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.
Kublicka A, Lorek D, Mikołajczyk-Martinez A, Chodaczek G, Chwirot A, Bażanów B, Matczuk AK.The process of viral entry into host cells is crucial for the establishment of infection and the determination of viral pathogenicity. A comprehensive understanding of entry pathways is fundamental for the development of novel therapeutic strategies. Standard techniques for investigating viral entry include confocal microscopy and flow cytometry, both of which provide complementary qualitative and quantitative data. Imaging flow cytometry, which integrates the advantages of both methodologies, offers significant potential in virological studies. In this investigation, we employed imaging flow ...
Fegraeus K, Riihimäki M, Nordlund J, Akula S, Wernersson S, Raine A.Single-cell RNA sequencing (scRNA-seq) is a valuable tool for investigating cellular heterogeneity in diseases such as equine asthma (EA). This study evaluates the HIVE™ scRNA-seq method, a pico-well-based technology, for processing bronchoalveolar lavage (BAL) cells from horses with EA. The HIVE method offers practical advantages, including compatibility with both field and clinical settings, as well as a gentle workflow suited for handling sensitive cells. Our results show that the major cell types in equine BAL were successfully identified; however, the proportions of T cells and macropha...
Wang J, Ren W, Li Z, Ma S, Li L, Wang R, Zeng Y, Meng J, Yao X.This study aims to analyze the whole-genome DNA methylation differences in Yili horses before and after racing, with the goal of identifying differentially methylated genes associated with racing performance and exploring the epigenetic mechanisms underlying exercise in horses. Blood samples were collected from the jugular veins of the top 3 Yili horses in a 5000 m race, which included 25 competitors, both prior to and within 5 min after the race. Genomic DNA was extracted, followed by sequencing using Whole-Genome Bisulfite Sequencing (WGBS) to assess DNA methylation levels, differentially me...
Hickman E, Carberry V, Carberry C, Cooper B, Mordant AL, Mills A, Sokolsky M, Herring LE, Alexis NE, Rebuli ME, Jaspers I, Sheats K, Rager JE.Growing evidence supports the importance of extracellular vesicle (EV) as mediators of communication in pathological processes, including those underlying respiratory disease. However, establishing methods for isolating and characterizing EVs remains challenging, particularly for respiratory samples. This study set out to address this challenge by comparing different EV isolation methods and evaluating their impacts on EV yield, markers of purity, and proteomic signatures, utilizing equine/horse bronchoalveolar lavage samples. Horses can serve as effective translational animal models for respi...
The AnalystJanuary 23, 2025
doi: 10.1039/d4an01501k
Göktaş EF, Kabil E, Söylemez Yeşilçimen E, Dirikolu L.Various technical methodologies are required to accurately detect substances of different chemical and pharmacological properties in biological samples, which are increasing in number and variety daily. Therefore, laboratories where many samples and different factors are analyzed simultaneously need methods with easy sample preparation, short analysis times and low analysis costs. In this study, the objective was to scan substances susceptible to chemical degradation, amenable to analysis without hydrolysis, and exhibiting short-term stability by employing a straightforward, expeditious, and c...
Whitlock F, van Dijk J, Hodgkinson JE, Grewar JD, Newton JR.Anthelmintic resistance (AR) threatens effective equine parasite control. Quarterly data summaries from faecal worm egg count testing (FWECT) performed by UK laboratories have appeared in Equine Quarterly Disease Surveillance Reports (EQDSR) since 2007, but have not previously been assessed. Objective: To assess strongyle FWECT methods and thresholds used by UK laboratories. To investigate factors associated with quarterly laboratory FWECT positivity rates between 2007 and 2023. Methods: Laboratory surveys and analysis of laboratory summary data. Methods: Laboratories were surveyed in Q3 2018 ...
Nakase M, Thapa J, Batbaatar V, Khurtsbaatar O, Enkhtuul B, Unenbat J, Lkham B, Fujita S, Koshikawa A, Tuanyok A, Saechan V, Higashi H, Hayashida K....Glanders and melioidosis are contagious zoonotic diseases caused by Burkholderia mallei and B. pseudomallei, respectively. Bacterial isolation and polymerase chain reaction (PCR) have been used to detect these bacteria in animals suspected of infection; however, both methods require skilled experimental techniques and expensive equipment. These obstacles make it difficult to diagnose B. mallei and B. pseudomallei infections in areas where reagents and equipment are difficult to procure. To solve this problem, we developed an easy and ready-to-use dried-format diagnostic tool based on loop-medi...
Palomino Lago E, Ross AKC, McClellan A, Guest DJ.Bone fractures are a significant problem in Thoroughbred racehorses. The risk of fracture is influenced by both genetic and environmental factors. To determine the biological processes that are affected in genetically susceptible horses, we utilised polygenic risk scoring to establish induced pluripotent stem cells (iPSCs) from horses at high and low genetic risk. RNA-sequencing on iPSC-derived osteoblasts revealed 112 genes that were significantly differentially expressed. Forty-three of these genes have known roles in bone, 27 are not yet annotated in the equine genome and 42 currently have ...
Nemoto M, Kawanishi N, Kambayashi Y, Bannai H, Yamanaka T, Tsujimura K.Equine influenza virus (EIV) can be transmitted by inhalation of aerosolized droplets, direct contact, and contaminated fomites. However, to our knowledge, there are no reports of the recovery of EIV from the air surrounding infected horses. Here, we evaluated whether EIV can be recovered from the air in the stalls of experimentally infected horses by using an air sampler. Furthermore, we examined whether rapid molecular test kits with reaction times of less than 30 min can detect EIV from air samples for potential field application. Two horses kept in individual stalls were experimentally i...
Hidalgo M.Sperm vitrification is an alternative freezing method, which includes high cooling rates and non-permeable cryoprotectants agents. The first attempt in equids was using the spheres technique by directly dropping small volumes of the sperm into liquid nitrogen. Later, vitrification was developed using 0.25 mL straws with outer covers, which resulted in similar progressive motility when compared to conventional freezing in donkeys (44.3 ± 15.0 % vs. 44.7 ± 18.2 %) or even higher in horses (48.2 ± 2.3 % vs. 37.3 ± 2.2 %). Subsequently, the vitrification of larger volumes of sperm in 0.5 mL st...
Birckhead EM, Raidal SL, Das S, Raidal SR.Sepsis is a main cause of death in neonatal foals. While the syndrome is not completely understood, sepsis is a dysregulated inflammatory response of the host to infection. It can be difficult to diagnose because of varying and non-specific clinical signs and imperfect diagnostic tests. Increased circulating nucleosome levels have been detected in humans and animals with sepsis, but this has not been documented in foals. Nucleosomes are released extracellularly during neutrophil extracellular trap formation, as well as from damaged and dead cells. We analysed plasma samples from clinically hea...
Cutarelli A, De Falco F, Serpe F, Izzo S, Fusco G, Catoi C, Roperto S.BPV1, BPV2, BPV13, and BPV14 are all genotypes of bovine delta papillomaviruses (δPV), of which the first three cause infections in horses and are associated with equine sarcoids. However, BPV14 infection has never been reported in equine species. In this study, we examined 58 fresh and thawed commercial semen samples from healthy stallions. In 34 (58.6%), bovine δPV DNA was detected and quantified using droplet digital polymerase chain reaction (ddPCR). Real time quantitative PCR (qPCR) was able to identify bovine δPV DNA in 5 samples (8.6%). Of the BPV-infected semen samples, 15 were posi...
Königová A, Babják M, Kuzmina TA, Burcáková Ľ, Syrota Y, Várady M.The study aimed to conduct a survey on the occurrence of benzimidazole (BZ) resistance in strongyles by in vitro egg hatch test (EHT) and larval development test (LDT) and to identify the effective indicators of early resistance detection on horse farms with associated risk factors analysis appraisal. In total, 203 horses from 8 farms underwent the fecal sampling of which 77 horses were selected for in vitro testing. Simultaneously, 18 horses were chosen to analyse the results of in vitro tests compared to the in vivo fecal egg count reduction test (FECRT). The EHT indicated the presence of re...
Fernandes T, Robin M, Freeman KP.Biological variation in laboratory results refers to physiological fluctuations that occur around a homeostatic setpoint (HSP) for various laboratory measurands. Assessment of biological variation includes determining individual variation (CV), group variation (CV), and analytical variation (CV). Reference change value (RCV) is an objective tool for an evidence-based approach to interpret data by assessing the significance of consecutive results in an individual for the diagnosis, prognosis, and monitoring of disease. The individualised reference intervals (iRI) are derived from the determinat...
Ekinci G, Deniz O, Tüfekci E, Timur MC, Onmaz AC, Günes V, Citil M, Keles I, Hoven RVD.Rhodococcus equi (R. equi) is a primary cause of pyogranulomatous pneumonia of foals between three weeks and five months of age. Early diagnosis of rhodococcal pneumonia has always been considered a preferable approach as it can lead to more successful treatment and better outcomes. Horse stud farms where the disease is common, neonatal foals are subjected to tests such as complete blood count and fibrinogen analysis at certain intervals. However, new biomarkers are needed in addition to blood count and fibrinogen measurement in this field for early diagnosis of diseases. Based on this need, i...
Parsley AL, Hollingshead NA, Gruber EJ.To minimise storage-related artefacts, EDTA-anticoagulated whole blood should be evaluated promptly or refrigerated, a challenge in ambulatory practice. The effects of suboptimal blood storage on equine leukocyte parameters are not well-documented. Objective: Document changes in leukocyte number, automated and manual differential counts, and morphology in equine blood stored at four temperatures for up to 5 days. Methods: In vitro experimental. Methods: EDTA-anticoagulated whole blood from 13 horses was stored in a refrigerator (4°C), cooler with icepacks (3°C-19°C), room temperature (22Â...
Baghdadi HBA, Abdelsalam M, Attia MM.Strongylus vulgaris, a devastating parasitic nematode in equids, causes life-threatening verminous aneurysms that are challenging to diagnose early. This study pioneered integrating nanotechnology into an indirect enzyme-linked immunosorbent assay (i-ELISA) system to enhance the sensitivity and specificity for detecting S. vulgaris larval antigens in equine serum samples, with PCR confirmation of the species. A conventional i-ELISA and an innovative nano-based ELISA were developed using excretory-secretory antigens from adult S. vulgaris worms. The nano-ELISA incorporated gold nanoparticles (1...
Hermans LM, Bonsergent C, Josson A, Rocafort-Ferrer G, Le Guyader M, Angelloz-Pessey S, Leblond A, Malandrin L.Equine piroplasmosis is a tick-borne disease mainly caused by Theileria equi and Babesia caballi. The objectives of this study were to analyse the frequency and routes of vertical transmission of these blood parasites from 179 asymptomatic mares to their foals. Foals were sampled within 72 h post-partum. The seroprevalences determined by Indirect Immunofluorescent Antibody Test (IFAT) and based on a subset sample of 107 couples, were 59.8% and 42.1% for T. equi and B. caballi, respectively in the mare population, and 54.2% and 40.2% in the foal population. A species-specific nested PCR was per...
Ferreira CMC, Simões BD, Marques EAS, Carbas RJC, da Silva LFM.Despite technological advancements in various industries, the equine sector still relies on old methods like horseshoeing. Although traditional, the industry is dynamic and well-funded. Therefore, there is a need to modernize these methods with more reliable and less invasive solutions for attaching horseshoes to horse hooves. There are currently several commercial adhesive solutions in the market specifically tailored to this application. In this work, the mechanical properties of two acrylic adhesives were characterized under quasi-static conditions. In the characterization process, tensile,...
Valderrama-Martinez C, Packham A, Zheng S, Smith W, Plancarte M, Aleman M.Evaluating antibody titers for Sarcocystis neurona for the diagnosis of equine protozoal myeloencephalitis from serum samples is a common practice. However, ensuring timely and proper refrigeration is not always possible. Objective: To evaluate immunofluorescent antibody (IFA) titers for S. neurona from serum samples stored at room temperature and 4°C. Methods: Twenty-two serum samples. Methods: Prospective longitudinal study. Two serum aliquots of 1 mL each were stored at room temperature (20-23.3°C) and 4°C. The unrefrigerated aliquot was immediately tested for IFA titers. Both aliquots...
Broeckl CV, Hiereth S, Straubinger RK.Diagnosis of equine Lyme borreliosis (LB), an infection caused by members of the Borrelia burgdorferi sensu lato complex (Bbsl), is challenging due to the nonspecific clinical signs of the disease and due to the variety of non-standardized serological tests. Specific vaccine-induced antibodies against LB, providing an effective protection against the infection, complicate the issue further. The standard for the detection of specific antibodies against Bbsl is a two-tier test system based on an enzyme-linked immunosorbent assay (ELISA) or indirect fluorescent antibody test (IFA) for antibody sc...
Chen JW, Stefanovski D, Haughan J, Jiang Z, Soma LR, Robinson MA.Interleukin 1 beta (IL-1β) and IL-1 receptor antagonist (IL-1RA) are both upregulated following traumatic injury. As IL-1RA blocks inflammatory signaling by IL-1β, overexpression of IL-1β relative to IL-1RA may drive inflammatory diseases. As such, determination of the relationship between IL-1β to IL-1RA expression levels in horses may provide insight into disease states or serve as a therapeutic readout of response to medical interventions. As techniques to detect plasma concentrations of IL-1β and IL-1RA in horses lack sensitivity, we developed and validated novel enzyme-linked immunos...
Crisà A, Cardinali I, Giontella A, Silvestrelli M, Lancioni H, Buttazzoni L.Lipizzan is a famous horse breed dating back to 1580 when the original stud of Lipica was established by the Hasburg Archduke Charles II. Currently, the Italian State Stud of Lipizzan Horses (ASCAL) is a conservation nucleus managed through strict mating rules where mitochondrial DNA sequences are used to verify the correct assignment of mares to a historical pedigree maternal lineage. Here, we analyzed the D-loop sequences of Lipizzan horses from the ASCAL in Monterotondo (Rome, Italy) in order to confirm their pedigree assignment to known female founder families. The concurrent investigation...
Wong JKY, Choi TLS, Wong COL, Curl P, Wan TSM, Ho ENM.Methylsulfonylmethane (MSM), also known as dimethyl sulfone, is a naturally occurring sulphur-containing compound that can be found in plants, animals and humans. MSM can also be a metabolite of dimethyl sulfoxide (DMSO). Due to their anti-inflammatory and analgesic effects, both MSM and DMSO are prohibited substances in horseracing. As both substances are naturally occurring, their misuse in horses is controlled by International Residue Limits (IRL) of 1200 and 15 μg/mL, respectively, in horse urine as established by the International Federation of Horseracing Authorities. The elimination ...
Koziy RV, Katselis GS, Yoshimura S, Simko E, Bracamonte JL.Prompt diagnosis of equine septic arthritis is crucial for successful treatment. Serum amyloid A (SAA) has been suggested as a reliable biomarker. However, we previously found that synovial fluid SAA increases in nonaffected joints of horses with septic arthritis. We hypothesized that systemic SAA may leak into the nonaffected joints. If this is the case, we also hypothesized that locally produced joint SAA isoforms may be better candidates for septic arthritis biomarkers. Thus, our objectives were 1) to evaluate the temporal kinetics of systemic and synovial fluid SAA in horses with septic ar...
McCreight KA, Barbosa LN, Odoi A, Reed P, Rajeev S.We estimated the seroprevalence in dogs, cats, and horses from Tennessee, USA, using the microscopic agglutination test (MAT) against 12 serovars. We observed seropositivity in 110 of 374 (29.4%) dogs, 21 of 170 (12.4%) cats, and 42 of 88 (47.7%) horses. The highest seroprevalence was observed for serovars Autumnalis (74.6%) in dogs, and Bratislava in cats (42.9%) and horses (95.2%). We found a significant level of potential cross-reactivity between multiple serovars tested, with highest cross-reactivity to serovar Autumnalis in dogs. seroprevalence was significantly higher in vaccinated ...
An T, Dugarjaviin M, Han H.The Mongolian horse, one of the oldest and most genetically diverse breeds, exhibits a wide variety of coat colors and patterns, including both wild-type and unique features. A notable characteristic of dun Mongolian horses is the presence of Bider markings-symmetrical, black-mottled patterns observed on the shoulder blades. These markings are also seen in Przewalski's horses. The dun coat color, a common wild-type phenotype in domestic horses, is characterized by pigment dilution with distinct dark areas and is regulated by mutations in the gene. This study aimed to investigate the role of ...
Samper JC, Hernandez Aviles JC, Ramirez-Agamez LF, Love CC, Gonzalez-Marin C, Fleury P, Dini P, De La Fuente A, Foss R, Campos FL, Ross PJ.The application of sex-sorted semen in horses has historically lagged the bovine industry due to differences in the reproductive physiology and grater variability in equine semen quality. Recent advancements, including SuperGen™ sorters and Ultraplus™ semen processing methods developed by STgenetics® (Navasota, Texas), have significantly improved the efficiency of the sex-sorting process and reduced the sperm damage previously reported. To facilitate the broader use sex-sorted semen in the equine industry, it is essential to address challenges such as shipping cooled semen to a central so...
Bernick A, Wehrend A.Uroperitoneum is a typical disease in newborn foals. It occurs rarely but regularly. The aim of the present study is to present clinical signs, laboratory findings and prognosis of foals with uroperitoneum. Methods: Medical records of 33 foals suffering from uroperitoneum that were treated between 2006 and June 2018 were reviewed retrospectively. Results: The incidence was 2.3% based of all presented foals during the first 14 days of life in the investigation period. Colts (78.8%) were significant more often affected (p=0,012) than fillies (21.2%). Common clinical signs were disturbed general ...
Steagall WK, Robek MD, Perry ST, Fuller FJ, Payne SL.The retrovirus equine infectious anemia virus (EIAV) encodes a dUTPase situated between reverse transcriptase and integrase. We have described the inability of EIAV with a 270-bp dUTPase deletion, delta DU EIAV, to replicate to wild-type (WT) levels in equine macrophages (D. S. Threadgill, W. K. Steagall, M. T. Flaherty, F. J. Fuller, S. T. Perry, K. E. Rushlow, S. F. J. LeGrice, and S. L. Payne, J. Virol. 67, 2592-2600, 1993). Here we describe the construction of a second dUTPase-deficient virus (DUD71E) containing a single amino acid substitution in dUTPase. delta DU and DUD71E replicate to ...
Graham JK.For many years, scientists have sought to develop laboratory assays that accurately predict the fertilizing capacity of a semen sample. This goal, however, has proven elusive and will most likely be very difficult to achieve, due to the complex nature of the problem. Part of the problem results from the many attributes that a spermatozoon must possess to fertilize an egg, and how laboratory assays can evaluate all of these attributes simultaneously. The percentage of motile sperm in a sample is most commonly used to evaluate semen quality. This assay, however, is not highly correlated with the...
Frampton AR, Uchida H, von Einem J, Goins WF, Grandi P, Cohen JB, Osterrieder N, Glorioso JC.To initiate infection, equine herpesvirus type 1 (EHV-1) attaches to heparan sulfate on cell surfaces and then interacts with a putative glycoprotein D receptor(s). After attachment, virus entry occurs either by direct fusion of the virus envelope with the plasma membrane or via endocytosis followed by fusion between the virus envelope and an endosomal membrane. Upon fusion, de-enveloped virus particles are deposited into the cytoplasm and travel to the nucleus for viral replication. In this report, we examined the mechanism of EHV-1 intracellular trafficking and investigated the ability of EH...
Shukla AK, Schauer R.A method is described for all quantitative determination of free or glycosidically bound sialic acids with special reference to erythrocyte membranes. Sialic acids, unsubstituted in their side chains, quantitatively yield formaldehyde after mild periodate oxidation (1 mM NaIO4, 15 min, 4 degrees C, in the dark). The formaldehyde is determined by the reaction with acetylacetone and ammonium acetate which leads to a sensitive fluorogen (F 410/510 nm). Sialic acids O-acetylated at C-9 or C-8 are not oxidized under these conditions. Therefore, they can be determined quantitatively by measuring the...
Wang X, Rikihisa Y, Lai TH, Kumagai Y, Zhi N, Reed SM.Anaplasma phagocytophilum immunodominant polymorphic major surface protein P44s have been hypothesized to go through antigenic variation, but the within-host dynamics of p44 expression has not been demonstrated. In the present study we investigated the composition and changes of p44 transcripts in the blood during the acute phase of well-defined laboratory A. phagocytophilum infections in naive equine hosts. Three traveling waves of sequential population changeovers of the p44 transcript species were observed within a single peak of rickettsemia of less than 1 month. During the logarithmic inc...
Johns IC, Adams EL.This study aimed to identify changing antimicrobial resistance patterns in isolates commonly obtained from equine clinical submissions. Laboratory records from 1999 to 2012 were searched for equine samples from which Escherichia coli or Streptococcus species was isolated. Susceptibility to enrofloxacin, ceftiofur, gentamicin, penicillin G, trimethoprim sulfamethoxazole (TMPS) and tetracyclines was noted. Isolates were divided into those identified between 1999 and 2004 (Early) and between 2007 and 2012 (Late). The proportion of isolates resistant to each antimicrobial and multiple drug-resista...
Taylor SE, Smith RK, Clegg PD.The goal in the therapeutic use of mesenchymal stem cells (MSCs) in musculoskeletal disease is to harness the regenerative nature of these cells focussing on their potential to grow new tissues and organs to replace damaged or diseased tissue. Laboratory isolation of MSCs is now well established and has recently been demonstrated for equine MSCs. Stem cell science has attracted considerable interest in both the scientific and clinical communities because of its potential to regenerate tissues. Research into the use of MSCs in tissue regeneration in general reflects human medical needs, however...
Hessel LN, Bosch G, van Weeren PR, Ionita JC.Autologous platelet concentrates (APCs) are being used increasingly in horses to enhance regeneration in tissues that have poor natural healing capabilities. Numerous APC systems, which are based on different preparation techniques and were originally developed for human patients, are now routinely used in equine cases. However, preliminary process validation and adequate in vitro biochemical characterisation of most of these systems do not exist for horses. Objective: To compare haematological findings and growth factor concentrations of equine APCs obtained with 4 commercially available syst...
Bauer N, Nakagawa J, Dunker C, Failing K, Moritz A.The automated laser-based hematology analyzer Sysmex XT-2000iV™ provides a 5-part differential count and specific cytograms that are of great interest for large veterinary laboratories. The aim of the study was to validate the Sysmex XT-2000iV compared to the laser-based hematology analyzer ADVIA® 2120 and manual differential in dogs, cats, and horses as well as the impact of anticoagulant (heparin, ethylenediamine tetra-acetic acid [EDTA], and citrate) and storage at 22°C and 4°C. Consecutive fresh K(3)-EDTA blood samples from 216 cats, 314 dogs, and 174 horses were included. The impact ...
Sun Y, Wang C, Wang Y, Zhang T, Ren L, Hu X, Zhang R, Meng Q, Guo Y, Fei J, Li N, Zhao Y.Based on the recently released horse genome, we have characterized the genomic organization of the horse Ig gene loci. The horse IgH locus in genomic scaffold Un0011 contains 40 D(H) segments, 8 J(H) segments and 50 V(H) segments. The Igkappa locus contains only a single C(kappa) gene, 5 J(kappa) segments and a 60 V(kappa) segments, whereas the Iglambda locus contains 7 C(lambda) genes each preceded by a J(lambda) gene segment. A total of 110 V(lambda) segments with the same transcriptional polarity as J(lambda)-C(lambda) were identified upstream of the J(lambda)-C(lambda) cluster. However, 34...
Moore AI, Squires EL, Graham JK.Cryopreservation induces partially irreversible damage to equine sperm membranes. Part of this damage occurs due to membrane alterations induced by the membrane changing from the fluid to the gel-state as the temperature is reduced lower than the membrane transition temperature. One way to prevent this damage is to increase the membrane fluidity at low temperatures by adding cholesterol to the membrane. Different concentrations of cholesterol-loaded-cyclodextrins (CLC) were added to stallion sperm to determine the CLC concentration that optimizes cryosurvival. Higher percentages of motile sper...
Paras KL, George MM, Vidyashankar AN, Kaplan RM.Gastrointestinal nematode parasites are important pathogens of all domesticated livestock species. Fecal egg counts (FEC) are routinely used for evaluating anthelmintic efficacy and for making targeted anthelmintic treatment decisions. Numerous FEC techniques exist and vary in precision and accuracy. These performance characteristics are especially important when performing fecal egg count reduction tests (FECRT). The objective of this study was to compare the accuracy and precision of three commonly used FEC methods and determine if differences existed among livestock species. In this study, ...
Tijms MA, Snijder EJ.Non-structural protein 1 (nsp1), the N-terminal subunit of the replicase polyprotein of the arterivirus Equine arteritis virus (EAV), is essential for viral subgenomic mRNA synthesis, but fully dispensable for genome replication. However, at the molecular level, the role of nsp1 in EAV subgenomic mRNA synthesis is poorly understood. A yeast two-hybrid screen did not reveal interactions between EAV nsp1 and other viral non-structural proteins or the nucleocapsid protein, although both nsp1 and the nucleocapsid protein were found to form homomers. Subsequently, a yeast two-hybrid screen of a HeL...
Wagner B.Antibodies of the horse were studied intensively by many notable immunologists throughout the past century until the early 1970's. After a large gap of interest in horse immunology, additional basic studies on horse immunoglobulin genes performed during the past 10 years have resulted in new insights into the equine humoral immune system. These include the characterization of the immunoglobulin lambda and kappa light chain genes, the immunoglobulin heavy chain constant (IGHC) gene regions, and initial studies regarding the heavy chain variable genes. Horses express predominately lambda light c...
Li F, Puffer BA, Montelaro RC.Equine infectious anemia virus (EIAV) contains the simplest genome among lentiviruses in that it encodes only three putative regulatory genes (S1, S2, S3) in addition to the canonical gag, pol, and env genes, presumably reflecting its limited tropism to cells of monocyte/macrophage lineage. Tat and Rev functions have been assigned to S1 and S3, respectively, but the specific function for the S2 gene has yet to be determined. Thus, the function of S2 in virus replication in vitro was investigated by using an infectious molecular viral clone, EIAVUK. Various EIAVUK mutants lacking S2 were constr...
Vanin AF, Serezhenkov VA, Mikoyan VD, Genkin MV.The parameters of EPR signal from dinitrosyl-iron complexes (DNIC) with bovine serum albumin (BSA), horse hemoglobin (Hb), and apometallothionein (apo-Mt) of horse kidney incorporating one (BSA, Hb) or two thiol-containing ligands (apo-Mt) were compared. The EPR signal from DNIC-BSA was characterized by the rhombic symmetry of g tensor at room temperature of signal recording (ambient temperature) or at 77K in the solution frozen in the presence of glycerol. In freezing of the solution in the absence of glycerin, under the exposure of DNIC-BSA to negatively charged sodium dodecyl sulfate (SDS) ...
Zhou M, Wang Q, Sun J, Li X, Xu L, Yang H, Shi H, Ning S, Chen L, Li Y, He T, Zheng Y.The importance of microRNAs at the post-transcriptional regulation level has recently been recognized in both animals and plants. We used the simple but effective sequential method of first Blasting known animal miRNAs against the horse genome and then using the located candidates to search for novel miRNAs by RNA folding method in the vicinity (+ -500 bp) of the candidates. Here, a total of 407 novel horse miRNA genes including 354 mature miRNAs were identified, of these, 75 miRNAs were grouped into 32 families based on seed sequence identity. MiRNA genes tend to be present as clusters in som...
Novak S, Smith TA, Paradis F, Burwash L, Dyck MK, Foxcroft GR, Dixon WT.The global proteome of sperm and seminal plasma of fertile stallions was investigated to determine whether associations with relative in vivo fertility exist. Seven stallions at stud in a commercial breeding station were collected throughout the breeding season and bred to a total of 164 mares to determine conception rates. On three occasions during the breeding season, raw semen was obtained from a regular collection for proteomic analysis using two-dimensional electrophoresis and also assessed for routine semen quality end points. First cycle conception rate was negatively related to ejacula...
Wallner B, Piumi F, Brem G, Müller M, Achmann R.Y chromosome polymorphisms such as microsatellites or single nucleotide polymorphisms represent a paternal counterpart to mitochondrial DNA (mtDNA) for evolutionary and phylogeographic studies. The use of Y chromosome haplotyping in natural populations of species other than humans is still hindered by the lack of sequence information necessary for polymorphism screening. Here we used representational difference analysis (RDA) followed by a screen of a bacterial artificial chromosome (BAC) library for repetitive sequences to obtain polymorphic Y-chromosomal markers. The procedure was performed ...
Finley MR, Li Y, Hua F, Lillich J, Mitchell KE, Ganta S, Gilmour RF, Freeman LC.In dogs and in humans, potassium channels formed by ether-a-go-go-related gene 1 protein ERG1 (KCNH2) and KCNQ1 alpha-subunits, in association with KCNE beta-subunits, play a role in normal repolarization and may contribute to abnormal repolarization associated with long QT syndrome (LQTS). The molecular basis of repolarization in horse heart is unknown, although horses exhibit common cardiac arrhythmias and may receive drugs that induce LQTS. In horse heart, we have used immunoblotting and immunostaining to demonstrate the expression of ERG1, KCNQ1, KCNE1, and KCNE3 proteins and RT-PCR to det...
Allen G, Yeargan M, Costa LR, Cross R.An experimental system that permits sensitive and reproducible detection of equine herpesvirus 1 (EHV-1)-specific cytotoxic T-lymphocyte (CTL) activity in the horse was developed. Peripheral blood mononuclear cells (PBMC) collected from immune horses were restimulated in vitro by culture with live EHV-1. Cytotoxic activity against virus-infected, pokeweed mitogen-stimulated lymphoblast targets was assessed in a 4-h 51Cr release assay. The optimal conditions for in vitro stimulation of equine memory CTLs and for preparation of EHV-1-infected target cells expressing viral antigens were systemati...
Freeman K, Woods E, Welsby S, Percival SL, Cochrane CA.Evidence of biofilms in human chronic wounds are thought to be responsible for preventing healing in a timely manner. However, biofilm evidence in horse wounds has not yet been documented. Consequently, this study aimed to determine whether biofilms could be detected in wounds, and to investigate the microbiology of chronic wounds in horses. Prior to analysis, wound surfaces were irrigated with 5 mL of sterile saline to remove debris. All wounds were swabbed twice (1 cm2 area) using sterile cotton-tipped swabs. In addition to this, 2 tissue biopsies were taken to investigate evidence of biofil...
Deregt D, de Vries AA, Raamsman MJ, Elmgren LD, Rottier PJ.Monoclonal antibodies (MAbs) to equine arteritis virus (EAV) proteins were produced and characterized. The protein specificities of eight MAbs were determined definitively by immunoprecipitation of EAV proteins expressed from vaccinia virus recombinants (VVRs). Included were two new VVRs produced for this study, expressing the M and the GL proteins, respectively. Three MAbs were determined to be N-specific and five MAbs recognized the GL protein. One GL-specific MAb, 17F5, of the IgA class, efficiently neutralized EAV infectivity. In competitive binding assays (CBAs), the N-specific MAbs defin...
Suire S, Stewart F, Beauchamp J, Kennedy MW.The equine conceptus is surrounded by a fibrous capsule that persists until about day 20 of pregnancy, whereupon the capsule is lost, the conceptus attaches to the endometrium and placentation proceeds. Before attachment, the endometrium secretes in abundance a protein of the lipocalin family, uterocalin. The cessation of secretion coincides with the end of the period during which the conceptus is enclosed in its capsule, suggesting that uterocalin is essential for the support of the embryo before direct contact between maternal and foetal tissues is established. Using recombinant protein and ...
Cook RF, Cook SJ, Li FL, Montelaro RC, Issel CJ.A single-tube reverse transcriptase-polymerase chain reaction (RT-PCR) using a fluorogenic real-time PCR detection method is described for the quantitation of equine infectious anemia virus (EIAV) RNA in the plasma of equids. To compensate for variations inherent in sample preparation a multiplex real-time RT-PCR system was developed that permitted the simultaneous calculation of the nucleic acid recovery rate along with the copy number of viral RNA molecules. Detection of EIAV RNA was linear from 10(9) to 10(1) molecules with intra- and inter-assay variability of less than 1% at 10(8), 10(6),...
Lin YZ, Sun LK, Zhu DT, Hu Z, Wang XF, Du C, Wang YH, Wang XJ, Zhou JH.Human schlafen11 is a novel restriction factor for HIV-1 based on bias regarding relative synonymous codon usage (RSCU). Here, we report the cloning of equine schlafen11 (eSLFN11) and the characteristics of its role in restricting the production of equine infectious anemia virus (EIAV), a retrovirus similar to HIV-1. Overexpression of eSLFN11 inhibited EIAV replication, whereas knockdown of endogenous eSLFN11 by siRNA enhanced the release of EIAV from its principal target cell. Notably, although eSLFN11 significantly suppressed expression of viral Gag protein and EIAV release into the culture ...
Roberts MM, Oroszlan S.Capsids of equine infectious anemia virus have been isolated as cone-shaped particles 60 x 120 nm in size. Detergent treatment of whole virus followed by two cycles of rate-zonal centrifugation in Ficoll produces these capsids in a yield of approximately 10%. The major protein components are the gag-encoded p11 nucleocapsid protein and p26 capsid protein, which are present in equimolar amounts. Substantial cleavage of p11 to p6 and p4 can be observed under conditions where the viral protease packaged in the capsid is enzymatically active.
