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.
Medica AJ, Gibb Z, Aitken RJ.In vitro semen purification techniques have been developed that seek to mimic the in vivo selection process in order to generate the highest possible chance of oocyte fertilization following artificial insemination. Numerous methods have been developed to isolate functional spermatozoa for artificial insemination, yet only one method, single-layer centrifugation using commercial preparations like EquiPure, has been widely employed. In this study, we have introduced a novel approach for isolating spermatozoa and compared their quality to those isolated using EquiPure. The AI port system (Mempha...
Cheong SH, Franklin-Guild R, Goodman LB, Lago-Alvarez Y, Lee YL, de Aguiar LH, de Amorim MD, Altier C.Fungal and bacterial culture is currently the primary method for pathogen detection and identification. Next-generation sequencing is a powerful method for detecting and identifying the presence of microbial DNA in samples. We evaluated the correlation between fungal and bacterial culture with next-generation sequencing in equine uterine samples. Fungal cultures (n=63) were evaluated based on their culture results. In culture positive (n=16) samples, next-generation sequencing identified the same organism in 10 samples (62.5%), 5 samples did not identify fungal agents, and 1 sample identified ...
Gabryś J, Pietras N, Kowal-Mierzwa W, Karnas E, Andronowska A, Nowak A, Kochan J, Bugno-Poniewierska M.The efficacy of in vitro embryo production (IVEP) in equines is relatively limited compared to other species due to the lack of a reliable superovulation technique, limited availability of cumulus oocyte complexes (COCs), low in vitro oocyte maturation (IVM) and fertilization rates. Extracellular vesicles (EVs), which are nanoparticles involved in intercellular signaling in the ovarian environment, have shown potential as supplements to improve oocyte development during IVM. This study tested the hypothesis that EVs from small (< 20 mm) ovarian follicles could enhance fertilization rates in m...
Scupham AJ, Tong C.In 2018, a new virus, named equine parvovirus-hepatitis (EqPV-H), was discovered in a biologic product that had been administered to horses that subsequently developed clinical signs of equine serum hepatitis (Theiler disease). Further correlation of the virus with the disease sparked federal requirements that all equine biologics be free of EqPV-H. The initial quantitative real-time PCR (qPCR) test for EqPV-H has proved to be sensitive to co-extracted PCR inhibitors in template nucleic acids, causing false-negative results. We investigated the use of digital PCR (dPCR) as a more robust test. ...
Ohnuma K, Hirano-Kodaira M, Bannai M, Shimizu Y, Yamada M, Kinoshita K, Ngai-Wa Leung G, Ishii H.The abuse of prohibited peptide-based drugs with a broad spectrum of chemical characteristics poses a significant concern for the horseracing industry. Recently, there has been a notable increase in positive cases of small-peptide drugs reported in equine and canine sports. In addition to small peptides, large peptides (over 2 kDa) with structural diversity have also entered the market in increasing numbers as drugs for humans and livestock. However, the simultaneous analysis of both small- and large-peptide-based drugs is still challenging. In this study, a screening method was developed to c...
Ortiz-Rodriguez JM, Bucci D, Tovar-Pascual L, Granata S, Spinaci M, Nesci S.Sperm metabolism consists of a sophisticated network of biochemical reactions and varies between species, resulting in different metabolic strategies for ATP production to maintain sperm functionality. ATP can be produced through glycolysis or in the mitochondria by oxidative phosphorylation (OXPHOS). Since OXPHOS is the predominant metabolic pathway in horses spermatozoa, various assessments of mitochondrial activity are used to evaluate fertility, utilizing techniques such as fluorescent probes analysed via microscopy or flow cytometry, and polarographic electrode assays to measure current f...
Botía M, Martín-Cuervo M, Martínez-Subiela S, Cerón JJ, Ayala I, Hansen S, Muñoz-Prieto A.Equine Gastric Ulcer Syndrome (EGUS) is a widespread disease with a very high prevalence and importance in horses. It includes two conditions: Equine squamous gastric disease (ESGD), linked to acid disturbances, and Equine Glandular Gastric Disease (EGGD), associated with inflammation and immune system issues. In saliva, increased levels of immunological markers have been observed in this disease, indicating immune system involvement. In addition, changes in biomarkers of oxidative stress have been described. In horses, IgG and IgA are proteins that appear in saliva and serum and are considere...
Martin-Pelaez S, Fuente A, Takahashi K, Perez IT, Orozco J, Okada CTC, Neto CR, Meyers S, Dini P.Intracytoplasmic sperm injection (ICSI) is the current clinical practice for the in vitro production of equine embryos. The use of conventional fertilization methods such as in vitro fertilization (IVF), has historically been associated with poor success in horses. However, recent improvements have led to better outcomes with IVF, though only when using fresh semen, which limits its use in clinical practice. IVF remains in its infancy in equine reproduction, and several unknowns remain about the technique. One significant gap in knowledge concerns the morphokinetics of IVF embryos and how they...
Mersich I, Bishop RC, Diaz Yucupicio S, Nobrega AD, Austin SM, Barger AM, Fick ME, Wilkins PA.Coagulopathy is common in equine critical illness, with its early recognition being crucial for patient management and prognosis. In vitro viscoelastic (VE) hypercoagulability with decreased RCM/PCV has been demonstrated in dogs but not horses. Our objective was to evaluate the effects of acepromazine-induced (0.1 mg/kg IV) decreased RCM on VE and plasma coagulation parameters using a prospective interventional study of eight adult horses. Complete blood count (CBC), fibrinogen, prothrombin time (PT), partial thromboplastin time (PTT), packed cell volume (PCV), total solids (TS), and VCM Vet...
Hernández-Avilés C, Ramírez-Agámez L, Varner DD, Raudsepp T, Love CC.Cases of stallion subfertility due to acrosome dysfunction have been recognized since the 1990s. While some of these were observed in stallions with reduced sperm motility and morphology, a more severe form has been reported in stallions with normal-to-excellent sperm quality parameters, which is also uniquely observed in individuals of the Thoroughbred registry. These stallions carry a susceptibility genotype (A/A-A A in the gene FKBP6, exon 5) for Impaired Acrosomal Exocytosis (IAE). Current clinical observations from our group have identified a few highly subfertile stallions from other bre...
Lehman ML, Domenig O, Ames MK, Morgan JM.Furosemide, a commonly used diuretic, activates the renin-angiotensin-aldosterone system (RAAS) in other species. Little is known about RAAS peptide activation in horses. Objective: To evaluate equilibrium analysis as a practical method for RAAS quantification in horses and describe the RAAS response to a single dose of furosemide. We hypothesize that furosemide would cause transient increase in RAAS peptides in horses. Methods: 14 healthy adult thoroughbreds from a university teaching herd. Methods: Horses received either furosemide (1 mg/kg IV) or saline IV in a crossover study design. Pro...
Craig NM, Munguia NS, Trujillo AD, Chan AM, Wilkes R, Dorr M, Marsella R.This study investigated the effects of recombinant equine IL-31 (eIL-31) in vivo and in vitro. Methods: Equine IL-31 mRNA sequences were verified by sequencing. Recombinant eIL-31 was produced using mammalian and bacterial expression systems. From November 2019 through February 2021, 12 normal horses, 6 to 10 years old with no history or clinical signs consistent with allergic skin disease, were injected ID with eIL-31 and saline in 2 challenge studies. Pruritus-associated behaviors were recorded for a minimum of 15 minutes preinjection and 4 hours postinjection. Adherent monocytes from 3 prur...
Luis-Calero M, Muñoz-García CC, Fernández-Hernández P, Macías-García B, González-Fernández L.A protocol for conventional in vitro fertilization (IVF) in horses using fresh semen has been described, using a prolonged incubation in FERT-TALP medium (22 h) at 38.2°C in the presence of penicillamine, hypotaurine and epinephrine (PHE). Our work aimed to develop a protocol that maintains quality parameters in frozen-thawed equine spermatozoa incubated for 22 h in the presence of PHE using different media (FERT-TALP and INRA96) and incubation temperatures (30 and 38.2°C). Twelve frozen ejaculates from four stallions were thawed and then incubated in either FERT-TALP or INRA96 with PHE...
Vokes JR, Gedye KR, Lovett AL, de Kantzow MC, Shan R, Steel CM, Sykes BW.Gastrin is an important hormone involved in gastric acid secretion. Despite its importance, validated methods other than radioimmunoassay (RIA) to assess serum gastrin concentrations in horses are lacking. This study aims to determine the agreement between ELISA and RIA in quantifying equine serum gastrin concentrations. Serum gastrin concentrations were quantified using two ELISA kits and RIA. Samples (196) from 14 horses at different time points were analyzed using one ELISA kit and RIA, selected samples (7) were analyzed using a second ELISA kit, and the correlation between methods was calc...
Velineni S, Schiltz P, Chang KH, Peng YM, Cowles B.Abnormal blood glucose (BG) levels often seen in critically ill horses are significantly associated with adverse patient outcomes and increased mortality. Rapid and accurate BG monitoring is now considered an essential component of evidence-based equine practice and can provide critical information quickly for treatment. Although several point-of-care (POC) BG monitoring hand-held devices are commercially available for veterinary use, none contains a unique algorithm validated for use in horses. The AlphaTrak 3 (AT3) BG monitoring system is a first-of-its-kind device with an equine-specific al...
Brito LFC, Felix MR, Linardi RL, Martinez de Andino EV, Balamurugan NS, Hernández-Avilés C, Hinrichs K.Intracytoplasmic sperm injection (ICSI) is a valuable assisted reproduction technology in clinical practice, especially when semen availability is limited. Since the number of sperm required per ICSI cycle is much less than the number of sperm available in a standard straw of frozen semen, refreezing semen at lower sperm concentrations could yield multiple straws for ICSI use. However, there is little data on the effect of sperm refreezing on ICSI outcomes, especially on the effect of extender used for refreezing. The objective of the present study was to evaluate the effect of refreezing exte...
Peña FJ, Martín-Cano FE, Becerro-Rey L, da Silva-Álvarez E, Gaitskell-Phillips G, Ortega-Ferrusola C, Aparicio IM, Gil MC.Although stallion spermatozoa are now recognized as highly dependent on oxidative phosphorylation for ATP production in the mitochondria, most extenders in use contain supraphysiological concentrations of glucose as the main energy source. While the toxicity of cryoprotectants has been well documented in the literature, the potential toxicity of excessive glucose in extenders is largely ignored. However, the toxicity of excess glucose, known as "carbotoxicity", is well-established in many areas of medicine. In this paper, we review the basic aspects of stallion spermatozoa metabolism, focusing...
Vokes JR, Lovett AL, de Kantzow MC, Rogers CW, Wilkins PA, Sykes BW.Viscoelastic monitoring of horse coagulation is increasing due to its advantages over traditional coagulation testing. The use of a point-of-care viscoelastic coagulation monitor (VCM Vet™) has been validated for use in horses using native whole blood (NWB) but has not been assessed using citrated whole blood (CWB), a technique that might have advantages in practicality and precision. Blood was collected from 70 horses, tested in duplicate immediately using NWB (T0), and stored at room temperature as CWB for testing in duplicate at 1 (T1) and 4 (T4) hours after venipuncture for comparison to...
Hellstrom E, McKinney-Aguirre C, Gonzalez L, Ziegler A, Blikslager A.Inflammatory postoperative conditions of equine colic (acute abdomen) contribute not only to increased client cost, patient discomfort, and hospitalization time, but in many cases, prove to be life-threatening. A unique population of intestinal cells, enteric glia, are increasingly acknowledged for their roles in sensing the gastrointestinal environment and communicating with surrounding cell types. Interactions between enteric glia and intestinal epithelia may prove critical in establishing how equine enteric glia can alter the mucosal barrier to modulate inflammation in health and colic. To ...
Buschmann E, Van Steenkiste G, Duytschaever M, Segers P, Ibrahim L, van Loon G, Decloedt A.Radiofrequency ablation is a promising technique for arrhythmia treatment in horses. Due to the thicker myocardial wall and higher blood flow in horses, it is unknown if conventional radiofrequency settings used in human medicine can be extrapolated to horses. The study aim is to describe the effect of ablation settings on lesion dimensions in equine myocardium. To study species dependent effects, results were compared to swine myocardium. Right ventricular and right and left atrial equine myocardium and right ventricular swine myocardium were suspended in a bath with circulating isotonic sali...
Share ER, Mastellar SL, Suagee-Bedore JK, Eastridge ML.The measurement of fecal cortisol/corticosterone metabolites (FCMs) is often used to quantify the stress response. The sampling method is relatively non-invasive, reduces concern for elevation of cortisol from the sampling method, and has been shown to measure cortisol more consistently without the daily diurnal rhythm observed in blood. Commercial ELISA (enzyme-linked immunoassay) kits offer benefits over previously validated immunoassay methods but lack validation. The objective of this study was to evaluate a commercial ELISA kit (Arbor Assays DetectX Cortisol ELISA kit, K003-H1, Ann Arbor,...
Sergazy S, Zhetkenev S, Shulgau Z, Chulenbayeva L, Kamyshanskiy Y, Nurgaziyev M, Nurgozhina A, Mukhanbetzhanova Z, Berikkhanova K, Gulyayev A....Exosomes are cell-derived, membrane-surrounded particles that deliver bioactive molecules to various cells. Due to their small size, low immunogenicity, extended blood circulation, and involvement in cellular communication, they hold potential as effective drug carriers. Exosomes are present in various biological fluids, including mare's milk, a traditional drink in Central Asia. This study aims to compare exosome isolation methodologies and determine the stability of mare's milk-derived exosomes as potential therapeutic carriers. Three extraction methods-immunoprecipitation, size exclusion ch...
Ali YH, Mohieddeen TAG, Abdellatif MM, Ahmed BM, Saeed IK, Attaalfadeel HM, Ali AA.Rabies is endemic in Sudan with continuing outbreaks occurring annually, the most common animals affected are dogs, followed by goats and equids. This work focused on equid rabies, to elucidate the current situation of the disease through analysis of reports of equid rabies outbreaks in Sudan during 2010-2022 supported by laboratory confirmation of the disease. During the study period, 66 animals were affected during 35 equid rabies outbreaks. The highest incidences were found in Al Gezira (30.3%), followed by Darfur (24.2%) and Kordofan (15.2%). The highest incidence rate was observed during ...
Loup B, André F, Leuenberger N, Marchand A, Barnabé A, Delcourt V, Garcia P, Popot MA, Bailly-Chouriberry L.Detection and monitoring of biomarkers related to doping is particularly suitable for the development of analytical strategies dedicated to indirect detection of banned substances. Previous studies in horses have already allowed the investigation of transcriptomic biomarkers in equine blood associated with reGH and rHuEPO administrations. Our most recent developments continue to focus on the discovery and monitoring of transcriptomic biomarkers for the control of ESAs, and a collaborative study with WADA-accredited doping control laboratories has recently been initiated to conduct a pilot stud...
Neufang L, Ramos J, Eda S, Flatland B, Giori L.Pituitary pars intermedia dysfunction (PPID) is a neurodegenerative disease of senior horses. Loss of dopaminergic inhibition of the melanotropes of the pars intermedia leads to increased concentrations of pro-opiomelanocortin (POMC)-derived peptides. Diagnosis is challenging due to pre-analytical variables, such as sample storage, handling, and time to analysis. Our objective was to develop an ELISA for ACTH measurement, which could ultimately form the basis for a stall-side equine ACTH test. We selected 2 ACTH-specific monoclonal antibodies, CBL57 and EPR20361-248, based on the recognition o...
Delcourt V, Garcia P, Chabot B, Aber N, Pescher M, Cacault M, Scholtes P, Loup B, Barnabé A, Popot MA, Bailly-Chouriberry L.Thymosin 4 (TB4) is a ubiquitous, highly conserved and abundant peptide among mammals with a critical role in cytoskeleton organization. In spite of its yet non-authorized use as a medicine and being forbidden by the IFHA, the FEI, and the WADA, intelligence and doping control laboratories reported numerous products available online claiming to contain a synthetic acetylated fragment of TB4 or TB4 itself, promoted as a growth factor with regenerative properties. In this article, the first estimation of the endogenous TB4 concentration in racing horses' blood samples was performed through a po...
Cheung HW, Wong KS, Tam JCL, Farrington AF, Bond AJ, Wan TSM, Ho ENM.Erythropoiesis-stimulating agents (ESAs) continue to be a significant threat to the integrity of human and equine sports. Besides conventional direct testing, monitoring the biomarkers associated with the effects of ESAs may provide a complementary approach via indirect detection to enhance doping control. In this study, we applied label-free proteomics to discover plasma protein biomarkers in Thoroughbred geldings after administration with a long-acting form of recombinant human erythropoietin (rhEPO), methoxy polyethylene glycol epoetin beta, Mircera. Increased haematocrit, haemoglobin and r...
Bussiman F, Alves AAC, Richter J, Hidalgo J, Veroneze R, Oliveira T.Gait scores are widely used in the genetic evaluation of horses. However, the nature of such measurement may limit genetic progress since there is subjectivity in phenotypic information. This study aimed to assess the application of machine learning techniques in the prediction of breeding values for five visual gait scores in Campolina horses: dissociation, comfort, style, regularity, and development. The dataset contained over 5000 phenotypic records with 107,951 horses (14 generations) in the pedigree. A fixed model was used to estimate least-square solutions for fixed effects and adjusted ...
Arifianto D, Esfandiari A, Wibawan IWT, Amrozi A, Maharani M, Darsono D, Setiadi H, Setiyono A.Horses used for antisera production are repeatedly hyperimmunized to produce high levels of specific antibodies. This prolonged process can lead to various health issues, including amyloidosis, which involves the accumulation of amyloid proteins in organs and tissues, potentially causing organ dysfunction and failure. These horses are often retired when they no longer produce adequate antibody levels. This study aimed to evaluate the impact of prolonged antisera production on the health of retired horses by examining their blood biochemistry and serum amyloid A (SAA) levels, which are indicato...
Aguiar LH, Pinto CRF.Sperm capacitation is considered the main factor limiting conventional in vitro fertilization (IVF) in horses. A recent scientific breakthrough in sperm processing for IVF in horses has resulted in embryos and foals being produced; however, various aspects of the IVF process remain to be fully elucidated. Lysine acetylation has been shown to play a role in sperm capacitation in several species and the objective of this study was to detect and evaluate this process in the horse. Ejaculates of two stallions were collected and incubated in different conditions with deacetylase inhibitors to induc...
Warnken T, Huber K, Feige K.Exact analysis of equine insulin in blood samples is the key element for assessing insulin resistance or insulin dysregulation in horses. However, previous studies indicated marked differences in insulin concentrations obtained from sample analyses with different immunoassays. Most assays used in veterinary medicine are originally designed for use in human diagnostics and are based on antibodies directed against human insulin, although amino acid sequences between equine and human insulin differ. Species-specific assays are being used more frequently and seem to provide advantages compared to ...
Argüelles D, Carmona JU, Pastor J, Iborra A, Viñals L, Martínez P, Bach E, Prades M.The aim of this study was to evaluate single and double centrifugation tube methods for concentrating equine platelets. Whole blood samples were collected from clinically normal horses and processed by use of single and double centrifugation tube methods to obtain four platelet concentrates (PCs): PC-A, PC-B, PC-C, and PC-D, which were analyzed using a flow cytometry hematology system for hemogram and additional platelet parameters (mean platelet volume, platelet distribution width, mean platelet component concentration, mean platelet component distribution width). Concentrations of transformi...
Yi J, Heinecke J, Tan H, Ford PC, Richter-Addo GB.It is now well-established that mammalian heme proteins are reactive with various nitrogen oxide species and that these reactions may play significant roles in mammalian physiology. For example, the ferrous heme protein myoglobin (Mb) has been shown to reduce nitrite (NO(2)(-)) to nitric oxide (NO) under hypoxic conditions. We demonstrate here that the distal pocket histidine residue (His64) of horse heart metMb(III) (i.e., ferric Mb(III)) has marked effects on the mode of nitrite ion coordination to the iron center. X-ray crystal structures were determined for the mutant proteins metMb(III) H...
Allen GP, Coogle LD.The molecular structure of the equine herpesvirus type 1 (EHV-1) gene encoding glycoprotein 13 (gp13) was analyzed. The gene is contained within a 1.8-kilobase AccI-EcoRI restriction fragment mapping at map coordinates 0.136 to 0.148 in the UL region of the EHV-1 genome and is transcribed from right to left. Determination of the nucleotide sequence of the DNA fragment revealed a complete transcriptional unit composed of typical regulatory promoter elements upstream to a long open reading frame (1,404 base pairs) that encoded a 468-amino-acid primary translation product of 51 kilodaltons. The p...
Lannergård J, Guss B.Streptococcus equi ssp. equi is the causative agent of strangles, a highly contagious and serious disease in the upper respiratory tract of horses. The present study describes the characterization of IdeE, a homolog of the secreted IgG-specific protease IdeS/Mac of Streptococcus pyogenes. The activity of IdeE is compared with the activity of IdeZ, the corresponding enzyme of the closely related S. equi ssp. zooepidemicus. A study of the proteolytic activity of recombinant IdeE and IdeZ on IgG from a selection of mammals shows that only antibodies containing the substrate site of IdeS/Mac are c...
Kulikov E, Kropinski AM, Golomidova A, Lingohr E, Govorun V, Serebryakova M, Prokhorov N, Letarova M, Manykin A, Strotskaya A, Letarov A.Lytic coliphage vB_EcoP_G7C and several other highly related isolates were obtained repeatedly from the samples of horse feces held in the same stable thus representing a component of the normal indigenous intestinal communities in this population of animals. The genome of G7C consists of 71,759 bp with terminal repeats of about 1160 bp, yielding approximately 73 kbp packed DNA size. Seventy-eight potential open reading frames, most of them unique to N4-like viruses, were identified and annotated. The overall layout of functional gene groups was close to that of the original N4 phage, with som...
Hamada T, Watanabe G, Kokuho T, Taya K, Sasamoto S, Hasegawa Y, Miyamoto K, Igarashi M.A sensitive radioimmunoassay (RIA) for the determination of inhibin in peripheral plasma and tissue homogenates of different species has been developed using antisera to partially purified bovine follicular fluid (bFF) inhibin and 125I-labelled bFF 32 kDa inhibin. Antisera were produced by immunization of rabbits with partially purified bFF inhibin prepared by immunoaffinity chromatography. Increasing doses of a high titre antiserum could neutralize the suppressing effect of bFF, porcine follicular fluid and rat ovarian homogenate on FSH secretion from rat anterior pituitary cells in culture. ...
Van de Walle GR, Peters ST, VanderVen BC, O'Callaghan DJ, Osterrieder N.Equine herpesvirus 1 (EHV-1) is a member of the Alphaherpesvirinae, and its broad tissue tropism suggests that EHV-1 may use multiple receptors to initiate virus entry. EHV-1 entry was thought to occur exclusively through fusion at the plasma membrane, but recently entry via the endocytic/phagocytic pathway was reported for Chinese hamster ovary cells (CHO-K1 cells). Here we show that cellular integrins, and more specifically those recognizing RGD motifs such as alphaVbeta5, are important during the early steps of EHV-1 entry via endocytosis in CHO-K1 cells. Moreover, mutational analysis revea...
Decourt C, Tillet Y, Caraty A, Franceschini I, Briant C.To determine if kisspeptin could be implicated in the control of reproduction in equine species, we studied the distribution of kisspeptin neurons and their anatomical interactions with GnRH neurons in the hypothalamus of pony mares. Brains were collected in three pony mares between 2 and 4h after ovulation. One major population of kisspeptin immunoreactive cell bodies was found in the arcuate nucleus (ARC), where they extended from the middle of the nucleus to the premammillary recess. Kisspeptin immunoreactive varicose fibers extended from the preoptic area to the mammillary nuclei, with imp...
Giannitti F, Diab S, Mete A, Stanton JB, Fielding L, Crossley B, Sverlow K, Fish S, Mapes S, Scott L, Pusterla N.Equine coronavirus (ECoV) is a Betacoronavirus recently associated clinically and epidemiologically with emerging outbreaks of pyrogenic, enteric, and/or neurologic disease in horses in the United States, Japan, and Europe. We describe the pathologic, immunohistochemical, ultrastructural, and molecular findings in 2 horses and 1 donkey that succumbed to natural infection with ECoV. One horse and the donkey (case Nos. 1, 3) had severe diffuse necrotizing enteritis with marked villous attenuation, epithelial cell necrosis at the tips of the villi, neutrophilic and fibrinous extravasation into th...
Hdud IM, Mobasheri A, Loughna PT.The metabolic activity of articular chondrocytes is influenced by osmotic alterations that occur in articular cartilage secondary to mechanical load. The mechanisms that sense and transduce mechanical signals from cell swelling and initiate volume regulation are poorly understood. The purpose of this study was to investigate how the expression of two putative osmolyte channels [transient receptor potential vanilloid 4 (TRPV4) and large-conductance Ca(2+)-activated K(+) (BKCa)] in chondrocytes is modulated in different osmotic conditions and to examine a potential role for MAPKs in this process...
Steel CM.The most important application for synovial fluid (SF) analysis in the horse is in the diagnosis of synovial sepsis. Misdiagnosis of synovial sepsis is costly, and SF analysis makes correct diagnosis more likely, although far from certain. The precision of diagnosis may be increased with polymerase chain reaction analysis for detection of bacterial DNA in SF and with assays for various enzymes and cytokines. These tests are currently not widely available, however, and routine SF analysis remains of prime importance in diagnosis.
Nunokawa Y, Fujinaga T, Taira T, Okumura M, Yamashita K, Tsunoda N, Hagio M.Serum amyloid A protein (SAA) was isolated from equine acute-phase serum by repeating Sephadex G-75 gel filtration 3 times. Quantitative measurement of equine SAA was performed by the single radial immunodiffusion technique with rabbit anti-equine SAA serum. In clinically normal horses, the SAA concentration remained relatively high from immediately after birth up to 1 week of age. After this the concentration showed periodic fluctiation in the range of approximately 13 to 30 micrograms/ml. The mean (+/- SD) concentration of SAA in foals ( or = 18 months old) was 19.37 +/- 9.41 and 21.53 +/- 9...
Iqbal K, Chitwood JL, Meyers-Brown GA, Roser JF, Ross PJ.Formation of the inner cell mass (ICM) and trophectoderm (TE) marks the first differentiation event in mammalian development. These two cell types have completely divergent fates for the remainder of the developmental process. The molecular mechanisms that regulate ICM and TE formation are poorly characterized in horses. The objective of this study was to establish the transcriptome profiles of ICM and TE cells from horse blastocysts using RNA sequencing (RNA-seq). A total of 12 270 genes were found to be expressed in either lineage. Global analysis of the transcriptome profiles by unsupervi...
Liu ZP.The diagnosis of lead poisoning combined with cadmium in sheep and horses living on farmland in the vicinity of non-ferrous metal smelters in Baiyin of Gansu province in China was based on laboratory findings as well as clinical signs. The concentrations of lead, cadmium, copper and zinc in soils, water, forages, feed and blood, hair and tissues of affected sheep and horses were determined in the surroundings of the smelters in Baiyin and controls. Haematological values were also measured. The concentrations of these elements in soils, forages, water and feed were significantly higher than tho...
Sales KG, Costa PL, de Morais RC, Otranto D, Brandão-Filho SP, Cavalcanti Mde P, Dantas-Torres F.Phlebotomine sand flies are blood-feeding insects of great medical and veterinary significance acting as vectors of Leishmania parasites. Studying the blood-feeding pattern of these insects may help in the understanding of their interactions with potential reservoir hosts of Leishmania parasites. In this study, we developed real time PCR assays for the identification of sand fly blood meal. Methods: Six pairs of primers were designed based on cytochrome b gene sequences available in GenBank of the following potential hosts: dog, cat, horse, chicken, black rat, and human. Firstly, SYBR Green-ba...
Genzel Y, Behrendt I, König S, Sann H, Reichl U.The production of equine influenza in Madin-Darby canine kidney (MDCK) cells in large-scale microcarrier culture is described with detailed on- and off-line analytical data during cell growth and virus replication. Metabolite concentration profiles for glucose, glutamine, lactate and ammonium are shown. Lactate and ammonium concentrations were always below inhibiting levels. Concentration profiles for essential and non-essential amino acids of the cell culture medium are discussed. During cell growth proline was released into the medium with a significant rate while two amino acids, serine and...
Lin M, Zhang C, Gibson K, Rikihisa Y.Neorickettsia risticii is an obligate intracellular bacterium of the trematodes and mammals. Horses develop Potomac horse fever (PHF) when they ingest aquatic insects containing encysted N. risticii-infected trematodes. The complete genome sequence of N. risticii Illinois consists of a single circular chromosome of 879 977 bp and encodes 38 RNA species and 898 proteins. Although N. risticii has limited ability to synthesize amino acids and lacks many metabolic pathways, it is capable of making major vitamins, cofactors and nucleotides. Comparison with its closely related human pathogen N. senn...
Ladrón N, Fernández M, Agüero J, González Zörn B, Vázquez-Boland JA, Navas J.The actinomycete Rhodococcus equi is an important pathogen of horses and an emerging opportunistic pathogen of humans. Identification of R. equi by classical bacteriological techniques is sometimes difficult, and misclassification of an isolate is not uncommon. We report here on a specific PCR assay for the rapid and reliable identification of R. equi. It is based on the amplification of a fragment of the choE gene encoding cholesterol oxidase. The choE-based PCR was assessed by using a panel of strains comprising 132 isolates from different sources and of different geographical origins, all i...
Billich C, Sauder C, Frank R, Herzog S, Bechter K, Takahashi K, Peters H, Staeheli P, Schwemmle M.The recent observation that Borna disease virus (BDV)-reactive antibodies from psychiatric patients exhibit only low avidity for BDV antigen called into question their diagnostic value and raised the possibility that antigenically related microorganisms or self antigens caused the production of these antibodies. We further characterized the specificity of these antibodies. Methods: We established a peptide array-based screening test that allows the identification of antibodies directed against linear epitopes of the two major BDV proteins, the nucleoprotein (N) and the phosphoprotein (P). Resu...
Balasuriya UB, Leutenegger CM, Topol JB, McCollum WH, Timoney PJ, MacLachlan NJ.A one-tube real-time TaqMan reverse transcription-polymerase chain reaction (RT-PCR) assay was developed for the detection of equine arteritis virus (EAV). The test was validated using the seminal plasma and nasal secretions of infected horses that were proven to contain EAV by traditional virus isolation in rabbit kidney thirteen (RK-13) cells, as well as a variety of cell culture-propagated European and North American strains of EAV. The primers and a fluorogenic TaqMan probe were designed to amplify and detect a highly conserved region of open reading frame 7 (ORF7) of EAV. The real-time Ta...
Ellegren H, Johansson M, Sandberg K, Andersson L.We have isolated equine microsatellites by screening a genomic library with (TG)n and (TC)n probes. TG microsatellites were found to be more abundant than TC repeats, with an estimated frequency of one per 100,000bp. Sequence analysis of eight TG-positive clones revealed varying structures of the repeat regions; perfect stretches of TG repeats, imperfect stretches of TG repeats and compound regions of TG and TC repeats. Five loci were analysed by PCR and showed extensive polymorphism; three to seven alleles and heterozygosities of 0.40-0.76 were observed when screening 20-30 unrelated individu...
Westermann CM, Dorland L, Votion DM, de Sain-van der Velden MG, Wijnberg ID, Wanders RJ, Spliet WG, Testerink N, Berger R, Ruiter JP, van der Kolk JH.The aim of the current study was to assess lipid metabolism in horses with atypical myopathy. Urine samples from 10 cases were subjected to analysis of organic acids, glycine conjugates, and acylcarnitines revealing increased mean excretion of lactic acid, ethylmalonic acid, 2-methylsuccinic acid, butyrylglycine, (iso)valerylglycine, hexanoylglycine, free carnitine, C2-, C3-, C4-, C5-, C6-, C8-, C8:1-, C10:1-, and C10:2-carnitine as compared with 15 control horses (12 healthy and three with acute myopathy due to other causes). Analysis of plasma revealed similar results for these predominantly...
Silaghi C, Liebisch G, Pfister K.Equine Granulocytic Anaplasmosis (EGA) is caused by Anaplasma phagocytophilum, a tick-transmitted, obligate intracellular bacterium. In Europe, it is transmitted by Ixodes ricinus. A large number of genetic variants of A. phagocytophilum circulate in nature and have been found in ticks and different animals. Attempts have been made to assign certain genetic variants to certain host species or pathologies, but have not been successful so far. The purpose of this study was to investigate the causing agent A. phagocytophilum of 14 cases of EGA in naturally infected horses with molecular methods o...
Chotwiwatthanakun C, Pratanaphon R, Akesowan S, Sriprapat S, Ratanabanangkoon K.The purpose of this study was to prepare a potent polyvalent antivenom against three elapids namely, the Thai cobra (Naja kaouthia, NK), the King cobra (Ophiophagus hannah, OH) and the banded krait (Bungarus fasciatus, BF). Two groups of horses were immunized. Group 1, comprising five horses, was immunized twice with a mixture of postsynaptic neurotoxins followed by an additional six immunizations with a mixture of crude venoms of the three elapids. Group 2, comprising four horses, was immunized with a mixture of crude venoms throughout the course. For the first immunization, the immunogens we...
Chowdhary BP, Raudsepp T, Kata SR, Goh G, Millon LV, Allan V, Piumi F, Guérin G, Swinburne J, Binns M, Lear TL, Mickelson J, Murray J, Antczak DF....A first-generation radiation hybrid (RH) map of the equine (Equus caballus) genome was assembled using 92 horse x hamster hybrid cell lines and 730 equine markers. The map is the first comprehensive framework map of the horse that (1) incorporates type I as well as type II markers, (2) integrates synteny, cytogenetic, and meiotic maps into a consensus map, and (3) provides the most detailed genome-wide information to date on the organization and comparative status of the equine genome. The 730 loci (258 type I and 472 type II) included in the final map are clustered in 101 RH groups distribute...
Hartmann J, Lindau M.Exocytosis begins with formation of a small fusion pore which then expands allowing rapid release of granular contents. We studied the influence of cytoplasmic free Ca2+ ([Ca2+]i) on the conductance of the initial pore and on the dynamics of subsequent expansion in horse eosinophils using the patch clamp technique. The mean initial conductance is approximately 200 pS independent of [Ca2+]i. This value is close to that previously found in beige mouse mast cells. The pore subsequently expands by 18 nS/s at [Ca2+]i < 10 nM, by 40 nS/s at [Ca2+]i = 1.5 microM and by 90 nS/s at [Ca2+]i = 10 micr...
Baquero-Perez B, Kuchipudi SV, Nelli RK, Chang KC.Current methods of isolation of muscle satellite cells from different animal species are highly variable making inter-species comparisons problematic. This variation mainly stems from the use of different proteolytic enzymes to release the satellite cells from the muscle tissue (sometimes a single enzyme is used but often a combination of enzymes is preferred) and the different extracellular matrix proteins used to coat culture ware. In addition, isolation of satellite cells is frequently laborious and sometimes may require pre-plating of the cell preparation on uncoated flasks or Percoll cent...
