urine using ultra-high performance liquid chromatography-high resolution mass spectrometry (UHPLC/HRMS). A simple mixed-mode cation exchange solid-phase extraction (SPE) cartridge was...
Leung DKK, Wong ASY, Zhou QL, Wan TSM, Ho ENM.A non-target variable Data Independent Acquisition (vDIA) workflow based on accurate mass measurements using a Q Exactive OrbiTrap is presented for the first time for equine doping control testing. The vDIA workflow uses a combination of MS1 events (1 to 2) and multiple vDIA events to cover the analytes of interest. The workflow basically captures a digital image of a sample allowing all relevant MS1 and MS2 data to be recorded. In theory, the workflow can accommodate an unlimited number of analytes as long as they are amenable to the sample extraction protocol and fall within the mass limits ...
Garcia P, Pinètre J, Morel S, Jaubert M, Deruy X, Perot I, Delcourt V, Loup B, Popot MA, Bailly-Chouriberry L.Bisphosphonates are prohibited drugs according to Article 6 of the International Agreement on Breeding, Racing and Wagering of the International Federation of Horseracing Authorities (IFHA) and the International Equestrian Federation (FEI). These compounds are used for the treatment of lameness, navicular and bone diseases in horses and are divided into two groups: non-nitrogen-containing bisphosphonate drugs (e.g. clodronic acid) and nitrogen-containing bisphosphonate drugs (e.g. zoledronic acid). Their hydrophilic properties and the high affinity for the bone matrix make the control of their...
Tomlinson JE, Wolfisberg R, Fahnøe U, Sharma H, Renshaw RW, Nielsen L, Nishiuchi E, Holm C, Dubovi E, Rosenberg BR, Tennant BC, Bukh J, Kapoor A....Pegiviruses frequently cause persistent infection (as defined by >6 months), but unlike most other Flaviviridae members, no apparent clinical disease. Human pegivirus (HPgV, previously GBV-C) is detectable in 1-4% of healthy individuals and another 5-13% are seropositive. Some evidence for infection of bone marrow and spleen exists. Equine pegivirus 1 (EPgV-1) is not linked to disease, whereas another pegivirus, Theiler's disease-associated virus (TDAV), was identified in an outbreak of acute serum hepatitis (Theiler's disease) in horses. Although no subsequent reports link TDAV to disease, an...
Cruz A, Sullivan DB, Doty KF, Hess RA, Canisso IF, Reddi PP.The acrosome plays a critical role in sperm-oocyte interactions during fertilization. SP-10 is an acrosomal matrix protein, which is evolutionarily conserved among mammals. The SP-10 antibody has been shown to be useful for staging the seminiferous cycle in the mouse and human. A canonical acrosomal marker; however, has never been used for staging in the horse. The objectives of the present study were to investigate the presence of SP-10 within the horse acrosome using an anti-mouse SP-10 antibody, to classify spermatids based on the shape of the acrosome, and then to use that information to a...
Chidlow H, Giguère S, Camus M, Wells B, Berghaus R, McConachie Beasley E.Serial cerebrospinal fluid (CSF) analysis might be required in clinical neurologic disease. The effect of lumbosacral (LS) or cervical (C1-C2) centesis on subsequent CSF cytologic analyses has not been investigated in horses. Objective: To evaluate the effect of thecal puncture on subsequent CSF analyses ANIMALS: Ten healthy adult horses. Methods: Prospective study. Horses were randomly assigned to undergo CSF collection twice, 14 days apart, from either the C1-C2 or LS space. After a 4-month washout period, CSF collection was repeated from the alternate site. Continuous data were analyzed u...
Marr N, Hopkinson M, Hibbert AP, Pitsillides AA, Thorpe CT.Three-dimensional imaging modalities for optically dense connective tissues such as tendons are limited and typically have a single imaging methodological endpoint. Here, we have developed a bimodal procedure utilising fluorescence-based confocal microscopy and x-ray micro-computed tomography for the imaging of adult tendons to visualise and analyse extracellular sub-structure and cellular composition in small and large animal species. Results: Using fluorescent immunolabelling and optical clearing, we visualised the expression of the novel cross-species marker of tendon basement membrane, lam...
Göktaş EF, Kabil E, Arıöz F.Nonsteroidal anti-inflammatory drugs (NSAIDs) are commonly used in therapeutic doses in human and veterinary medicine for the treatment of inflammation, pain, and fever. A method for the simultaneous determination of nine NSAIDs, known as therapeutic prohibited substances, in equine urine was developed and fully validated according to the European Commission Decision 2002/657/EC and Association of Official Racing Chemists criteria. The validation was performed for naproxen, flunixin, ketoprofen, diclofenac, eltenac, meclofenamic acid, phenylbutazone, vedaprofen, and carprofen in equine urine i...
Donkey placenta does not allow the passage of immunoglobulins; thus, foals are born hypogammaglobulinemic and an adequate intake of high-quality colostrum in the first 24 hours of life is crucial for the surviving. The study aims to assess the relation between colostrum immunoglobulin G (IgG) concentration evaluated by the single radial immunodiffusion (SRID) test and the Brix refractometer in donkeys to establish a cutoff value for high quality of colostrum based on Brix refractometry. Colostrum was collected at foaling, and at 6, 12, and 24 hours after foaling from the left and the right h...
Lombardi F, Palumbo P, Augello FR, Giusti I, Dolo V, Guerrini L, Cifone MG, Giuliani M, Cinque B.The ability of a collagen-based matrix to support cell proliferation, migration, and infiltration has been reported; however, the direct effect of an aqueous collagen suspension on cell cultures has not been studied yet. In this work, the effects of a high-concentration aqueous suspension of a micronized type I equine collagen (EC-I) have been evaluated on a normal mouse fibroblast cell line. Immunofluorescence analysis showed the ability of EC-I to induce a significant increase of type I and III collagen levels, parallel with overexpression of crucial proteins in collagen biosynthesis, matura...
Aitken RJ, Gregoratos D, Kutzera L, Towney E, Lin M, Wilkins A, Gibb Z.MTT is widely used in biology as a probe for cell viability by virtue of its ability to generate deposits of insoluble formazan at sites of intense oxidoreductase activity. This response is generally held to reflect mitochondrial redox activity; however, extra-mitochondrial MTT reduction has also been recorded in certain cell types. Given this background, we set out to determine the major sites of formazan deposition in mammalian spermatozoa. In the mouse, most MTT reduction took place within the extensive mitochondrial gyres, with a single minor site of formazan deposition on the sperm head. ...
Valle-Casuso JC, Gaudaire D, Martin-Faivre L, Madeline A, Dallemagne P, Pronost S, Munier-Lehmann H, Zientara S, Vidalain PO, Hans A.RNA viruses are responsible for a large variety of animal infections. Equine Arteritis Virus (EAV) is a positive single-stranded RNA virus member of the family Arteriviridae from the order Nidovirales like the Coronaviridae. EAV causes respiratory and reproductive diseases in equids. Although two vaccines are available, the vaccination coverage of the equine population is largely insufficient to prevent new EAV outbreaks around the world. In this study, we present a high-throughput in vitro assay suitable for testing candidate antiviral molecules on equine dermal cells infected by EAV. Using t...
Data in briefJune 20, 2020
Volume 31 105887 doi: 10.1016/j.dib.2020.105887
Martín-Cano FE, Gaitskell-Phillips G, Ortiz-Rodríguez JM, Silva A, Gil C, Ortega-Ferrusola C, Peña FJ.This paper provides the dataset of proteins of stallion ejaculates before and after cryopreservation. The data report the analysis and identification of stallion sperm proteins obtained from the same ejaculates and split in two subsamples. The first aliquot consisted on fresh spermatozoa and the second aliquot was frozen and thawed spermatozoa. Samples were analyzed using a UHPLC/MS/MS system consisting of an Agilent 1290 infinity series UHPLC coupled to an Agilent 6550 Q-TOF mass spectrometer (Agilent Technologies, Santa Clara, CA, USA). We provide a data set of 2226 different proteins, with ...
Journal of proteomicsJune 19, 2020
Volume 225 103880 doi: 10.1016/j.jprot.2020.103880
Chiaradia E, Miller I.Equine medicine represents a relevant field of veterinary science and the horse industry generates a significant economic impact. Horses can be involved in different sport disciplines, meat production, work and recreational purposes. Horses are also important for human health as they can be used as animal models for studying human diseases and in equine-assisted therapy. This review summarizes the data related to body fluids such as plasma/serum, urine, cerebrospinal fluid, synovial fluid, saliva, bronchoalveolar lavage fluid and peritoneal fluid obtained using proteomic analysis. Horse body f...
Rahnama S, Vathsangam N, Spence R, Anderson ST, de Laat MA, Bailey S, Sillence MN.Prolonged hyperinsulinemia is thought to be the cause of equine endocrinopathic laminitis, a common and crippling disease of the foot, for which there are no pharmacologic treatments other than pain relief. It has been suggested that insulin causes its effects on the lamellae by activating IGF-1 receptors (IGF-1R), as insulin receptors (InsR) are scarce in this tissue, whereas IGF-1R are abundant and become downregulated after prolonged insulin infusion. As a first step toward confirming this mechanism and beginning to develop a therapeutic anti-IGF-1R monoclonal antibody (mAb) for horses, it ...
Torquato RBC, Iamamoto K, Fernandes ER, Achkar S, Silva SR, Katz ISS, Guedes F.Laboratory diagnosis of rabies in equines is essential for distinguishing the disease from other sources of encephalitis. Diagnosis by conventional techniques such as a direct fluorescent antibody test (dFAT) or viral isolation in mice or cell culture can be difficult, and the application of molecular biological methods may be necessary. We performed an indirect rapid immunohistochemistry test (iRIT) for the detection of the rabies virus (RABV) antigen in the central nervous system (CNS) of equines and compared the results with those of other diagnostic techniques. We reviewed result records f...
Gautier C, Scarlet D, Ertl R, Walter I, Wulf M, Nagel C, Aurich J, Aurich C.The aim of the present study was to characterise key enzymes involved in polyunsaturated fatty acid (PUFA) synthesis in the testis and epididymis collected from 2-year-old healthy warmblood stallions (n=10). The mRNA expression of fatty acid synthase, the Δ9-, Δ6-, Δ5- and Δ4-desaturases and elongases 6, 5 and 2 (encoded by the fatty acid synthase (FASN), the stearoyl-CoA desaturase (SCD), the fatty acid desaturase 2 (FADS2), the fatty acid desaturase 1 (FADS1), the delta 4-desaturase, sphingolipid 1 (DEGS1), ELOVL fatty acid elongase 6(ELOVL6), ELOVL fatty acid elongase 5 (ELOVL5), ELOVL ...
Schaefer RJ, McCue ME.High-quality genomic tools have been integral in understanding genomic architecture and function in the modern-day horse. The equine genetics community has a long tradition of pooling resources to develop genomic tools. Since the equine genome was sequenced in 2006, several iterations of high throughput genotyping arrays have been developed and released, enabling rapid and cost-effective genotyping. This review highlights the design considerations of each iteration, focusing on data available during development and outlining considerations in selecting the genetic variants included on each arr...
Cakir-Koc R, Budama-Kilinc Y, Ustun E, Babur C.Toxoplasma gondii is a parasite that causes severe health problems in the world. Toxoplasmosis, an infection caused by T. gondii, leads to high risk of mortality in patients with immunodeficiency, transplantation, and cancer. Besides that, it causes miscarriages in pregnancy, various abnormalities such as hydrocephalus in infants and congenital diseases. Because the clinical indication of the disease is not specific, it is confused with many diseases, and this leads to the necessity of directly detecting the presence of the toxoplasmosis. Therefore, various diagnostic assays are needed for th...
Waraksa E, Owczarek K, Kubica P, Kłodzińska E, Ozimek M, Wrzesień R, Bobrowska-Korczak B, Namieśnik J.Bromhexine (BH), expectorant used in the treatment of respiratory disorders associated with viscid or excessive mucus, is not permitted for use in the competing horse by many authorities in horseracing and Olympic disciplines. Metabolic studies are of the great importance in anti-doping field because they allow for updating the selection of the most appropriate markers for prohibited substances, such as metabolites present at higher concentration levels and/or lasted for a longer period of time in biological samples than a parent drug. This study describes LC-MS/MS-based method for simultaneou...
Rubessa M, Feugang JM, Kandel ME, Schreiber S, Hessee J, Salerno F, Meyers S, Chu I, Popescu G, Wheeler MB.The capacity for microscopic evaluation of sperm is useful for assisted reproductive technologies (ART), because this can allow for specific selection of sperm cells for in vitro fertilization (IVF). The objective of this study was to analyze the same sperm samples using two high-resolution methods: spatial light interference microscopy (SLIM) and atomic force microscopy (AFM) to determine if with one method there was more timely and different information obtained than the other. To address this objective, there was evaluation of sperm populations from boars and stallions. To the best of our k...
Li N, Dou S, Feng L, Zhu Q, Lu N.In matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), the analyte is usually distributed unevenly throughout the sample spot. The area with aggregated analyte molecules contributing abundant signal, is termed as "sweet spot", which results in poor detection reproducibility and makes it impossible to quantify analytes without internal standards. We proposed a strategy to eliminate sweet spot in MALDI-MS by using a hydrophobic ordered structure as target. The target is fabricated by creating a hydrophobic silicon nanopillar array and subsequently decorating it uniformly wi...
Marcos R, Ribeiro T, Santos M, Correia-Gomes C, Macedo S, Guimarães T, Blielbernicht M, Caniatti M, Rocha A.Uterine lavage (UL) is a routine diagnostic procedure for endometritis. In UL the fluid is centrifuged and the sediment smeared. Samples prepared in cytocentrifuges, the so-called "cytospins", are useful for evaluating cells in fluids, but never been used in UL. The aim of this study was to assess the usefulness of cytospins after UL, comparing automatic versus manual cytocentrifuges, and to determine their value for the diagnosis of endometritis. The study was divided in two parts. Firstly, UL was performed in 16 mares and a small part of the retrieved fluid was cytocentrifuged in an automati...
Durham AE, Kemp-Symonds J.Serology is commonly used as a means of identifying horses that might be chronic and silent carriers of S. equi but its sensitivity is rarely examined. Objective: The study was designed to investigate the sensitivity of serological testing for antibodies against S. equi antigens A and C to detect guttural pouch carriers of S. equi. Methods: Retrospective clinical study. Methods: As part of routine surveillance and quarantine procedures horses arriving at a welfare charity quarantine unit were subject to both microbiological sampling of guttural pouches and also serological testing for antibo...
Shibany KA, Pratt SL, Aldurdunji M, Totemeyer S, Paine SW.Horses are exposed to various kinds of medication, however, there are limited determinations of plasma clearance (CL) for the drugs used due to the high cost of equine studies.Many of the CL values generated come from the equine sports industry for determining drug plasma screening limits in the control of medications at the time of competition.The kinetics of omeprazole metabolism were investigated in freshly isolated and cryopreserved equine hepatocytes and hepatic microsomes ( = 3 horses).The V, K and intrinsic clearance (CL) of omeprazole were determined via the substrate depletion me...
Podstawski P, Witarski W, Szmatoła T, Bugno-Poniewierska M, Ropka-Molik K.Sarcoids are the most common skin neoplasm in the Equidae family. Sarcoids are benign, but may cause severe damage in affected animals. Due to the high risk of post-treatment recurrence and the lack of an effective method of treatment, it is reasonable to perform studies on the molecular aspects of this neoplasm. Therefore, the present studies analyzed five genes (cell cycle control binding protein alpha, coronin 1b, metalloproteinase 2, tissue inhibitor of metalloproteinases 3 and vimentin) related to cell mobility and invasion traits. Primary healthy fibroblasts and sarcoid cells were obtain...
Wong JKY, Chan GHM, Choi TLS, Kwok KY, Lau MY, Leung GNW, Wan TSM, Ho ENM.A high-throughput method has been developed for the doping control analysis of 124 drug targets, processing up to 154 horse urine samples in as short as 4.5 h, from the time the samples arrive at the laboratory to the reporting deadline of 30 min before the first race, including sample receipt and registration, preparation and instrument analysis and data vetting time. Sample preparation involves a brief enzyme hydrolysis step (30 min) to detect both free and glucuronide-conjugated drug targets. This is followed by extraction using solid-supported liquid extraction (SLE) and analysis using liq...
Roberts D, Cotter HVT, Cubeta M, Gilger BC.To determine minimum inhibitory concentrations (MICs) of four fungal species isolated from horses presented with equine fungal keratitis (EFK) in the southeastern United States to previously untested azole, echinocandin, and carboxamide antifungal drugs. Methods: In vitro assays were performed to determine the susceptibility of Aspergillus flavus, A. fumigatus, Fusarium falciforme, and F. keratoplasticum to five antifungal drugs representing three modes of action. Results: Luliconazole exhibited increased growth inhibition against both Aspergillus and Fusarium compared to commonly used, stan...
Cordoni G, Williams A, Durham A, Florio D, Zanoni RG, La Ragione RM.Strangles is one of the most common equine infectious diseases with serious health, welfare and socio-economic impact. However, the detection of Streptococcus equi subspecies equi can be challenging and persistently infected carriers are common. Furthermore, the use of classical microbiology can result in an underestimation of the prevalence of the disease. The difficulties associated with the slow diagnosis of Strangles can result in rapid spread of the disease. Therefore, rapid and economical diagnostic tests are urgently required. Here, two multiplex assays, were developed and validated for...
von Teichman BF, Smit TK.The polyvalent African Horsesickness (AHS) attenuated live vaccine (ALV) produced by Onderstepoort Biological Products (OBP) Ltd., South Africa, has been associated with some safety concerns and alleged cases of vaccine failure or vaccine-induced disease. The risk of reassortment and reversion to virulence is a common concern associated with the use of ALVs, and a phenomenon reported for viruses with segmented RNA genomes. The purpose of this study was to determine whether or not reassortment of AHS vaccine strains could result in reassortants and reversion to virulence and therefore cause AHS...
Nemoto M, Morita Y, Niwa H, Bannai H, Tsujimura K, Yamanaka T, Kondo T.A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the rapid detection of equine coronavirus (ECoV). This assay was conducted at 60 °C for 40 min. Specificity of the RT-LAMP assay was confirmed using several equine intestinal and respiratory pathogens in addition to ECoV. The novel assay failed to cross-react with the other pathogens tested, suggesting it is highly specific for ECoV. Using artificially synthesized ECoV RNA, the 50% detection limit of the RT-LAMP assay was 10(1.8)copies/reaction. This is a 50-fold greater sensitivity than conventio...
Borchers K, Wolfinger U, Goltz M, Broll H, Ludwig H.Equine herpesvirus type 2 (EHV-2) is a slow-growing, cytopathogenic gammaherpesvirus, which is suggested to be ubiquitous in the equine population. However, its precise role as a pathogen and its tissue tropism remains uncertain. To estimate the prevalence of EHV-2 in Germany and to investigate the possible pathogenicity of the virus, peripheral blood leucocytes (PBL) from 172 horses were examined for EHV-2 DNA by a sensitive and specific nested PCR based on the EcoRI-N genomic fragment and by classical cocultivation. PBL samples from 51% of the horses were positive by PCR and virus was isolat...
Svala E, Löfgren M, Sihlbom C, Rüetschi U, Lindahl A, Ekman S, Skiöldebrand E.The molecular aspects of inflammation were investigated in equine articular cartilage explants using quantitative proteomics. Articular cartilage explants were stimulated with interleukin (IL)-1β in vitro for 25 days, and proteins released into cell culture media were chemically labeled with isobaric mass tags and analyzed by liquid chromatography-tandem mass spectrometry. A total of 127 proteins were identified and quantified in media from explants. IL-1β-stimulation resulted in an abundance of proteins related to inflammation, including matrix metalloproteinases, acute phase proteins, comp...
Eder C, Curik I, Brem G, Crameri R, Bodo I, Habe F, Lazary S, Sölkner J, Marti E.To investigate whether allergen-specific IgE production is influenced by environmental and genetic factors, IgE levels against 2 mould extracts (Alternaria alternata [Alt a] and Aspergillus fumigatus [Asp f]) and against recombinant (r) rAlt a 1, rAsp f 7 and rAsp f 8 were determined by ELISA in sera from 448 Lipizzan horses living in 6 studfarms. Statistical evaluation showed a significant effect of studfarm-specific environment on IgE levels against the different allergens, but genetic factors also influenced allergen-specific IgE production: an heritability of 0.33 was found for IgE levels ...
Adelson DL, Raison JM, Garber M, Edgar RC.The interspersed repeat content of mammalian genomes has been best characterized in human, mouse and cow. In this study, we carried out de novo identification of repeated elements in the equine genome and identified previously unknown elements present at low copy number. The equine genome contains typical eutherian mammal repeats, but also has a significant number of hybrid repeats in addition to clade-specific Long Interspersed Nuclear Elements (LINE). Equus caballus clade specific LINE 1 (L1) repeats can be classified into approximately five subfamilies, three of which have undergone signifi...
Rogers HJ.Under a variety of conditions of concentration, Eh, and pH, horse anti- serum and normal horse serum exerted similar bacteriostatic effects against Type A. Ferric iron abolished the bacteriostatic effect when added during the first 2 hours of incubation at Eh+60 mV. Ferrous iron abolished the bacteriostatic effect when added after 3 hours. Ferric iron abolished the bacteriostatic effect at—140 mV. A mixture consisting of horse β- and γ-globulins together with human transferrin exerted a bacteriostatic effect similar to that of whole serum. This system responded in the same way as whole se...
Melnick JL, Hampil B.This paper summarizes the results of co-operative studies undertaken by the WHO International Reference Centre for Enteroviruses and a number of WHO Regional Reference Centres for viruses, WHO Virus Collaborating Laboratories, or other laboratories in a comprehensive testing programme of enterovirus equine antisera. The studies were designed to appraise the specificity of immune serum prepared in horses against five representative prototype enteroviruses (poliovirus 1, coxsackieviruses A9 and B3, and echoviruses 4 and 11). Tests for neutralizing antibody were performed not only against the hom...
Holl HM, Vanhnasy J, Everts RE, Hoefs-Martin K, Cook D, Brooks SA, Carpenter ML, Bustamante CD, Lafayette C.Genetic markers are important resources for individual identification and parentage assessment. Although short tandem repeats (STRs) have been the traditional DNA marker, technological advances have led to single nucleotide polymorphisms (SNPs) becoming an attractive alternative. SNPs can be highly multiplexed and automatically scored, which allows for easier standardization and sharing among laboratories. Equine parentage is currently assessed using STRs. We obtained a publicly available SNP dataset of 729 horses representing 32 diverse breeds. A proposed set of 101 SNPs was analyzed for DNA ...
Hackett ES, Lunn DP, Ferris RA, Horohov DW, Lappin MR, McCue PM.Neonatal sepsis is a common problem in foals and is a primary cause of death in the post natal period. Transient bacteraemia and subsequent host responses have not been described in the equine neonate. Objective: The primary objective of this study was to determine if transient bacteraemia occurs in foals within the first 72 h of life. Additional objectives included description of bacterial organisms associated with transient bacteraemia and concurrent cytokine gene expression in healthy foals. Methods: Prospective observational study in healthy foals. Methods: Blood was aseptically collecte...
Arosalo BM, Raekallio M, Rajamäki M, Holopainen E, Kastevaara T, Salonen H, Sankari S.The aim of the study was to investigate urine matrix metalloproteinase (MMP-2 and -9) activity, alkaline phosphatase/creatinine (U-AP/Cr) and gamma-glutamyl-transpeptidase/creatinine (U-GGT/Cr) ratios, glucose concentration, and urine protein/creatinine (U-Prot/Cr) ratio and to compare data with plasma MMP-2 and -9 activity, cystatin-C and creatinine concentrations in colic horses and healthy controls. Horses with surgical colic (n = 5) were compared to healthy stallions (n = 7) that came for castration. Blood and urine samples were collected. MMP gelatinolytic activity was measured by zymogra...
Brinkmeyer-Langford C, Raudsepp T, Lee EJ, Goh G, Schäffer AA, Agarwala R, Wagner ML, Tozaki T, Skow LC, Womack JE, Mickelson JR, Chowdhary BP.A high-resolution (1 marker/700 kb) physically ordered radiation hybrid (RH) and comparative map of 122 loci on equine homologs of human Chromosome 19 (HSA19) shows a variant evolution of these segments in equids/Perissodactyls compared with other mammals. The segments include parts of both the long and the short arm of horse Chromosome 7 (ECA7), the proximal part of ECA21, and the entire short arm of ECA10. The map includes 93 new markers, of which 89 (64 gene-specific and 25 microsatellite) were genotyped on a 5000-rad horse x hamster RH panel, and 4 were mapped exclusively by FISH. The orie...
Suchan T, Kusliy MA, Khan N, Chauvey L, Tonasso-Calvière L, Schiavinato S, Southon J, Keller M, Kitagawa K, Krause J, Bessudnov AN, Bessudnov AA....DNA hybridization-capture techniques allow researchers to focus their sequencing efforts on preselected genomic regions. This feature is especially useful when analysing ancient DNA (aDNA) extracts, which are often dominated by exogenous environmental sources. Here, we assessed, for the first time, the performance of hyRAD as an inexpensive and design-free alternative to commercial capture protocols to obtain authentic aDNA data from osseous remains. HyRAD relies on double enzymatic restriction of fresh DNA extracts to produce RNA probes that cover only a fraction of the genome and can serve a...
Dieckmann SM, Winkler M, Groebel K, Dieckmann MP, Hofmann-Lehmann R, Hoelzle K, Wittenbrink MM, Hoelzle LE.Haemotrophic mycoplasmas (HM) are parasites on the surface of red blood cells and known to infect a wide range of animals. However, there are no previous evidences of HM infections in horses. In this study HM were detected for the first time in the blood of two horses suffering from poor performance, apathy, weight loss, and anaemia. Using a HM specific PCR assay and subsequent sequencing the infective agents isolated from the blood of said horses were confirmed as closely related to the HM species Mycoplasma haemofelis and 'Candidatus Mycoplasma haemobos'.
Schott HC, Gallant LR, Coyne M, Murphy R, Cross J, Strong-Townsend M, Szlosek D, Yerramilli M, Li J.Symmetric dimethylarginine (SDMA) is considered a more sensitive indirect estimate of glomerular filtration rate (GFR) than creatinine (Cr). Symmetric dimethylarginine is not affected by sex or muscle mass in small animals. Objective: To validate a commercial SDMA immunoassay (IA) for equine serum; to compare SDMA and Cr in cohorts of draft horse breeds; and to assess effects of age, sex, and breed. Methods: One hundred and sixty-five healthy draft horses (0.5-16 years), including 63 Percherons, 52 Clydesdales, and 50 Belgians. Methods: Cross-sectional study. The SDMA IA was validated for eq...
Bernet A, Bastien A, Soulet D, Jerczynski O, Roy C, Bianchi Rodrigues Alves M, Lecours C, Tremblay MÈ, Bailey JL, Robert C, Belleannée C.Where are primary cilia (PC) organelles located during postnatal epididymal development? Our findings unveil the existence of PC sensory organelles in different epididymal cell types according to postnatal development stage. Primary cilia are sensory organelles that orchestrate major signaling pathways during organ development and homeostasis. Epididymal PC have been detected in the horses, donkey and mules but their cell-lineage specificity has never been investigated in this organ. A longitudinal study was performed by examining tissue from n = 3 to n = 10 transgenic mice at different times ...
Moczygemba C, Guidry J, Wittung-Stafshede P.Native myoglobin (Mb) consists of two populations which differ in the orientation of the heme by 180 degrees rotation (as verified by nuclear magnetic resonance) but have identical absorption spectra and equilibrium-thermodynamic stability. Here, we report that these two fractions of native oxidized Mb (from horse) both unfold and refold (chemical denaturant, pH 7, 20 degrees C) in two parallel kinetic reactions with rate constants differing 10-fold. In accord, the oxidized heme remains coordinated to unfolded horse Mb in up to 4 M guanidine hydrochloride (pH 7, 20 degrees C).
Gaynor AM, Zhu KW, Dela Cruz FN, Affolter VK, Pesavento PA.Bovine papillomaviruses (BPV1/BPV2) have long been associated with equine sarcoids; deciphering their contribution has been difficult due to their ubiquitous presence on skin and in the environment, as well as the lack of decent techniques to interrogate their role in pathogenesis. We have developed and characterized an in situ hybridization (ISH) assay that uses a pool of probes complementary to portions of the E5, E6, and E7 genes. This assay is highly sensitive for direct visualization of viral transcript and nucleic acid in routinely processed histopathologic samples. We demonstrate here t...
Schroeder B, Boyle MD, Sheerin BR, Asbury AC, Lottenberg R.Our laboratory previously demonstrated that group C streptococcal isolates from humans and horses secrete streptokinases that preferentially activate plasminogens reflecting the origin of the isolates. To analyze the significance of these findings, series of streptokinase-producing Streptococcus equisimilis isolates recovered from humans and horses were examined. Southern blot analysis revealed that chromosomal DNA of the streptococcal isolates from humans reacted exclusively with a skc(hu) probe and that chromosomal DNA of streptococcal isolates from horses reacted preferentially with an skc(...
Stankov B, Cozzi B, Lucini V, Fumagalli P, Scaglione F, Fraschini F.Melatonin receptors were characterized in the brains of three mammals (rabbit, horse and sheep) by an in vitro binding technique, using 2-[125I]iodomelatonin as labelled ligand. Although binding sites for melatonin have been described recently in several vertebrate species (including the sheep), the rabbit and the horse have not been the subject of investigation so far. Apart from characterization, the present report describes receptor distribution in a number of brain regions, thus allowing for direct interspecies comparison under the same methodological conditions. 2-[125I]iodomelatonin labe...
Evenberg A, Meyer H, Gaastra W, Verheij HM, De Haas GH.The complete amino acid sequence of phosphlipase A2 (EC 3.1.1.4) from horse pancreas was determined. The protein controls of a single polypeptide chain of 125 amino acids and has a molecular weight of 13,927. The chain is crosslinked by seven disulfide bridges. The sequence was determined by automated Edman degradation of the intact protein and several of the large peptide fragments. Smaller peptides were analyzed by manual Edman degradation. Fragmentation of the peptide chain was accomplished by enzymatic digestion with trypsin, chymotrypsin, and thermolysin. The final overlap was found by di...
Hussain KA, Issel CJ, Schnorr KL, Rwambo PM, West M, Montelaro RC.Monoclonal antibodies (MCAbs) were used to dissect the antigenic sites of the surface glycoproteins of the prototype cell-adapted Wyoming strain of equine infectious anemia virus (EIAV). Serologic reactivities of these MCAbs were determined by ELISA, additive ELISA, competitive ELISA, and Western blot assays. The results indicated that antigenic reactivity of gp90 was localized on at least four distinct epitopes, two of which were important in neutralization. Our studies also revealed that these epitopes were localized on overlapping antigenic sites on gp90. On the other hand, only two distinc...
Rombouts C, Jeanneau C, Camilleri J, Laurent P, About I.Adequate revascularization is a prerequisite for successful healing of periodontal bone defects. This study characterized three different xenogeneic bone grafting materials: Gen-Os of equine and porcine origins, and anorganic Bio-Oss. We also investigated their angiogenic potential. All materials were composed of poorly crystalline calcium oxide phosphate, with Bio-Oss exhibiting a carbonated phase and larger particle size and both Gen-Os showing the presence of collagen. Both Gen-Os materials significantly enhanced vascular endothelial growth factor (VEGF) secretion by PDL cells. A significan...
Chagas JR, Hirata IY, Juliano MA, Xiong W, Wang C, Chao J, Juliano L, Prado ES.The present studies demonstrate the importance of subsite interactions in determining the cleavage specificities of kallikrein gene family proteinases. The effect of substrate amino acid residues in positions P3-P'3 on the catalytic efficiency of tissue kallikreins (rat, pig, and horse) and T-kininogenase was studied using peptidyl-pNA and intramolecularly quenched fluorogenic peptides as substrates. Kinetic analyses show the different effects of D-amino acid residues at P3, Pro at P'2, and Arg at either P'1 or P'3 on the hydrolysis of substrates by tissue kallikreins from rat and from horse o...
Lawrence WC.Autoradiographic analyses of deoxyribonucleic acid (DNA) synthesis in randomly growing KB cell cultures infected with equine abortion virus (EAV) suggested that viral DNA synthesis was initiated only at times that coincided with the entry of noninfected control cells into the S phase of the cell cycle. Synchronized cultures of KB cells were infected at different stages of the cell cycle, and rates of synthesis of cellular and viral DNA were measured. When cells were infected at different times within the S phase, viral DNA synthesis was initiated 2 to 3 hr after infection. However, when cells ...
Reimers TJ, Lamb SV, Bartlett SA, Matamoros RA, Cowan RG, Engle JS.Veterinary diagnostic endocrinology laboratories frequently receive hemolyzed plasma, serum, or blood samples for hormone analyses. However, except for the previously reported harm done by hemolysis to canine insulin, effects of hemolysis on quantification of other clinically important hormones are unknown. Therefore, these studies were designed to evaluate effects of hemolysis on radioimmunoassay of thyroxine, 3,5,3'-triiodothyronine, progesterone, testosterone, estradiol, cortisol, and insulin in equine, bovine, and canine plasma. In the first experiment, hormones were measured in plasma obt...
Albrecht RA, Kim SK, O'Callaghan DJ.The equine herpesvirus 1 (EHV-1) EICP27 protein cooperates with either the immediate-early (IE) or the EICP0 protein to synergistically trans-activate viral promoters. GST-pulldown and co-immunoprecipitation assays revealed that the EICP27 protein's cooperation with the IE or the EICP0 protein involves its physical interaction with these viral proteins. In the case of the IE-EICP27 protein interaction, IE residues 424 to 826 and EICP27 residues 41 to 206 harbor the interactive domains. Electrophoretic mobility shift assays (EMSA) suggested that the EICP27 protein is not a sequence-specific DNA...
Davis DG, Schaefer DM, Hinchcliff KW, Wellman ML, Willet VE, Fletcher JM.Hypogammaglobulinemia as a result of failure of transfer of passive immunity (FTPI) is an important risk factor for infectious disease in neonatal foals. The current gold standard for determining serum immunoglobulin concentrations is radial immunodiffusion (RID). The purpose of this study was to compare immunoglobulin concentrations measured by RID with those determined by an automated turbidimetric immunoassay (TIA), which has a much shorter turnaround time. Immunoglobulin concentrations were measured by both RID and TIA in serum collected from 84 neonatal foals. Sixty-seven foals had result...
