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.
Kikuchi M, Murase H, Urata K, Ishige T, Nagata SI, Tozaki T, Kakoi H, Ishiguro-Oonuma T, Kizaki K.Circulating microRNAs (miRNAs) are stable in body fluids and can serve as biomarkers for various diseases and physiological states. Although pregnancy-related miRNAs have been identified in various mammals, studies on parturition-related circulating miRNAs in mares are limited. Therefore, this study aimed to identify parturition-related miRNAs and examine their potential applications in the prediction of parturition date. miRNAs were extracted from the plasma of Thoroughbred mares 30 days (295-326 days pregnant) and 5 (323-352 days pregnant) - 0 (328-357 days pregnant) days before parturition,...
Fukuda K, Mita H, Kuroda T, Tamura N, Kuwano A, Sato F, Takahashi T.To clarify the anti-inflammatory effect of platelet lysate (PL) on equine persistent synovitis by using a model of synovitis induced by monoiodoacetic acid (MIA). Methods: Nonseptic synovitis was induced by administering MIA into both antebrachiocarpal joints of 6 clinically healthy horses on day 0. On days 23, 30, and 37, carpal circumference measurement and synovial fluid collection for assays (leucocytes, LDH, tumor necrosis factor-α, and TGF-β1) were performed, after which PL was injected into 1 antebrachiocarpal joint and saline into the contralateral joint. Synovium and synovial fluid ...
Hobbs KJ, Le Sueur ANV, Hallowell K, Martin E, Sheats MK, Ueda Y.An 8-year-old American Quarter Horse gelding was treated with extracorporeal hemoperfusion (HP) therapy for treatment of Clostridioides difficile (C. difficile) colitis-induced systemic inflammatory response syndrome (SIRS). The gelding developed C. difficile associated peracute colitis and severe SIRS as evidenced by a positive fecal C. difficile PCR and tachypnea, tachycardia, fever, neutropenia, altered mucous membrane color, and hyperlactatemia. Concurrent acute kidney injury in the horse limited the use of routine anti-inflammatory and anti-lipopolysaccharide treatments, including flunixi...
de Souza AF, Pereira CAM, Fürst A, Kümmerle JM, De Zoppa ALDV.The implantation of unicortical cortex screws in the proximal hole of locking compression plates (LCP) has been recommended for proximal interphalangeal (PIP) arthrodesis in horses to prevent fractures resulting from stress risers in the proximal phalanx (P1). However, this cortex screw fixation technique may limit efficient dorsal compression of the PIP joint by the plate, potentially affecting the stability of the construct. In this study, we aimed to measure stress and strain in P1 and the plate using an ex vivo model of PIP arthrodesis in horses. We employed various implantation methods an...
Bell J, Raidal S, Peters A, Hughes KJ.Management of diarrhoea in horses is usually non-specific and supportive. Faecal microbiota transplantations (FMT) are used to manage dysbiosis in horses with diarrhoea. There are few studies investigating the effects of storage on prepared FMT solutions. This study was an in vitro non-randomised controlled experiment that investigated the effects of FMT solution preparation and storage on the faecal microbiota. Fresh faeces were collected from five healthy adult horses and used for DNA extraction and preparation of FMT. From each FMT, seven aliquots were collected and DNA was extracted immedi...
Woo PCY, Mheiri FA, Cavalleri J, Joseph S, Tang JYM, Joseph M, Tsang CC, Lau SKP, Wernery U.Epizootic lymphangitis (EL) is a highly prevalent and contagious infectious disease affecting horses in many parts of Ethiopia caused by Histoplasma capsulatum sensu lato ('var. farciminosum'). In this study, 12 suspected isolates of H. capsulatum sensu lato or yeasts unidentified by conventional biochemical tests isolated from Ethiopian horses with EL were characterised by ITS sequencing. Six of the 12 isolates were identified to be members of H. capsulatum sensu lato and the other six were Pichia kudriavzevii (synonym: Candida krusei) (n = 3), Trichosporon asahii (n = 1), Geotrichum silv...
Richards S, Palmer D, Cawley A, Wainscott M, Keledjian J.An improved screening workflow and a robust capillary flow LC-MS confirmatory method for the detection of recombinant human erythropoietin (rHuEPO) has been implemented to increase the sensitivity of rHuEPO detection and to reduce the number of suspect samples committed to confirmatory testing. The influence of repeated dosing of epoetin-β on the detection window of rHuEPO in equine plasma was assessed using the optimised method. Samples were initially assessed using an economical R&D Human EPO Duo-Set ELISA Development System. Samples indicating a result greater than the batch baseline were ...
Hobbs KJ, Cooper BL, Dembek K, Sheats MK.Cell-free DNA (cfDNA) is fragmented extracellular DNA that is released from cells into various body fluids. Previously published data from adult horses supports cfDNA as a potential disease biomarker, but also shows that direct measurement in plasma is inaccurate due to matrix effect. It is currently unknown whether a similar matrix effect exists in foal plasma. Given this, the objectives of the current study were to investigate foal plasma for potential matrix effect during fluorescence measurement of cfDNA using a Qubit fluorometer, and to determine whether neat and/or extracted plasma cfDNA...
Chiara M, Mariaelena C, Alessandro C, Davide B, Lavinia C, Paola MM, Barbara L, Chiara DP, Flagiello F, Pia PM.The advanced-PRF+ (A-PRF+) is a platelet concentrate, showing a higher concentration of growth factors, an increased number of cells and looser structure of the fibrin clot than leukocyte-PRF. A high variability in the size of PRF associated with patients, haematological features and centrifugation protocols was reported. The aims of this study were to evaluate the feasibility of A-PRF+ production in the field and the correlation between haematological parameters, macroscopic and microscopic features in equine A-PRF+. Samples from twenty Standardbred horses (3-7 years) were harvested with gla...
Lomnicka I, Dubey S, Waller P, Vora D, Dirikolu L.The screening of drugs in plasma and urine often requires initial extraction (such as liquid-liquid extraction and solid-phase extraction) before the samples are submitted to instrumental analyses. These extraction procedures are often laborious and time-consuming. In this manuscript, a high-throughput automated assay based on liquid chromatography-high-resolution mass spectrometry (LC-HRMS) suitable for use as an initial testing procedure covering multiple classes of compounds prohibited in horse racing is described. The assay requires a 600-μL plasma aliquot, which is subjected to solid pha...
Shams F, Jokar M, Abdous A, Mohammadi P, Abbassioun A, Seuberlich T, Rahmanian V.Neospora spp. and Toxoplasma gondii are two closely related protozoan parasites that are widely distributed throughout the world. Horses can act as intermediate hosts for both parasites and can acquire disease. Blood samples were taken from 487 clinically healthy horses from 17 different mechanized stables in Tehran, the capital of Iran, during September and November of 2022. IFAT and ELISA were employed to detect antibodies directed against Neospora spp. and T. gondii. The anti-N. caninum antibodies were detected in 52 of the horses (10.67%) based on IFAT and in 86 of the 487 horses (17.65%) ...
Lacante SA, Jiang C, Mustamir AA, Mizuno T, Bi X, Syafruddin D, Tokoro M., a human intestinal protozoan parasite of the diplomonad group, has been overlooked because of its commensal features; therefore, molecular studies on this parasite are limited. To address this gap, we designed a molecular screening protocol using polymerase chain reaction (PCR) and DNA sequencing targeting the 18S small subunit ribosomal RNA gene and applied this screening method to the molecular epidemiological analysis of spp. in humans and various livestock. We validated our methodology using stool samples collected from 215 humans and 270 animal hosts (buffaloes, pigs, dogs, goats, hors...
Winther MF, Johnsson J, Madsen PK, Pihl TH, Paltrinieri S, Cerón JJ, Scavone D, Pardo-Marin L, Jacobsen S.Paraoxonase-1 (PON-1) has been suggested as a marker of inflammation and oxidative stress in horses and could potentially be used for prognostication in horses with colitis. Objective: Assessment of PON-1 in horses with colitis and comparison of two methods. Methods: Serum PON-1 was measured by two methods (paraoxon and p-nitrophenyl acetate) in 161 horses with colitis and 57 controls. Follow-up samples obtained during hospitalization were available from 106 horses with colitis. The two methods were compared. Results: Serum PON-1 was significantly lower in horses with colitis than in healthy h...
Diekmann I, Blazejak K, Krücken J, Strube C, von Samson-Himmelstjerna G.Surveillance of Strongylus vulgaris and other Strongylus spp. in equids is important for targeted intervention in parasite control, requiring reliable routine diagnostic methods. Objective: Comparing morphological examination and PCR analyses of larval cultures to identify Strongylus spp. species based on German diagnostic samples from 2018. Methods: Method comparison. Methods: During the routine diagnostic investigations, in total 712 strongyle-egg positive equine faecal samples were cultured. Third-stage larvae (L3) were morphologically differentiated. For molecular validation, samples were ...
Ing NH, Konganti K, Ghaffar N, Johnson CD, Forrest DW, Love CC, Varner DD.Males of some species, from horses to humans, require medical help for subfertility problems. There is an urgent need for novel molecular assays that reflect spermatozoal function. In the last 25 years, studies examined RNAs in spermatozoa as a window into gene expression during their development and, more recently, for their functions in early embryo development. In clinics, more dense spermatozoa are isolated by density gradient centrifugation before use in artificial insemination to increase pregnancy rates. The objectives of the current study were to discover and quantify the microRNAs i...
Gray B, Lubbock K, Love C, Ryder E, Hudson S, Scarth J.The analytical approaches taken by laboratories to implement robust and efficient regulation of horseracing medication and doping control are complex and constantly evolving. Each laboratory's approach will be dictated by differences in regulatory, economic and scientific drivers specific to their local environment. However, in general, laboratories will all be undertaking developments and improvements to their screening strategies in order to meet new and emerging threats as well as provide improved service to their customers. In this paper, the published analytical advances in horseracing me...
Pihl TH, Engelhart KE, Houen G.Polyclonal antibodies are relatively easy to produce and may supplement monoclonal antibodies for some applications or even have some advantages.The choice of species for production of (peptide) antisera is based on practical considerations, including availability of immunogen (vaccine) and animals. Two major factors govern the production of antisera: the nature of adaptive immune responses, which take place over days/weeks and ethical guidelines for animal welfare.Here, simple procedures for immunization of mice, rabbits, sheep, goats, pigs, horses, and chickens are presented.
Giraldo A, Koch TG, Madan P, Lepage S, Monteith G, Alizadeh AH, Tran A, Mortagy N, Koenig JB.There is a knowledge gap regarding the effect of extracorporeal shockwave treatment (ESWT) on the stress response and immunomodulatory and anti-inflammatory properties of equine umbilical cord blood mesenchymal stromal cells (CB-MSCs). The objective of this study was to investigate the presence of cellular oxidative stress, inflammatory response, and production of growth factors in CB-MSCs after treatment with ESWT. We hypothesized that CB-MSCs treated with ESWT will experience higher levels of cellular stress and increased production of anti-inflammatory cytokines and growth factors compared ...
Teles Filho ACA, Sanchez DJD, Viana AGA, Sheheryar S, Guerreiro DD, Bustamante-Filho IC, Martins AMA, Sousa MV, Ricart CAO, Fontes W, Moura AA.The present study was conducted to investigate the global proteome of 8-day-old equine blastocysts. Follicular dynamics of eight adult mares were monitored by ultrasonography and inseminated 24 h after the detection of a preovulatory follicle. Four expanded blastocysts were recovered, pooled, and subjected to protein extraction and mass spectrometry. Protein identification was conducted based on four database searches (PEAKS, Proteome Discoverer software, SearchGUI software, and PepExplorer). Enrichment analysis was performed using g:Profiler, Panther, and String platforms. After the elimina...
Maniego J, Harding C, Habershon-Butcher J, Hincks P, Stewart G, Proudman C, Ryder E.Due to the ease of collection, transport and storage, the use of dried blood spots (DBS) offers an attractive alternative matrix for detection of the abuse of gene therapy, otherwise known as gene doping. This study evaluated the recovery, extraction efficiency and resulting detection capability of DNA from DBS by evaluating different target types, DNA extraction kits, the number of punches and blood tube preservatives. The long-term storage stability of low-copy-number transgene targets in DBS was not assessed in this study but would be noteworthy to investigate further. DNA was quantified us...
Ramírez-Agámez L, Castaneda C, Hernández-Avilés C, Grahn RA, Raudsepp T, Love CC.Two methods for preimplantation genetic testing (PGT) have been described for equine embryos: trophoblast cell biopsy (TCB) or blastocoele fluid aspiration (BFA). While TCB is widely applied for both in vivo- and in vitro-produced embryos, BFA has been mostly utilized for in vivo-produced embryos. Alternative methods for PGT, including analysis of cell-free DNA (CFD) in the medium where in vitro-produced embryos are cultured, have been reported in humans but not for equine embryos. In Experiment 1, in vivo- (n = 10) and in vitro-produced (n = 13) equine embryos were subjected to BFA, cultu...
Long AE, Pitta D, Hennessy M, Indugu N, Vecchiarelli B, Luethy D, Aceto H, Hurcombe S.Currently, lack of standardization for fecal microbiota transplantation (FMT) in equine practice has resulted in highly variable techniques, and there is no data on the bacterial metabolic activity or viability of the administered product. The objectives of this study were to compare the total and potentially metabolically active bacterial populations in equine FMT, and assess the effect of different frozen storage times, buffers, and temperatures on an equine FMT product. Fresh feces collected from three healthy adult horses was subjected to different storage methods. This included different ...
Goodrich EL, Webb JL.Previous hematologic and serum biochemistry reference interval (RI) values have been established for donkeys in various geographic regions, life-stages, or for specific donkey breeds. The last extensive investigation establishing RIs for adult donkeys in the United States (U.S.) was published over three decades ago. We aimed to establish updated robust RIs using a reference population of apparently healthy adult donkeys from across the U.S. Standard sized ( = 102), miniature ( = 17), and mammoth ( = 1) donkeys from four different states were enrolled, with 20% of the study population including...
Segabinazzi LGTM, Dell'Aqua CPF, Cavalero T, Frasson M, Lisboa FP, Papa FO, Alvarenga MA.Poor sperm quality in cooled-shipped semen has been related to subpar fertility in horses. Therefore, this study aimed to evaluate the ability of post-cooling sperm processing to improve sperm parameters of cooled-stored stallion semen for artificial insemination. For all experiments, ejaculates were collected, processed, and diluted in skimmed milk-based (SM) medium and stored at 5 °C/24h. In all experiments an aliquot of unprocessed cooled semen was used as a control. In the first experiment (Exp 1.), cooled-stored semen from 16 stallions (n = 32) was processed by SpermFilter or centrifu...
Kushida C, Tamura N, Kasashima Y, Sato K, Arai K.In equine regenerative medicine using bone marrow-derived mesenchymal stem/stromal cells (BM-MSC), the importance of the quality management of BM-MSC has been widely recognized. However, there is little information concerning the relationship between cellular senescence and the stemness in equine BM-MSC. In this study, we showed that stemness markers (NANOG, OCT4, SOX2 and telomerase reverse transcriptase) and colony forming unit-fibroblast apparently decreased accompanied with incidence of senescence-associated β-galactosidase-positive cells by repeated passage. Additionally, we suggested th...
Maniego J, Harding C, Habershon-Butcher J, Hincks P, Ryder E.Gene doping, which includes the non-therapeutic use of genes or genetic elements that have the capacity to enhance athletic performance, is prohibited in horseracing and equestrian sports. To provide a comprehensive assessment of matrix and detection techniques, a custom adeno-associated virus serotype 8 vector was designed to include PCR binding sites for multiple target genes and assay types. The vector was injected via an intramuscular route into two Thoroughbred horses and matrices collected at defined timepoints. DNA was analysed using 3 detection methods: qPCR, digital PCR, and NGS. Over...
Kabi L E, Zor TA, Göktaş EF, Güneş E, Yatanaslan L, Tektaş MH, İnceman B, Tufan M.The use of prohibited substances in horse racing is a major concern that jeopardizes both the fairness of competitions and the health of horses. This problem can stem from the use of licensed drugs for animal health, as well as unlicensed substances. Horse doping laboratories monitor the potential use of these substances in racehorses within the framework of regulations set by the International Federation of Horse Racing Authority. In this context, sildenafil and its major metabolite n-desmethyl sildenafil were detected in a post-race horse urine sample sent to the Pendik Veterinary Control In...
Núñez-Montero K, Leal K, Rojas-Villalta D, Castro M, Larronde C, Wagenknecht L, Contreras MJ.Studies on the bacterial composition of seminal samples have primarily focused on species isolated from semen and their effects on fertility and reproductive health. Culture-independent techniques, such as 16S rRNA gene sequencing and shotgun metagenomics, have revolutionized our ability to identify unculturable bacteria, which comprise >90% of the microbiome. These techniques allow for comprehensive analysis of microbial communities in seminal samples, shedding light on their interactions and roles. In this study, we characterized the taxonomic diversity of seminal microbial communities in he...
Hisaeda K, Ono T, Kadekaru S, Hata A, Miyama TS, Kutara K, Sugimoto K, Hiasa Y, Ohzawa E, Kunieda T, Iwata E, Kitagawa H.Plasma or serum amino acids are used to evaluate nutritional status and metabolic disorders. In this study, we aimed to set reference values of serum amino acid concentrations in the Noma horse, a Japanese native horse. Thirty-one horses were classified into six age groups: neonatal foal (0-4 days), foal (0.5-1 years), youth (5 years), middle age (10 years), old (15 years), and extra-old (>20 years). Horses >5 years of age were analyzed together as the adult group. In the adult horses, there were no significant differences among the serum amino acid concentrations of each age group. The ...
Tallmadge RL, Lear TL, Antczak DF.The availability of a contig of bacterial artificial chromosome (BAC) clones spanning the equine major histocompatibility complex (MHC) made possible a detailed analysis of horse MHC class I genes. Prior to this study, only a single horse MHC class I gene had been sequenced at the genomic level. Although many ( approximately 60) MHC class I cDNA sequences had been determined and published, from this information, it was not possible to determine how many class I loci are expressed in horses or to assign individual sequences to allelic series. In this study, 15 MHC class I genes were identified ...
Rwambo PM, Issel CJ, Hussain KA, Montelaro RC.A neutralization escape mutant (A/1 E) of equine infectious anemia virus was isolated after 13 passages in cell culture in the presence of serum containing antibodies to type- and group-specific determinants of EIAV envelope glycoproteins. Loss of neutralization by the selecting serum correlated with loss of two epitopes in the major envelope glycoprotein gp90 of A/1 E which were present in a parallel variant isolated from a persistently infected pony.
Corradin G, Chiller JM.Murine T-lymphocyte specificity was determined in a system of antigen driven in vitro T-cell proliferation using cytochrome c molecules from different species, their derived peptides and reconstituted hybrid proteins. It was observed that primed T cells could discriminate between peptide fragments which differed from each other at a single amino acid residue. These conclusions were substantiated by the pattern of cross-reactivity noted in the response of closely related cytochrome c proteins as well as when artificial hybrid molecules reconstituted by the covalent linkage of peptide fragments ...
Schamberger GP, Diez-Gonzalez F.Escherichia coli strains were screened for their ability to inhibit E. coli O157:H7. An initial evaluation of 18 strains carrying previously characterized colicins determined that only colicin E7 inhibited all of the E. coli O157:H7 strains tested. A total of 540 strains that had recently been isolated from humans and nine different animal species (cats, cattle, chickens, deer, dogs, ducks, horses, pigs, and sheep) were tested by a flip-plating technique. Approximately 38% of these strains were found to inhibit noncolicinogenic E. coli K12 strains. The percentage of potentially colicinogenic E...
Balasuriya UB, Timoney PJ, McCollum WH, MacLachlan NJ.The variation and phylogenetic relationship of open reading frame 5 (ORF5) of 3 different laboratory strains of the original prototype Bucyrus strain of equine arteritis virus (EAV), the modified live virus vaccine (ARVAC, Fort Dodge Laboratories), and 18 field isolates of EAV from North America and Europe were determined by comparison of their gene sequences. The viruses differed from the published sequence by between 3 (99.6% homology) and 94 (87.8%) nucleotides and by between 3 (98.8%) and 24 (90.6%) amino acids. The field isolates differed from each other by between 2 (99.7%) and 110 (85.7...
Lazzari G, Colleoni S, Crotti G, Turini P, Fiorini G, Barandalla M, Landriscina L, Dolci G, Benedetti M, Duchi R, Galli C.Assisted reproduction technologies (ART) are well developed in humans and cattle and are gaining momentum also in the equine industry because of the fact that the mare does not respond to superovulation but can donate large numbers of oocytes through ovum pick up (OPU). After collection, the oocytes can be fertilized by intracytoplasmic sperm injection (ICSI) using a variety of stallion semen samples, even of poor quality, and the resulting embryos can establish high pregnancy rates after cryopreservation and transfer. The discoveries that equine oocytes can be held at room temperature without...
Alber J, El-Sayed A, Lämmler C, Hassan AA, Weiss R, Zschöck M.The closely related streptococcal species Streptococcus equi subsp. zooepidemicus and S. equi subsp. equi were identified by polymerase chain reaction using oligonucleotide primers designed according to species-specific parts of the superoxide dismutase A encoding gene sodA. A further differentiation of both subspecies could be performed by amplification of the genes seeH and seeI encoding the exotoxins SeeH and SeeI, respectively, which could be detected for S. equi subsp. equi but not for S. equi subsp. zooepidemicus. A further simplification of the identification and differentiation of both...
Swegen A, Grupen CG, Gibb Z, Baker MA, de Ruijter-Villani M, Smith ND, Stout TAE, Aitken RJ.Early pregnancy in the mare is a poorly understood, high risk period during which the embryo communicates its presence to the maternal endometrium. Remarkably, the maternal recognition of pregnancy signal is unknown in the horse. This study aimed to profile the proteins secreted by equine blastocysts into their immediate environment, along with proteins contained in the blastocoel and within the acellular embryo capsule. Embryos were recovered on day 8 after ovulation and cultured for 48 hours. Secretomes of day 9 and day 10 embryos were analyzed by LC-MS/MS and supported by analysis of blast...
Isgren CM, Edwards T, Pinchbeck GL, Winward E, Adams ER, Norton P, Timofte D, Maddox TW, Clegg PD, Williams NJ.This study investigated changes over time in the epidemiology of extended-spectrum β-lactamase (ESBL) producing Escherichia coli within a single equine referral hospital in the UK. Faecal samples were collected from hospitalised horses in 2008 and 2017, processed using selective media and standard susceptibility laboratory methods. A novel real-time PCR with high resolution melt analysis was used to distinguish bla and bla within CTX-M-1 group. Results: In 2008, 457 faecal samples from 103 horses were collected, with ESBL-producing E. coli identified in 131 samples (28.7, 95% CI 24.6-33.1). I...
Dakin SG, Smith RK, Heinegård D, Önnerfjord P, Khabut A, Dudhia J.During inflammatory processes the extracellular matrix (ECM) is extensively remodeled, and many of the constituent components are released as proteolytically cleaved fragments. These degradative processes are better documented for inflammatory joint diseases than tendinopathy even though the pathogenesis has many similarities. The aims of this study were to investigate the proteomic composition of injured tendons during early and late disease stages to identify disease-specific cleavage patterns of the ECM protein cartilage oligomeric matrix protein (COMP). In addition to characterizing fragme...
Tanhauser SM, Yowell CA, Cutler TJ, Greiner EC, MacKay RJ, Dame JB.Studies designed to investigate the causative agent of equine protozoal myeloencephalitis and its life cycle have been hampered by the marked similarity of Sarcocystis neurona to other Sarcocystis spp. present in the same definitive host. Random-amplified polymorphic DNA techniques were used to amplify DNA from isolates of S. neurona and Sarcocystis falcatula. DNA sequence analysis of polymerase chain reaction (PCR) products was then used to design PCR primers to amplify specific Sarcocystis spp. DNA products. The ribosomal RNA internal transcribed spacer was also amplified and compared betwee...
Martin G, Plowright R, Chen C, Kault D, Selleck P, Skerratt LF.Hendra virus (HeV) is lethal to humans and horses, and little is known about its epidemiology. Biosecurity restrictions impede advances, particularly on understanding pathways of transmission. Quantifying the environmental survival of HeV can be used for making decisions and to infer transmission pathways. We estimated HeV survival with a Weibull distribution and calculated parameters from data generated in laboratory experiments. HeV survival rates based on air temperatures 24 h after excretion ranged from 2 to 10 % in summer and from 12 to 33 % in winter. Simulated survival across the distri...
Young R, Taylor JE, Kurioka A, Becker M, Louis EJ, Rudenko G.African trypanosomes (including Trypanosoma brucei) are unicellular parasites which multiply in the mammalian bloodstream. T. brucei has about twenty telomeric bloodstream form Variant Surface Glycoprotein (VSG) expression sites (BESs), of which one is expressed at a time in a mutually exclusive fashion. BESs are polycistronic transcription units, containing a variety of families of expression site associated genes (ESAGs) in addition to the telomeric VSG. These polymorphic ESAG families are thought to play a role in parasite-host adaptation, and it has been proposed that ESAG diversity might ...
Sheoran AS, Lunn DP, Holmes MA.This paper describes the production of a panel of monoclonal antibodies (mAbs) identifying the four recognised equine IgG subisotypes IgG, IgGa, IgGb, IgGc and IgG(T). Pure preparations of the subisotypes for use in immunisations and testing were produced using a combination of gel filtration, salt precipitation, ion exchange chromatography and protein A and Protein G affinity chromatography. The specificity of mAbs for the IgG subisotypes was confirmed using ELISA assays, by characterisation of affinity purified proteins recognised by the mAbs, and by Western blotting of equine serum proteins...
Chapman GE, Sherlock K, Hesson JC, Blagrove MSC, Lycett GJ, Archer D, Solomon T, Baylis M.There has been no evidence of transmission of mosquito-borne arboviruses of equine or human health concern to date in the UK. However, in recent years there have been a number of outbreaks of viral diseases spread by vectors in Europe. These events, in conjunction with increasing rates of globalisation and climate change, have led to concern over the future risk of mosquito-borne viral disease outbreaks in northern Europe and have highlighted the importance of being prepared for potential disease outbreaks. Here we assess several UK mosquito species for their potential to transmit arboviruses ...
Dias de Castro LL, Abrahão CLH, Buzatti A, Molento MB, Bastianetto E, Rodrigues DS, Lopes LB, Silva MX, de Freitas MG, Conde MH, Borges FA.The aim of this study was to compare two fecal egg count (FEC) techniques; McMaster (McM) and Mini-FLOTAC (mF), for the detection of cattle and horse gastrointestinal nematode eggs, in different locations. Experiment 1: feces were collected from 16 cattle and FEC was performed individually, using mF with the sensitivity of 5 eggs per gram of feces (EPG) and McM with a sensitivity of 50 EPG at Empresa de Pesquisa Agropecuária de Minas Gerais - EPAMIG and the Laboratory of Parasitic Diseases of the University of Parana - LDP/UFPR. Experiment 2: Fecal samples from 30 horses were analyzed with mF...
Liu C, Guo W, Lu G, Xiang W, Wang X.A new strain of equine herpesvirus type 8 (EHV-8), Wh, has been isolated from horses in China, and its complete genome has been sequenced and analyzed. The result indicates that the new strain has the same constitution and arrangement of open read frames as EHV-1 and EHV-9. This work is the first announced complete genome sequence of EHV-8.
Martens A, De Moor A, Ducatelle R.The purpose of this study was to examine if bovine papilloma virus (BPV) DNA can be detected in superficial swabs or scrapings from equine sarcoids. Samples were obtained from 92 sarcoids and 20 non-sarcoidal control lesions. The polymerase chain reaction technique was used with a first primer set to check whether DNA extraction was successful, and with a second primer set specific for BPV-DNA. DNA isolation was successful in 88% of the swabs and 93% of the scrapings. All control lesions were negative for BPV-DNA.
Todhunter PG, Kincaid SA, Todhunter RJ, Kammermann JR, Johnstone B, Baird AN, Hanson RR, Wright JM, Lin HC, Purohit RC.To use lipopolysaccharide (LPS) to create synovitis in the midcarpal joint of ponies, and to assess the morphologic, histochemical, and immunohistochemical effects of synovitis on articular cartilage of the third carpal bone. Methods: 2- to 3-year-old ponies, 6 control (group 1) and 6 treated (group 2). Methods: Synovitis was induced in 1 midcarpal joint of group-2 ponies by intra-articular injections of LPS (0.02 micrograms/kg of body weight), morphine (0.1 mg/kg), and saline solution (group 2a) and a morphine and saline solution alone in the contralateral midcarpal joint (group 2b). Articula...
Zwart L, Potgieter CA, Clift SJ, van Staden V.African horse sickness is a serious equid disease caused by the orbivirus African horse sickness virus (AHSV). The virus has ten double-stranded RNA genome segments encoding seven structural and three non-structural proteins. Recently, an additional protein was predicted to be encoded by genome segment 9 (Seg-9), which also encodes VP6, of most orbiviruses. This has since been confirmed in bluetongue virus and Great Island virus, and the non-structural protein was named NS4. In this study, in silico analysis of AHSV Seg-9 sequences revealed the existence of two main types of AHSV NS4, designat...
Li F, Leroux C, Craigo JK, Cook SJ, Issel CJ, Montelaro RC.Equine infectious anemia virus (EIAV) is genetically one of the simplest lentiviruses in that the viral genome encodes only three accessory genes, tat, rev, and S2. Although serological analyses demonstrate the expression of the S2 protein in persistently infected horses, the role of this viral gene remains undefined. We recently reported that the S2 gene is not essential for EIAV replication in primary equine macrophages, as EIAV mutants lacking the S2 gene replicate to levels similar to those of the parental virus (F. Li, B. A. Puffer, and R. C. Montelaro, J. Virol. 72:8344-8348, 1998). We n...
Carslake HB, Pinchbeck GL, McGowan CM.Many diagnostic tests for insulin dysregulation use reference intervals established with an insulin radioimmunoassay (RIA) that is no longer available. A chemiluminescent immunoassay (CLIA) is commonly used for the measurement of serum insulin concentration in clinical practice but requires further validation, especially at clinically relevant reference intervals. Objective: To evaluate the CLIA for measurement of equine insulin and compare it to the previously validated, but now unavailable RIA. Methods: Equine serum samples (n = 78) from clinical and experimental studies. Methods: In this ex...
Luo QH, Chen ML, Chen ZL, Huang C, Cheng AC, Fang J, Tang L, Geng Y.The aminolycoside Gentamicin is a widely used antibiotic, applied in equine medicine. Despite its clinical use, concerns remain regarding the potential toxic side-effects, such as nephrotoxicity. Early detection of renal damage is critical in preclinical drug development. This study was aimed to determine whether kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL) may be early indicators in the assessment of Gentamycin-induced nephrotoxicity. Methods: In our study, a model of Gentamicin-induced nephrotoxicity in male Sprague Dawley rats treated for up to 7 da...
Marklund S, Ellegren H, Eriksson S, Sandberg K, Andersson L.Ten (TG)n positive clones, isolated from an equine genomic library and sequenced, contained 12-19 uninterrupted TG repeats. Primers for polymerase chain reaction (PCR) were synthesized and nine of these (TG)n loci (HTG7-15) were successfully amplified and utilized in this study together with five previously reported equine microsatellite loci (HTG2-6). The PCR products were analysed by polyacrylamide gel electrophoresis followed by automated laser fluorescence detection or autoradiography. All microsatellites showed polymorphism and stable Mendelian inheritance. Differences in microsatellite v...
Swinburne JE, Boursnell M, Hill G, Pettitt L, Allen T, Chowdhary B, Hasegawa T, Kurosawa M, Leeb T, Mashima S, Mickelson JR, Raudsepp T, Tozaki T....A genetic linkage map of the horse consisting of 742 markers, which comprises a single linkage group for each of the autosomes and the X chromosome, is presented. The map has been generated from two three-generation full-sibling reference families, sired by the same stallion, in which there are 61 individuals in the F2 generation. Each linkage group has been assigned to a chromosome and oriented with reference to markers mapped by fluorescence in situ hybridization. The average interval between markers is 3.7 cM and the linkage groups collectively span 2772 cM. The 742 markers comprise 734 mic...
Gustafson MB, Martin RB, Gibson V, Storms DH, Stover SM, Gibeling J, Griffin L.This work determined whether mineral dissolution due to prolonged testing or storage of bone specimens in normal saline would alter their elastic modulus. In one experiment, small pieces of equine third metacarpal bone were soaked in normal saline supplemented with varying amounts of CaCl2. Changing Ca ion concentrations in the bath were monitored and the equilibrium concentration was determined. In a second experiment, the elastic moduli of twenty 4 x 10 x 100 mm equine third metacarpal beams were determined non-destructively in four-point bending. Half the beams were then soaked for 10 days ...
Ishida N, Hasegawa T, Takeda K, Sakagami M, Onishi A, Inumaru S, Komatsu M, Mukoyama H.The D-loop regions in equine mitochondrial DNA were cloned from three thoroughbred horses by polymerase chain reaction (PCR). The total number of bases in the D-loop region were 1114 bp, 1115 bp and 1146 bp. The equine D-loop region is A/T rich like many other mammalian D-loops. The large central conserved sequence block and small conserved sequence blocks 1, 2 and 3, that are common to other mammals, were observed. Between conserved sequence blocks 1 and 2 there were tandem repeats of an 8 bp equine-specific sequence TGTGCACC, and the number of tandem repeats differed among individual horses....
Boehm AK, Seth M, Mayr KG, Fortier LA.Many metabolic processes in chondrocytes thought to contribute to age-related changes in the extracellular matrix are influenced by known roles of Hsp90. Age-related decreases in the level of Hsp90 have been documented in numerous cell types and could contribute to cartilage degeneration. The aim of this study was to investigate the roles of age and Hsp90 in insulin-like growth factor 1 (IGF-1) and interleukin-1beta (IL-1beta) signaling in chondrocytes. Methods: Levels of Hsp90 messenger RNA (mRNA) and protein, with respect to age, were determined by quantitative real-time polymerase chain rea...
Mendoza L, Kaufman L, Standard PG.A practical, sensitive, and specific immunodiffusion test was developed for diagnosing and monitoring pythiosis in horses. Culture filtrates, a soluble cell mass, and trypsinized Pythium sp. antigens were evaluated against prepared rabbit anti-Pythium sp. serum and pythiosis horse case sera. The culture filtrate antigens demonstrated the greatest capacity for detecting precipitins and the greatest stability during storage. In contrast, the trypsinized antigens had the weakest capability for detecting multiple precipitins and the poorest stability. The 13 sera from horses with proven active pyt...
