The equine immune system is a complex network of cells, tissues, and organs that work collaboratively to defend against pathogens and maintain homeostasis. It consists of innate and adaptive components, each with distinct functions and mechanisms. The innate immune system provides the first line of defense through physical barriers, phagocytic cells, and the complement system. The adaptive immune system involves lymphocytes, such as B cells and T cells, which generate specific responses to antigens and provide immunological memory. Research in equine immunology explores the interactions between these components, the impact of genetic and environmental factors on immune function, and the development of vaccines and therapeutics. This page gathers peer-reviewed studies and scholarly articles focusing on the mechanisms, regulation, and clinical applications of the equine immune system in health and disease.
Springer NL, Smith E, Brooks MB, Stokol T.To develop a flow cytometric assay to quantify platelet-derived microparticles (PMPs) in equine whole blood and plasma. Methods: Citrate-anticoagulated whole blood from 30 healthy adult horses. Methods: Platelet-poor plasma (PPP) was prepared from fresh whole blood by sequential low-speed centrifugation (twice at 2,500 × g). Samples of fresh whole blood and PPP were removed and stored at 4° and 24°C for 24 hours. Platelet-derived microparticles were characterized in fresh and stored samples on the basis of the forward scatter threshold (log forward scatter < 10(1)) and labeling with anne...
Canty MJ, Fogarty U, Sheridan MK, Ensley SM, Schrunk DE, More SJ.Four primary mycotoxicosis have been reported in livestock caused by fungal infections of grasses or cereals by members of the Clavicipitaceae family. Ergotism (generally associated with grasses, rye, triticale and other grains) and fescue toxicosis (associated with tall fescue grass, Festuca arundinacea) are both caused by ergot alkaloids, and referred to as 'ergot alkaloid intoxication'. Ryegrass staggers (associated with perennial ryegrass Lolium perenne) is due to intoxication with an indole-diperpene, Lolitrem B, and metabolites. Fescue-associated oedema, recently described in Australia, ...
Pusterla N, Mackie S, Packham A, Conrad PA.The aim of the present study was to investigate the likelihood of transplacental transmission of Neospora hughesi and Sarcocystis neurona in foals, born from seropositive mares. Three broodmares with persistent N. hughesi infection gave birth to eight healthy foals over a period of 7 years. These foals were seropositive to N. hughesi prior to colostrum ingestion, with titers ranging between 640 and 20,480, measured by indirect fluorescence antibody test (IFAT). Of 174 foals born at another farm to mares with a high seroprevalence to S. neurona, only one (with a pre-colostrum antibody tit...
Braune S, Walter M, Schulze F, Lendlein A, Jung F.For in vitro studies assessing the interaction of platelets with implant materials, common and standardized protocols for the preparation of platelet rich plasma (PRP) are lacking, which may lead to non-matching results due to the diversity of applied protocols. Particularly, the aging of platelets during prolonged preparation and storage times is discussed to lead to an underestimation of the material thrombogenicity. Here, we study the influence of whole blood- and PRP-storage times on changes in platelet morphology and function. Blood from apparently healthy subjects was collected according...
Giles C, Vanniasinkam T, Ndi S, Barton MD.For decades researchers have been targeting prevention of Rhodococcus equi (Rhodococcus hoagui/Prescottella equi) by vaccination and the horse breeding industry has supported the ongoing efforts by researchers to develop a safe and cost effective vaccine to prevent disease in foals. Traditional vaccines including live, killed and attenuated (physical and chemical) vaccines have proved to be ineffective and more modern molecular-based vaccines including the DNA plasmid, genetically attenuated and subunit vaccines have provided inadequate protection of foals. Newer, bacterial vector vaccines hav...
Burk SV, Dangoudoubiyam S, Brewster-Barnes T, Bryant UK, Howe DK, Carter CN, Vanzant ES, Harmon RJ, Kazacos KR, Rossano MG.Currently, diagnosis of Parascaris equorum infection in equids is limited to patent infections. The goals of this study were to culture P. equorum larvae in vitro and identify excretory-secretory (ES) products for prepatent diagnostic testing. Parascaris equorum L2/L3 larvae were hatched and cultured for up to 3 weeks for ES product collection. Fifth stage (L5) P. equorum were also cultured for ES product collection. Examination of ES fractions by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and silver stain revealed L2/L3 products ranging from 12-94 kDa and L5 products...
Tanaka T, Kasai H, Yamashita A, Okuyama-Dobashi K, Yasumoto J, Maekawa S, Enomoto N, Okamoto T, Matsuura Y, Morimatsu M, Manabe N, Ochiai K....Equine hepacivirus (EHcV) has been identified as a closely related homologue of hepatitis C virus (HCV) in the United States, the United Kingdom, and Germany, but not in Asian countries. In this study, we genetically and serologically screened 31 serum samples obtained from Japanese-born domestic horses for EHcV infection and subsequently identified 11 PCR-positive and 7 seropositive serum samples. We determined the full sequence of the EHcV genome, including the 3' untranslated region (UTR), which had previously not been completely revealed. The polyprotein of a Japanese EHcV strain showed ap...
Boivin R, Vargas A, Lefebvre-Lavoie J, Lauzon AM, Lavoie JP.Overexpression of the (+)insert smooth muscle myosin heavy chain (SMMHC) isoform could contribute to airway bronchospasm by increasing the velocity of contraction. Whether the (+)insert isoform is present in the small airways and its expression is reversible in asthma are unknown. Objective: To determine the anatomical location and the expression kinetics of the (+)insert SMMHC isoform in airways of horses with heaves and to evaluate its modulation in response to disease status. Methods: We evaluated the (+)insert SMMHC isoform in the airways of horses with heaves during disease exacerbation a...
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...
Wilson AD.Horses are affected by a wide variety of arthropod ectoparasites, ranging from lice which spend their entire life on the host, through ticks which feed over a period of days, to numerous biting insects that only transiently visit the host to feed. The presence of ectoparasites elicits a number of host responses including innate inflammatory responses, adaptive immune reactions and altered behaviour; all of which can reduce the severity of the parasite burden. All of these different responses are linked through immune mechanisms mediated by mast cells and IgE antibodies which have an important ...
Brazil TJ, Dixon PM, Haslett C, Murray J, McGorum BC.The aim of this study was to characterise constitutive apoptosis in equine peripheral blood neutrophils, including assessment of factors that potentially modulate neutrophil survival through alteration of the rate of constitutive apoptosis. Cells underwent spontaneous time-dependent constitutive apoptosis when aged in culture for up to 36 h, developing the structural and functional features of apoptosis observed in many cell types, including human neutrophils. Neutrophils undergoing apoptosis also had diminished zymosan activated serum (ZAS)-stimulated chemiluminescence, but maintained respo...
Patan-Zugaj B, Gauff FC, Plendl J, Licka TF.To investigate effects of endotoxin on leukocyte activation and infiltration of the laminar tissue in isolated perfused equine limbs. Methods: 10 right forelimbs and 3 left forelimbs collected from 10 healthy adult horses after slaughter at a licensed abattoir. procedures: Isolated right forelimbs were randomly assigned to 2 groups (5 forelimbs/group): perfusion of the distal portion for 10 hours with 80 ng of endotoxin/L and perfusion under the same conditions without endotoxin. After perfusion, samples for immunohistochemical detection of leukocytes (by use of antibodies against calprotectin...
Yin X, Guo M, Gu Q, Wu X, Wei P, Wang X.Tetherin is an interferon-inducible host cell factor that blocks the viral particle release of the enveloped viruses. Most knowledge regarding the interaction between tetherin and viruses has been obtained using the primate lentiviral system. However, much less is known about the functional roles of tetherin on other lentiviruses. Equine infectious anemia virus (EIAV) is an important macrophage-tropic lentivirus that has been widely used as a practical model for investigating the evolution of the host-virus relationship. The host range of EIAV is reported to include all members of the Equidae ...
Boeer U, Buettner FF, Klingenberg M, Antonopoulos GC, Meyer H, Haverich A, Wilhelmi M.The limited biocompatibility of decellularized scaffolds is an ongoing challenge in tissue engineering. Here, we demonstrate the residual immunogenicity of an extensively decellularized equine carotid artery (dEAC(intens)) and identify the involved immunogenic components. EAC were submitted to an elaborated intensified decellularization protocol with SDS/sodium desoxycholate for 72 h using increased processing volumes (dEAC(intens)), and compared to dEAC(ord) prepared by an ordinary protocol (40 h, normal volumes). Matrix integrity was checked via correlative volumetric visualization which rev...
Dubuc J, Lavoie JP.The objective of this study was to determine whether eosinophils infiltrate the airway wall of horses with heaves. Eosinophils were evaluated using paraffin embedded lung tissues from six heaves-affected horses in crisis and six aged-matched controls. Slides were stained using Luna's method and eosinophils enumerated using histomorphometric techniques. Total eosinophil counts (expressed per mm(2) of basement membrane) were significantly higher in the airways of controls horses than in horses with heaves. Intraluminal, intraepithelial, and airway smooth muscle eosinophils counts were also incre...
Fouché N, Graubner C, Howard J.Various assays have been used as an aid to diagnose failure of passive transfer (FPT) of immunoglobulins in neonatal foals, but often lack sensitivity as screening tests, or are time consuming to perform and impractical as confirmatory tests. The aim of the present study was to evaluate whether measurement of serum total globulins (TG; i.e. total protein minus albumin) can be used to estimate the electrophoretic gamma globulin (EGG) fraction in hospitalised neonatal foals with suspected FPT. Sample data from 56 foals were evaluated retrospectively. The coefficient of rank correlation was 0.84....
Sanz MG, Oliveira AF, Page A, Horohov DW.Rhodococcus equi is the most common cause of pneumonia in young foals. A vaccine is not available and the use of R equi-specific hyperimmune plasma (HIP) is common. Despite its widespread use, the efficacy of HIP in preventing disease remains controversial. The objectives of this study were (1) to evaluate the virulence associate protein A (VapA)-specific IgG and IgG subclasses in commercially available R equi HIP and (2) to evaluate serum VapA-specific IgG and IgG subclasses in foals following administration of commercial R equi HIP. Three different lots from four commercial R equi HIP were s...
Liu C, Cook SJ, Craigo JK, Cook FR, Issel CJ, Montelaro RC, Horohov DW.Unlike other lentiviruses, EIAV replication can be controlled in most infected horses leading to an inapparent carrier state free of overt clinical signs which lasts for many years. While the resolution of the initial infection is correlated with the appearance of virus specific cellular immune responses, the precise immune mechanisms responsible for control of the infection are not yet identified. Since the virus undergoes rapid mutation following infection, the immune response must also adapt to meet this challenge. We hypothesize that this adaptation involves peptide-specific recognition sh...
Ma J, Wang SS, Lin YZ, Liu HF, Liu Q, Wei HM, Wang XF, Wang YH, Du C, Kong XG, Zhou JH, Wang X.The Chinese attenuated equine infectious anemia virus (EIAV) vaccine has successfully protected millions of equine animals from EIA disease in China. Given that the induction of immune protection results from the interactions between viruses and hosts, a better understanding of the characteristics of vaccine strain infection and host responses would be useful for elucidating the mechanism of the induction of immune protection by the Chinese attenuated EIAV strain. In this study, we demonstrate in equine monocyte-derived macrophages (eMDM) that EIAVFDDV13, a Chinese attenuated EIAV strain, indu...
Di Gennaro A, Lorusso A, Casaccia C, Conte A, Monaco F, Savini G.A serum neutralization assay (SN) was compared with the official plaque reduction neutralization test for the quantitation of West Nile virus antibodies. A total of 1,348 samples from equid sera and 38 from human sera were tested by these two methods. Statistically significant differences were not observed, thus supporting the use of SN for routine purposes.
Carmichael RC, Duell JR, Holbrook TC, Herrin BH, Leutenegger CM, O'Connor TP, Little SE.Abstract Tick infestations and infection with tick-borne agents are commonly recognized in horses in North America, but equine infection with true Ehrlichia spp. has not been described. To determine the degree to which horses in the south-central United States are naturally exposed to and infected with tick-borne disease agents, serum samples were collected at random (n=240) or from horses with active tick infestations (n=73) and tested by immunofluorescence antibody assay (IFA) and/or enzyme-linked immunosorbent assay (ELISA) for evidence of antibodies reactive to Ehrlichia spp., Anaplasma sp...
Hemphill DD, McIlwraith CW, Samulski RJ, Goodrich LR.Adeno-associated virus (AAV) receptors range from heparan sulfate proteoglycan to sialic acid moieties present on cell surfaces. Abundance of the glycan profiles is greatly influenced by animal species, cell type, and culture conditions. The objective of this study was to determine whether AAV serotypes' transduction efficiencies specifically in the equine monolayer culture model are an accurate representation of transduction efficiencies in tissue explants, a model more closely related to in vivo transduction. It was found that AAV 2 and 2.5 transduced cells more efficiently in explants than ...
Ons E, Van Brussel L, Lane S, King V, Cullinane A, Kenna R, Lyons P, Hammond TA, Salt J, Raue R.The efficacy of Zylexis®, an immunomodulator in horses based on inactivated Parapoxvirus ovis (iPPVO), was assessed using an equine herpesvirus type 1 (EHV-1) challenge model in the presence of a natural infection with Streptococcus equi equi (S. equi). Eleven horses were treated with iPPVO and twelve were kept as controls. Six horses were challenged with EHV-1 and commingled with the horses on study. Animals were dosed on Days -2, 0 (just before commingling) and Day 7. On Day 11 significantly less nasal discharge, enlarged lymph nodes, EHV-1 shedding and lower rectal temperatures were observ...
Manuja BK, Manuja A, Dahiya R, Singh S, Sharma RC, Gahlot SK.Equine influenza (EI) is primarily an infection of the upper respiratory tract and is one of the major infectious respiratory diseases of economic importance in equines. Re-emergence of the disease, species jumping by H3N8 virus in canines and possible threat of human pandemic due to the unpredictable nature of the virus have necessitated research on devising strategies for preventing the disease. The myxovirus resistance protein (Mx) has been reported to confer resistance to Orthomyxo virus infection by modifying cellular functions needed along the viral replication pathway. Polymorphisms and...
Manuja A, Kumar P, Kumar R, Kumar B, Singha H, Sharma RK, Yadav SC.Trypanosoma evansi is the causative agent of surra, which is the most common and widespread trypansomal disease. The infection is mainly restricted to animals, but it has also been documented in human. Trypanosomes possess the thick immunogenic surface coat known as variant surface glycoprotein (VSG). The parasite modifies the VSG constantly resulting in continuous antigenic variations and thus evades the host immune response. Due to antigenic variations, vaccination against trypanosomosis is not useful. Therefore, alternate strategies to augment the immune response are required. CpG-ODN class...
Khrustaleva TA, Khrustalev VV, Barkovsky EV, Kolodkina VL, Astapov AA.The SF23 peptide corresponding to the receptor binding fragment of diphtheria toxin (residues 508-530) has been synthesized. This fragment forming a protruding beta hairpin has been chosen because it is the less mutable B-cell epitope. Affine chromatography and ELISA show that antibodies from the sera of persons infected by toxigenic Corynebacterium diphtheriae and those immunized by diphtheria toxoid are able to bind the synthetic SF23 peptide. There are antibodies recognizing the SF23 peptide in the serum of horses hyperimmunized with diphtheria toxoid. Analysis of circular dichroism spectra...
Kanai Y, van Rijn PA, Maris-Veldhuis M, Kaname Y, Athmaram TN, Roy P.African horse sickness (AHS) is an equine disease with a mortality of up to 90% for susceptible horses. The causative agent AHS virus (AHSV) is transmitted by species of Culicoides. AHSV serogroup within the genus Orbivirus of the Reoviridae family consists of nine serotypes that show no or very limited cross-neutralization. Of the seven structural proteins (VP1-VP7) of AHSV, VP2 is the serotype specific protein, and the major target for neutralizing antibodies. In this report, recombinant VP2 proteins of all nine serotypes were expressed individually by the baculovirus expression system and t...
Galvão A, Tramontano A, Rebordão MR, Amaral A, Bravo PP, Szóstek A, Skarzynski D, Mollo A, Ferreira-Dias G.Metabolic hormones have been associated with reproductive function modulation. Thus, the aim of this study was: (i) to characterize the immunolocalization, mRNA and protein levels of leptin (LEP), Ghrelin (GHR) and respective receptors LEPR and Ghr-R1A, throughout luteal phase; and (ii) to evaluate the role of LEP and GHR on progesterone (P4), prostaglandin (PG) E2 and PGF2α , nitric oxide (nitrite), tumor necrosis factor-α (TNF); macrophage migration inhibitory factor (MIF) secretion, and on angiogenic activity (BAEC proliferation), in equine corpus luteum (CL) from early and mid-luteal sta...
Collins PJ, Vachieri SG, Haire LF, Ogrodowicz RW, Martin SR, Walker PA, Xiong X, Gamblin SJ, Skehel JJ.In 2004 an hemagglutinin 3 neuraminidase 8 (H3N8) equine influenza virus was transmitted from horses to dogs in Florida and subsequently spread throughout the United States and to Europe. To understand the molecular basis of changes in the antigenicity of H3 hemagglutinins (HAs) that have occurred during virus evolution in horses, and to investigate the role of HA in the equine to canine cross-species transfer, we used X-ray crystallography to determine the structures of the HAs from two antigenically distinct equine viruses and from a canine virus. Structurally all three are very similar with...
Wilkołek PM, Pomorski ZJ, Szczepanik MP, Adamek L, Pluta M, Taszkun I, Gołyński M, Rozwód A, Sitkowski W.The present study was designed to asses specific IgE towards environment allergens in 42 healthy horses. Determination of this immunoglobulin in serum serve as diagnostic tools in allergic diseases to improve efficacy of the treatment and proper allergen selection to specific immunotherapy. Serum levels of allergen specific IgE were measured with equine monoclonal antibody, using 15 individual and 5 mix allergens in North European Panel. The study revealed season dependent increased levels of allergen specific IgE in normal horses. It is noteworthy that healthy horses show high percentage of p...
Bailey GD, Love DN.Two hundred and seventy bacterial isolates were obtained from the pharyngeal tonsillar surface of 12 normal horses and 98 obligatory anaerobic bacteria were characterised. Of these, 57 isolates belonging to 7 genera (Peptostreptococcus (1); Eubacterium (9); Clostridium (6); Veillonella (6); Megasphera (1); Bacteroides (28); Fusobacterium (6)) were identified, and 16 of these were identified to species level (P. anaerobius (1); E. fossor (9); C. villosum (1); B. fragilis (1); B. tectum (2); B. heparinolyticus (2)). Three hundred and twenty isolates were obtained from 23 samples from horses with...
Fujimiya Y, Perryman LE, Crawford TB.Antibody-dependent cellular cytotoxicity and direct cytotoxicity assays were performed with equine infectious anemia virus-infected target cells, equine leukocytes, and equine anti-equine infectious anemia virus antibody to determine whether these mechanisms play a role in controlling viral replication in equine infectious anemia. Direct cytotoxicity was observed by using peripheral blood mononuclear cells from 7 of 10 infected horses. Antibody-dependent cellular cytotoxicity was not observed. The antibody-dependent cellular cytotoxicity reaction in horses was then studied by using sheep eryth...
Mencke N, Larsen KS, Eydal M, Sigurethsson H.Lice infestations in horses caused by the chewing louse Werneckiella (Damalinia) equi are observed worldwide. In the present study, the efficacy of 10% imidacloprid was tested on horses naturally infested with lice. Two groups of animals received a double application of 4 ml and 8 ml Advantage 10% spot-on on day 0 and 28 either. Horses, presenting dermatological signs with negative lice counts, were also included in this investigation. 40.86% of the horses presented positive lice counts and 84.21% of these animals showed clinical dermatologic signs. 65.45% of the lice-negative horses also show...
Hauck SM, Lepper MF, Hertl M, Sekundo W, Deeg CA.Equine recurrent uveitis is the only spontaneous model for recurrent autoimmune uveitis in humans, where T cells target retinal proteins. Differences between normal and autoaggressive lymphocytes were identified in this study by analyzing peripheral blood derived lymphocytes (PBL) proteomes from the same case with interphotoreceptor retinoid binding protein induced uveitis sampled before (Day 0), during (Day 15), and after uveitic attack (Day 23). Relative protein abundances of PBL were investigated in a quantitative, label-free differential proteome analysis in cells that were kept frozen for...
Miglio A, Morelli C, Gialletti R, Lauteri E, Sforna M, Marenzoni ML, Antognoni MT.The clinical, histological, and immunophenotypic findings are presented for 4 horses affected by different types of lymphoma. Diagnoses of a monomorphic epitheliotropic intestinal T-cell lymphoma, a diffuse splenic large B-cell lymphoma, a peripheral T-cell lymphoma, and a T-cell rich large B-cell lymphoma of the third eyelid were made. Constatations cliniques et immunophénotypiques pour quatre formes de lymphomes équins. Les constatations cliniques, histologiques et immunophénotypiques sont présentées pour quatre chevaux affectés par différents types de lymphome. Des diagnostics d’un...
Braverman Y, Ungar-Waron H, Frith K, Adler H, Danieli Y, Baker KP, Quinn PJ.A survey of sweet itch in horses in Israel based on a questionnaire to owners reported that 158 of 723 horses (21.8 per cent) had sweet itch lesions. The results indicated that the likelihood of a horse acquiring sweet itch decreased with increasing altitude but no definite association with rainfall zones was evident. Variation in the density of the horse population, however, obscured these observations. In the population surveyed, stallions were more sensitive than mares and pale horses appeared to be less sensitive than dark ones, but the sample size of this latter group was much smaller. In...
Hietala SK, Ardans AA.This study was undertaken to examine the neutrophil response to Corynebacterium (Rhodococcus) equi, and to assess the possibility of neutrophil immaturity or malfunction in predisposition to C. equi pneumonia in foals. Neutrophil phagocytosis of Corynebacterium (Rhodococcus) equi was studied in foals from birth to 6 months of age. Chemiluminescence (CL) and bactericidal assays were used to assay the phagocytic response of peripheral blood neutrophils to C. equi in vitro. Results of in vitro bactericidal and CL assays indicate that foal neutrophils are able to ingest and kill C. equi, however a...
Hamza E, Akdis CA, Wagner B, Steinbach F, Marti E.Insect bite hypersensitivity (IBH) is a recurrent allergic dermatitis of horses with similarities to human atopic eczema, caused by bites of insects of the genus Culicoides. Previous studies suggested a dysregulated T cell tolerance to Culicoides allergen in IBH-affected horses. Objective: We have investigated whether the suppressive function of CD4(+) CD25(high) cells is impaired in IBH-affected horses and possible ways to restore it. Methods: CD4(+) CD25(-) cells sorted from peripheral blood mononuclear cells (PBMC) were stimulated with irradiated autologous PBMC pulsed with Culicoides or te...
Parbhakar OP, Duke T, Townsend HG, Singh B.Pulmonary intravascular macrophages (PIMs) are present in horses and are believed to increase their sensitivity to endotoxin-induced cardio-pulmonary shock. However, owing to a lack of a marker for PIMs and the inability to isolate them, their precise contributions in the horse remain unknown. We designed this study to identify an immuno-phenotypic marker for PIMs and to develop a protocol for their transient depletion with gadolinium chloride (GC). GC is a lanthanide that has been used to deplete liver and lung macrophages. The horses (N = 15) were divided into control (n = 5) and GC-treated ...
Konishi E, Shoda M, Kondo T.Japanese encephalitis virus (JEV) causes fatal diseases in equines as well as humans. In Japan, racehorses are vaccinated with inactivated JE vaccine every year and no equine JE cases have been reported since 1986. However, the current reduction in JEV activity in nature has raised an argument against the requirement of continuous vaccination. Here, we studied natural infection rates in racehorses to address the issue. To identify naturally-infected individuals from vaccinated populations, we used an immunostaining method for detecting antibodies to JEV nonstructural 1 (NS1) protein. A total o...
Shao H, Robek MD, Threadgill DS, Mankowski LS, Cameron CE, Fuller FJ, Payne SL.The macrophage tropic lentivirus, equine infectious anemia virus (EIAV), encodes a dUTPase in the pol gene that is required for efficient replication in macrophages. Two naturally occurring variants of the enzyme were expressed as recombinant proteins in Escherichia coli; metal chelate affinity chromatography was used to purify histidine-tagged recombinant enzymes to greater than 80% homogeneity in a single chromatographic step. Biochemical and enzymatic analyses of these preparations suggest that this method yields dUTPase that is suitable for detailed mutational analysis. Specific activities...
Murray MJ, Eichorn ES, Dubovi EJ, Ley WB, Cavey DM.Whole blood and serum were collected from foals to determine the prevalence of Equine herpesvirus type 2 (EHV 2) infection in foals, age at which infection can first be identified and serological responses to infection. Equine herpesvirus type 2 was isolated from peripheral blood mononuclear cells (PBMC) from 68 of 69 foals, 1-8-months-old, sampled once. Virus isolation was performed twice at intervals of 2-7 months on PBMCs from 33 foals and EHV2 was isolated on both occasions in all but one foal (negative, then positive). Regression analysis of log2-transformed reciprocal serum EHV2 virus ne...
Anderson SL, Singh B.Horses that develop colitis invariably exhibit signs of a systemic inflammatory response syndrome (SIRS). A significant contributor to the development of SIRS in human subjects is delayed neutrophil apoptosis, but this has not been specifically studied in horses. Objective: To determine the occurrence of ex vivo neutrophil apoptosis and its contribution to the development of SIRS in an equine colitis model. Methods: Experiment using a colitis model. Methods: Neutrophils were isolated before and after the induction of colitis using an oligofructose overdose model, placed into culture for 12 h o...
Nolan MB, Schulman ML, Botha AE, Human AM, Roth R, Crampton MC, Bertschinger HJ.Commercial and regulatory limitations associated with native porcine zona pellucida (pZP) vaccines formulated with Freund's adjuvants may be overcome by developing effective recombinant ZP vaccines (reZP) and identifying alternative adjuvant formulations. A two-part preparatory study used 15 geldings and identified potentially effective alternative adjuvant formulations based on anti-pZP antibody response following treatment with pZP formulated with Addavax (AddaVax ™, Invivogen), Quil A (Quil-A® Adjuvant, Invivogen), Quil A and Poly (I:C) (HMW VacciGrade™, Invivogen), Pet Gel A (Montanid...
El-Sheikh Ali H, Legacki EL, Loux SC, Esteller-Vico A, Dini P, Scoggin KE, Conley AJ, Stanley SD, Ball BA.The current study aimed to elucidate the mechanisms underlying myometrial activation during equine placentitis related to progestogens and the progesterone receptor signaling pathways. Placentitis was induced via intracervical inoculation with Streptococcus equi ssp zooepidemicus in mares at approximately 290 days of gestation (placentitis group; n = 6) with uninoculated gestationally matched mares as controls (n = 4). Mares in the placentitis and control groups were euthanized, and myometrial samples were collected from two regions: region 1-parallel to active placentitis lesion with plac...
Hu Z, Guo K, Du C, Sun J, Naletoski I, Chu X, Lin Y, Wang X, Barrandeguy M, Samuel M, Wang W, Lau PI, Wernery U, Raghavan R, Wang X.Equine infectious anemia (EIA) is an important viral disease characterized by persistent infection in equids worldwide. Most EIA cases are life-long virus carriers with low antibody reactions and without the appearance of clinical symptoms. A serological test with high sensitivity and specificity is required to detect inapparent infection. In this study, a B-cell common epitope-based blocking ELISA (bELISA) was developed using a monoclonal antibody together with the EIAV p26 protein labelled with HRP. The test has been evaluated against the standard and with field serum samples globally. This ...
Timoney JF, Yang J, Liu J, Merant C.Streptococcus equi (S. equi) causes equine strangles, a highly contagious and widespread purulent lymphadenitis of the head and neck. Highly resistant to phagocytosis, it produces long extracellular chains in affected lymph nodes. In a screen of clones reactive with convalescent serum from a gene library of S. equi CF32 we identified IdeE, an IgG-endopeptidase and homologue of the leucocyte receptor Mac-1 (CD11b). IdeE is expressed during S. equi infection eliciting both serum and mucosal antibody responses which persisted at significant levels in serum for over 200 days. Release from S. equi ...
Smith DJ, Iqbal J, Purewal A, Hamblin AS, Edington N.IL-2 and equine chorionic gonadotrophin (eCG) initiated reactivation of equid herpesvirus-1 (EHV-1) from venous lymphocytes at a frequency of 1/10(-5). Indirect immunofluorescence showed that > 80% of virus-positive leukocytes were CD5+/CD8+ with the remaining 20% being CD5+/CD8-/CD4-. Cocultivation demonstrated that the reactivated virus was infectious. In addition, virus was reactivated in vitro from leukocytes of > 70% of horses by the mitogens phytohaemagglutinin (PHA) and pokeweed mitogen (PWM). Transfer of supernatants showed that IL-2 and eCG acted indirectly by causing the releas...
Azab W, Zajic L, Osterrieder N.Equine herpesvirus type 1 and 4 (EHV-1 and EHV-4) glycoprotein H (gH) has been hypothesized to play a role in direct fusion of the virus envelope with cellular membranes. To investigate gH's role in infection, an EHV-1 mutant lacking gH was created and the gH genes were exchanged between EHV-1 and EHV-4 to determine if gH affects cellular entry and/or host range. In addition, a serine-aspartic acid-isoleucine (SDI) integrin-binding motif present in EHV-1 gH was mutated as it was presumed important in cell entry mediated by binding to α4β1 or α4β7 integrins. We here document that gH is esse...
Benbarek H, Deby-Dupont G, Deby C, Caudron I, Mathy-Hartert M, Lamy M, Serteyn D.The activation of human polymorphonuclear leucocytes (the respiratory burst) can be studied by measuring their chemiluminescent response. This technique was adapted to equine leucocytes to investigate the effects of cell number, activator concentration, enhancers of chemiluminescence, pH, temperature and inhibitors. Leucocytes were isolated from citrated blood from healthy horses and chemiluminescence was measured with a Bio-Orbit luminometer sensitive to 900 nm light. The optimal cell density for the maximal chemiluminescent response ranged from 10(6) to 10(7) leucocytes 600 microliters-1. Ch...
Kinsley R, Scott SD, Daly JM.Serological assays provide an indirect route for the recognition of infectious agents via the detection of antibodies against the infectious agent of interest within serum. Serological assays for equine influenza A virus can be applied for different purposes: diagnosing infections; subtyping isolates; surveillance of circulating strains; and to evaluate the efficacy of vaccines before they reach the market. Haemagglutination inhibition (HI) and single radial haemolysis (SRH) assays are most commonly used in the equine field. This review outlines how both these assays together with virus neutra...
Yamanaka T, Nemoto M, Bannai H, Tsujimura K, Kondo T, Matsumura T, Gildea S, Cullinane A.We previously reported that horse antiserum against the Japanese equine influenza vaccine virus, A/equine/La Plata/1993 (LP93) exhibited reduced cross-neutralization against some Florida sublineage Clade (Fc) 2 viruses, for example, A/equine/Carlow/2011 (CL11). As a result, Japanese vaccine manufacturers will replace LP93 with A/equine/Yokohama/aq13/2010 (Y10, Fc2). To assess the benefit of updating the vaccine, five horses vaccinated with inactivated Y10 vaccine and five vaccinated with inactivated LP93 were challenged by exposure to a nebulized aerosol of CL11. The durations of pyrexia (≥3...
Schnabel CL, Fletemeyer B, Lübke S, Marti E, Wagner B, Alber G.Despite the relevance of adaptive immunity against equine pathogens antigen-specific T cell responses of horses are not well characterized and the lack of insight into T cell responses hampers the understanding of the pathogeneses of important diseases. In this study we used tetanus toxoid (TT) as a well-defined antigen to characterize antigen-reactive T cells. Six healthy adult horses received a routine booster against tetanus with an immune stimulating complex (ISCOM)-based vaccine and were followed for 28 days. TT-specific serum antibodies were quantified by ELISA and increased in all horse...
Wise LN, Kappmeyer LS, Knowles DP, White SN.The equine parasite Theilera equi continues to curtail global equine commerce due primarily to its ability to persist indefinitely in the immunocompetent horse. Details regarding the parasite life cycle, pathogenesis and mechanism of persistence remain unclear. The recently discovered T. haneyi is also capable of persistence in the horse, creating a potential reservoir for additional infections. These two divergent parasites share a unique gene family that expresses surface merozoite antigens, or equi merozoite antigens (EMAs). The EMA family was maintained in number and size in both parasites...
Kumar S, Malik P, Verma SK, Pal V, Gautam V, Mukhopadhyay C, Rai GP.Glanders, caused by the Gram-negative, nonmotile bacterium Burkholderia mallei, is a contagious and highly fatal disease of equines. During the last decade, the number of glanders outbreaks has increased steadily. The disease also has high zoonotic significance and B. mallei is listed biological warfare agent. The complement fixation test (CFT) is a routinely used and internationally recognized test to screen equine sera for the glanders. However, discrepant results have been observed using the CFT. The low sensitivity and specificity of the CFT and enzyme-linked immunosorbent assay (ELISA) ha...
Karàdi I, Kostner GM, Gries A, Nimpf J, Romics L, Malle E.Earlier studies demonstrated that lipoprotein (a), a lipoprotein of high atherogenicity, possesses proteolytic activity. In this report, we provide evidence that the lipoprotein (a)-specific antigen, apoprotein (a) is immunochemically related to plasminogen. This was demonstrated by polyclonal antisera from rabbit, sheep and horse, and with three monoclonal antibodies from mouse. Using immunospecific adsorbers against lipoprotein (a), all plasminogen could be adsorbed from lipoprotein (a)-positive and apparently lipoprotein (a)-negative plasma. As an additional similarity to plasminogen, lipop...
Lind C, Chen J, Byrjalsen I.We developed a rapid, competitive enzyme immunoassay (EIA) for measuring 25-hydroxyvitamin D3 [25(OH)D3] in serum. The EIA was based upon 25(OH)D3-3-hemisuccinate covalently coupled to secondary amino groups grafted onto the polystyrene surface of microtiter wells. Optimal coupling conditions were established, and we found that inclusion of 40 mumol/L chloramine T, an agent not previously described for use in coupling to these plates, resulted in both more reproducible coupling as well as more than a twofold increase in the coupling efficiency. Before EIA, 25(OH)D3 was extracted from the serum...
Bottoms GD, Johnson MA, Lamar CH, Fessler JF, Turek JJ.Dispersed equine vascular endothelial cells grown in tissue culture, and freshly isolated neutrophils were used to determine direct effects of endotoxin on cyclooxygenase and lipoxygenase products. Endothelial cells (10(7)/ml) or neutrophils (2 X 10(6)/ml) were incubated with (a) buffer, (b) endotoxin (10 micrograms/ml), (c) endotoxin + flunixin meglumine (10 micrograms/ml), or (d) calcium ionophore, A23187 (10 micrograms/ml). Thromboxane (TxB2), prostacyclin (6-keto-PGF1 alpha), and leukotriene C4 (LTC4) were determined in the incubation fluid by radioimmunoassay. Thromboxane and prostacyclin...
Chambers TM, Shortridge KF, Li PH, Powell DG, Watkins KL.The Directigen FLU-A enzyme immunoassay was tested for its ability to detect equine-2 influenza viruses in nasopharyngeal fluids from horses and ponies. A total of 125 swabs from experimental infections and from different sources of natural infection in the USA and Hong Kong were examined. The assay results were compared with the results of standard virus culture in embryonated chicken eggs or Madin-Darby canine kidney cells, and with the serology of the horses sampled. In comparison with virus culture the enzyme immunoassay exhibited 83 per cent sensitivity, 78 per cent specificity, 70 per ce...
Bullido R, Doménech N, Alvarez B, Alonso F, Babín M, Ezquerra A, Ortuño E, Domínguez J.A set of five monoclonal antibodies (mAb) against porcine major histocompatibility complex (MHC), or swine leukocyte antigens (SLA), class II molecules has been characterized. These mAbs appear to recognize monomorphic determinants on SLA-DR (2F4, 1F12 and 2E9/13) and SLA-DQ (BL2H5 and BL4H2) molecules, as assessed by flow cytometry and immunoprecipitation. By Western blot, the 2F4, 1F12, BL2H5 and BL4H2 epitopes were located on the beta-chains of these molecules. mAbs 2F4 and 1F12 crossreact with leucocytes of dog, cattle, horse and human; mAbs 2E9/13, BL2H5 and BL4H2 bind leucocytes of cattl...
