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.
Pinaud MA, Roser JF, Dybdal N.In vitro responsiveness of the horse anterior pituitary (AP) gonadotropes to single and multiple GnRH challenges was examined. The pituitaries were collected from reproductively sound mares in estrus (n = 5) and diestrus (n = 5). Uniform 0.5 mm AP slices were subdivided using a 3 mm biopsy punch and then bisected for use in the perifusion chamber. Four bisected sections per chamber were perifused at 0.5 ml/min at 37 C for 560 min in Medium 199 saturated with 95% 0(2)/5% CO2. Ten minute fractions were collected after an initial 2 hr equilibration period. Four different treatment regimes of GnRH...
Teifke JP, Weiss E.Unfixed and formalin-fixed frozen sections and paraffin-sections of histopathologically confirmed sarcoids of 20 horses were studied in the PCR. The used set of primers was located in the E5 open reading frame fitting both to bovine papillomavirus 1 (BPV-1) and BPV-2. Independent of the quality of the used tissues BPV-DNA was detected in all 20 sarcoids. By cleaving with restriction endonuclease Bst XI it was shown that the DNA-sequences amplified by PCR were identical with that of BPV-1. The results support the general view that BPV play an important role in equine sarcoids.
Posnett ES, Ambrosio RE.A genomic library of Babesia caballi DNA was constructed in the plasmid vector pUC13. The specificity of the clones for B. caballi was established by the lack of hybridization to Babesia equi, Babesia bovis, Babesia bigemina and equine DNA. Two probes, pBC11 and pBC191, were isolated that could detect 0.25 ng and 0.125 ng of B. caballi DNA, corresponding to a parasitaemia of 0.12% and 0.06% respectively. pBC191 could detect B. caballi parasites in the blood of an experimentally infected horse as well as in naturally infected horses.
Van den Berg JS.Clinically normal horses (n = 8) with ages ranging from 5 to 8 years, were starved for 12 h and their plasma ammonia concentrations were measured. The mean fasting plasma ammonia concentration was 17.8 +/- 3.8 mumol l-1. After dosing ammonium chloride at a dose rate of 0.02 g kg-1, there was a significant increase in plasma ammonia concentration, with a maximum rise after 20 min (P less than 0.05). To investigate the influence of temperature on plasma ammonia concentrations of stored samples, 8 plasma samples were stored at -20 degrees C and 4 degrees C respectively. The plasma ammonia concent...
Joris L, Quinton PM.Basic components of unstimulated electrolyte transport across equine tracheal mucosa were characterized. After the tissue was mounted in Ussing chambers, both current and tissue resistance gradually increased for approximately 60 min before reaching stable values. Thereafter, under open-circuit conditions, the tissue had a resistance of 250 +/- 14 omega.cm2, generated a transepithelial potential difference of -34 +/- 1.7 (SE) mV (referenced to the serosal side) and an equivalent short-circuit current (Ieqsc) of -149 +/- 10.2 microA/cm2. Even though 10(-5) M amiloride reduced the current by app...
Forrer R, Gautschi K, Lutz H.It was the goal of this paper to establish total selenium reference values for Switzerland in different animal species and in humans. To this purpose, a flameless atomic absorption method with deuterium background compensation utilizing a graphite furnace atomization system with a pyrolytic platform inside and palladium solution as matrix modifier was developed for the measurement of selenium in serum. The method was characterized by rapid performability, small sample requirement, acceptable detection limit (0.04 mumol/L) and precision and a linear range of measurement up to 4 mumol/L. The met...
Craig AM, Blythe LL, Rowe KE, Lassen ED, Walker LL.An oral vitamin E absorption test used in human beings was modified for use in horses. The most appropriate techniques with which to measure gastrointestinal tract absorption of vitamin E (alpha-tocopherol) in horses were developed. Vitamin E was administered orally, and serum values of alpha-tocopherol were measured by use of high-performance liquid chromatography at 0, 3, 6, 9, 12, and 24 hours after vitamin E administration. Variables included comparison of 2 dosages (45 and 90 IU/kg of body weight), routes of administration, and absorption dynamics of 3 preparations of dl-alpha-tocopherol....
Ross PF, Rice LG, Plattner RD, Osweiler GD, Wilson TM, Owens DL, Nelson HA, Richard JL.Ninety-eight samples of feeds associated with 44 cases of equine leukoencephalomalacia (ELEM) and 83 samples of feed associated with 42 cases of a porcine pulmonary edema syndrome (PPE) were analyzed for fumonisin B1 (FB1). For comparison purposes, 51 feed samples not associated with PPE or ELEM were also analyzed. Feed associated with ELEM contained FB1 ranging from less than 1 microgram/g to 126 micrograms/g with 75% of the cases having at least 1 sample above 10 micrograms/g. Feeds associated with PPE ranged from less than 1 microgram/g to 330 micrograms/g with 71% of the cases having at le...
den Boon JA, Snijder EJ, Chirnside ED, de Vries AA, Horzinek MC, Spaan WJ.The nucleotide sequence of the genome of equine arteritis virus (EAV) was determined from a set of overlapping cDNA clones and was found to contain eight open reading frames (ORFs). ORFs 2 through 7 are expressed from six 3'-coterminal subgenomic mRNAs, which are transcribed from the 3'-terminal quarter of the viral genome. A number of these ORFs are predicted to encode structural EAV proteins. The organization and expression of the 3' part of the EAV genome are remarkably similar to those of coronaviruses and toroviruses. The 5'-terminal three-quarters of the genome contain the putative EAV p...
Guillemette JG, Matsushima-Hibiya Y, Atkinson T, Smith M.A gene for expression of horse heart myoglobin in Escherichia coli has been constructed in one step from long synthetic oligonucleotides. The synthetic gene contains an efficient translation initiation signal and used codons that are commonly found in E. coli. Unique restriction sites are placed throughout the gene. It has been inserted in a phagemid vector and is expressed from the lac promoter in E. coli at high efficiency, the soluble heme protein representing approximately 10% of soluble protein. Two versions of horse heart myoglobin were produced with aspartic acid or asparagine at residu...
Marlin DJ, Harris RC.In vitro titration of muscle homogenates has been used to assess muscle buffering capacity (beta mtitr) in a variety of species. In the present study, factors likely to affect the estimation of beta mtitr were investigated. Also, values of beta mtitr from normal Thoroughbred horses are presented. A non-linear titration curve was obtained with addition of HCl to muscle homogenates. As a result, beta mtitr is expressed as the mumol H+ required to change the pH of 1g of dry muscle or wet muscle from 7.1 to 6.5. An effect of dilution on the initial pH was found below 40 mg wet muscle per ml homoge...
O'Keefe JS, Murray A, Wilks CR, Moriarty KM.Unpurified DNA derived from cultures of equine fetal kidney cells infected with either equine herpesvirus type 1 or equine herpesvirus type 4 was amplified by the polymerase chain reaction using one pair of oligonucleotide primers. Restriction endonuclease digestion of the amplified segments with PvuII, followed by electrophoresis, revealed restriction fragment length polymorphisms which enabled the two virus types to be differentiated.
Martens JG, Stephens KA, Kerchner LJ, Heck FC, Martens RJ.Plasma fibronectin concentrations were measured in clinically healthy mares and their neonatal foals, using a modified human fibronectin competitive enzyme-linked immunosorbent assay. Ranges of plasma fibronectin were established in clinically healthy horses, and the assay was reliable and reproducible. Plasma fibronectin concentrations were similar in mares and foals, both before and after colostrum ingestion.
Khittoo G, Vermette L, Nappert G, Lariviere N.In mammalian species studied previously, pepsinogen consisted of biochemically different groups of isozymogens. By use of gel filtration chromatography and electrophoresis, we isolated a predominant pepsinogen from the gastric mucosa of a horse. Peptide mapping with V8 protease revealed differences with its porcine homologue. However, porcine and equine pepsinogens, when activated to pepsin, had a similar pattern of activity when hemoglobin was used as substrate. Those results suggest that differences must exist in the primary structure of the pepsinogens of the 2 species.
Whitcomb RW, Schneyer AL, Roser JF, Hughes JP.Using a LH radioligand receptor assay (RRA) previously validated for use in serum and an equine monoclonal RIA, we have distinguished a subset of subfertile stallions with an elevated RRA/RIA ratio. After purification of the active moiety by anion exchange chromatography and immunoprecipitation with the equine LH (eLH) monoclonal antibody, RRA activity remained in the supernatant. This activity was also recognized by a polyclonal LH antibody (GDN 15) with wide cross-species recognition. This active fraction was further purified by gel filtration chromatography and shown to displace labeled eLH...
Gaĭdamovich SIa, Pomelova VG, Lavrova NA, Mel'nikova EE, Sokolova MV, Kharitonenkov IG, Zlobin VN.Potentialities of differentiation between Venezuelan equine encephalomyelitis (VEE) complex viruses by time-resolved fluoroimmunoassay and enzyme immunoassay were studied. For this, 4 test systems were used based on different combinations of native and labeled polyclonal antibodies to VEE virus, strain Trinidad, and monoclonal (MCA) antibody MAK 14-7 to protein EL of this virus. The maximal sensitivity and specificity was achieved in the test system formed from native MCA MAK 14-7 for sensitization of the solid phase and labeled polyclonal immunoglobulins for demonstration of the test results....
Takeda A, Suzuki E, Kamei K, Nakata H.Several kinds of loline-type alkaloids, norloline, loline, N-acetylnorloline, N-acetylloline, N-formylnorloline, N-formylloline and N-methylloline were detected in the urine of race-horses. Furthermore, a new compound of the alkaloids, N-senecioylnorloline, was also found and identified. These compounds were mainly identified by means of gas chromatography-mass spectrometry (GC-MS) and gas chromatography-fourier transform-infrared spectrometry (GC-FT-IR). A certain plant of Gramineae containing four kinds of loline-type alkaloids was found in a bale of hay used for the horse forage. The taxono...
Traub-Dargatz JL.Bacterial pneumonia in the horse often occurs secondary to viral respiratory infection; however, primary infections can occur. A diagnosis of bacterial pneumonia is made on the basis of history, physical examination, and laboratory and radiographic findings. Treatment consists of appropriate antimicrobial therapy for at least 7 to 10 days; further therapy may be needed in more severe cases. Prevention of bacterial pneumonia includes attempting to prevent viral respiratory infections and appropriate management to prevent stress to the respiratory tract.
Judson R, Songer JG.The minimal inhibitory concentrations of 39 antimicrobial agents for 54 isolates of Corynebacterium pseudotuberculosis in vitro have been determined. The most active agents were penicillins, macrolides, tetracyclines, cephalosporins, lincomycin, chloramphenicol, and rifampicin. Most isolates were resistant to aminoglycosides, nitrofurans, polymyxins, nalidixic acid, and cycloheximide.
Stewart F, Tuffnell PP.A 514 bp cDNA transcript coding for 78% of horse (Equus caballus.) GH has been cloned and sequenced. The deduced amino acid sequence corresponded precisely to that previously obtained by protein sequencing and, in addition, provided new sequence information for the signal peptide. The missing 3' fragment of the cDNA was reconstructed using synthetic oligonucleotides and site-specific directed mutagenesis. The complete cDNA sequence was then inserted into an expression vector (PIN-III-lppp-5) which utilizes a bacterial signal peptide to secrete the expressed product into the periplasmic space o...
Dunnett M, Harris RC, Orme CE.A simple, robust and reproducible analytical method for the determination of phosphocreatine (PCr), creatine (Cr) and creatinine (Cn) in equine skeletal muscle is presented. The technique used isocratic reverse-phase ion-pairing high-performance liquid chromatography. Neutralized perchloric acid extracts of equine muscle biopsies were analysed and the values obtained were compared with determinations from an established enzymic procedure. Good resolution of all three metabolites was achieved within a retention time of less than 11 min. Linearity for each metabolite within the concentration ran...
Borroto-Esoda K, Boone LR.The endogenous reverse transcriptase reaction of equine infectious anemia virus (EIAV) has been studied, and conditions allowing synthesis of full-length minus-strand DNA have been determined. In contrast to results reported for other retroviruses, synthesis of EIAV full-length minus-strand DNA was not impaired by high concentrations of Nonidet P-40, a nonionic detergent used to make the virion envelope permeable. All components of the reaction were titrated for maximum synthesis of complete minus strands, and a time course under the standardized conditions was determined. Minor subgenomic ban...
Schneider C, Müller FP, Bertschi I.We describe the utilization of the Abbott Vision system in a horse clinic and a comparison with the Compur M 2000 CS (Bayer Diagnostics and Electronics). Discrepancies were found in respect to precision and accuracy of results. Both systems showed good practicability during routine operation but different cost-effectiveness.
Peyton DH.Specific heme protons for the majority of resonances in the downfield resolved region of equine met-azido myoglobin have been assigned using solely the two-dimensional 1H NMR experiments NOESY and COSY. Metazido myoglobin provides a useful test case for the applicability of these techniques to paramagnetic proteins for the following reasons. First met-azido myoglobin is a mixed spin-state protein, with significantly shorter relaxation times and broadened lines relative to pure low-spin systems (eg., met-cyano myoglobin). Second, met-azido hemoglobin and met-azido myoglobin are important as mod...
Lumsden JH, Jacobs RM, Physick-Sheard P.A study was conducted to examine the relationship of delta bilirubin to traditional bilirubin fractions to determine if delta bilirubin might assist in differentiating causes of hyperbilirubinemia in the horse. A recently introduced thin-film method was used for delta bilirubin determination. Thin-film and the traditional diazo methods were used for determining total, unconjugated, and conjugated bilirubin fractions. Sera from 38 healthy and 85 sick horses were examined.Thin-film serum delta bilirubin determination does not appear to assist in differentiating causes of unconjugated hyperbiliru...
Troyer DL, Oyster RO, Hunt MC.The purpose of this study was to find a combination histochemical staining technique for the evaluation of equine skeletal muscle that is reliable and effective, while offering a substantial reduction in the labor and cost involved with currently used individual histochemical methods. Several combinations under varying conditions of pH were studied. The most uniform results were obtained using an acid preincubation step at an optimal pH of 4.2 followed by reduced nicotinamide adenine dinucleotide-tetrazolium reductase (NADH-TR) and the remainder of the acid-ATPase procedure.
Barakat-Walter I, Deloulme JC, Sensenbrenner M, Labourdette G.We have previously shown that neuroblasts from cerebral hemispheres of 6-day-old chick embryos are able to proliferate when grown in the presence of fetal calf serum. We report here that in the presence of horse serum alone the proliferative rate of neuroblasts is strongly reduced. A high proliferative rate is restored upon the addition of bovine transferrin and to a lesser extent with added FeSO4 or hemin. These findings suggest that the transferrin of horse serum cannot be used by chick neuroblasts in vitro, while bovine transferrin exogenously added is active in promoting cell proliferation...
Yalamanchili RR, Raengsakulrach B, O'Callaghan DJ.We have previously reported the sequence of the equine herpesvirus one genomic termini that are homologous to the genomic termini of other herpesviruses. In this paper, we present the nucleotide sequence adjacent to the left terminus sequence (map units 0.0087 to 0.0237). This sequence codes for two open reading frames (ORF) which are homologous to ORF2 and ORF3 of the varicella-zoster virus genome and are located at colinear positions. The L region sequence presented here also contains a segment that is involved in the generation of the genome of EHV-1 DI particles through recombination with ...
Kong XG, Pang H, Sugiura T, Sentsui H, Onodera T, Matsumoto Y, Akashi H.Equine infectious anemia virus (EIAV) core proteins were obtained from a baculovirus expression system. Recombinant baculoviruses (rBVs) highly expressed the Gag precursor and p26 antigens in an rBV-infected Sf21 cell culture supernatant. Enzyme-linked immunosorbent assay (ELISA) and agar gel immunodiffusion (AGID) were conducted using the expressed proteins to detect antibodies from experimentally infected horses. The expressed antigens showed low background levels, high specificity and sensitivity in ELISA and AGID. The results of the serological tests using the expressed antigens were ident...
Koupai-Abyazani MR, Esaw B, Laviolette B.Urine and serum samples collected from four standard-bred mares after and oral regimen administration of methocarbamol were extracted and analyzed. The method consisted of enzyme hydrolysis followed by a one-step liquid-liquid extraction, separation on a reversed-phase (RP-18) column, and detection using an ultraviolet (UV) detector. The confirmation was carried out using a liquid chromatography-atmospheric pressure ionization-mass spectrometry (LC-API-MS) system. Maximum methocarbamol concentrations of 1498, 1734, 1547, 2322 micrograms/mL in urine and 4.9, 1.7, and 3.6 micrograms/mL in serum ...
Pereira B, Dorado J, Diaz-Jimenez M, Consuegra C, Ortiz I, Gosalvez J, Hidalgo M.The acquisition of equine oocyte developmental capacity is ensured by the follicular environment, such as granulosa cells, which could reflect the meiotic development potential of immature oocytes. This study evaluated the relationship between DNA fragmentation of granulosa cells, using the chromatin dispersion test, and equine oocyte meiotic development after in vitro maturation. Granulosa cells and cumulus-oocytes complexes (n = 50) were recovered from slaughterhouse-derived ovaries. Oocytes were in vitro matured, stained and evaluated under fluorescence microscopy. Maturation rates were c...
Bouwman FG, van Ginneken MM, van der Kolk JH, van Breda E, Mariman EC.The aim of the study was to identify new biomarkers for acute tying-up in horses. Skeletal muscle biopsies were taken from 3 horses suffering from acute tying-up and 3 healthy horses. We performed 2D gel electrophoresis and mass spectrometry for identification of proteins that are differentially expressed in tying-up. 2D gel electrophoresis of skeletal muscle sequential extracts yielded more than 350 protein spots on each gel, of which 14 were differentially expressed more than two-fold (p<0.05). In-gel digestion followed by peptide mass fingerprinting enabled identification of three significa...
Hu Z, Zhu C, Chang H, Guo W, Liu D, Xiang W, Wang X.The objective of this study was to develop a novel EvaGreen (EG) based real-time PCR technique for the simultaneous detection of Equine herpesvirus 1 (EHV-1) and Equine herpesvirus 4 (EHV-4) genomes from equine nasal swabs. Viral genomes were identified based on their specific melting temperatures (T m), which are 88.0 and 84.4 °C for EHV-1 and EHV-4, respectively. The detection limitation of this method was 50 copies/μl or 0.15 pg/μl for EHV-1 and 5 copies/μl or 2.5 fg/μl for EHV-4. This assay was 50-1,000 times more sensitive than the SYBR Green (SG)-based assay using the same primer...
Bailey MT, Christman SA, Wheaton JE, Troedsson MH, O'Brien TD, Ababneh MM, Santschi E.The aim of this study was to examine inhibin production in granulosa-theca cell tumours (GTCT). The experimental aims were: (i) to determine GTCT cell types that produce inhibin alpha- and betaA-subunits; (ii) to determine whether alpha- and betaA-subunit forms differ in GTCT fluid and normal equine follicular fluid (eFF); and (iii) to determine whether dimeric inhibin (alpha betaA) is present in GTCT plasma and tumour fluid. Plasma, tumour fluid and tumour tissue were collected from mares (n=6) with GTCT. Plasma and eFF were collected during the follicular phase from mares (n=4) undergoing no...
Stanley SM, Wee WK, Lim BH, Foo HC.Direct-injection LC-LC hybrid tandem MS methods have been developed for undertaking broad-based screening for acidic drugs in protein-precipitated plasma and neutral doping agents in equine urine. In both analyses, analytes present in the matrix were trapped using a HLB extraction column before being refocused and separated on a Chromolith RP-18e monolithic analytical column using a controlled differential gradient generated by proportional dilution of the first column's eluent with water. Each method has been optimised by the adoption of a mobile phase and gradient that was tailored to enhanc...
Becht JL, Page EH, Morter RL, Boon GD, Thacker HL.A series of modified (field) tests were compared to a crossmatch between mare and foal for their reliability in predicting neonatal isoerythrolysis (NI) in eight foals born to experimentally alloimmunized mares. In the field tests, mare's serum, plasma and colostrum were combined with foal erythrocytes washed by a modified procedure to determine which combination was the best predictor of impending NI. A consistent grading system for agglutination and hemolysis was employed. The field tests using mare's plasma demonstrated less agglutination and hemolysis than tests where serum was employed. I...
Birmingham SSW, Mason RM.Serum amyloid A (SAA) is one of the major acute phase proteins in horses. It serves as a marker for systemic inflammation and infection, as the concentration can increase 100- to even 1000-fold during systemic disease processes. The objective of this study was to evaluate the effect of sedation, oral examination, and odontoplasty on systemic inflammation as measured by SAA in the adult performance horse. This study included 32 clinically healthy adult performance horses. Blood samples were collected immediately prior to sedation, oral examination, and odontoplasty and 48 and 72 hours afterward...
Capistrano da Silva E, Gibson DJ, Jeong S, Zimmerman KL, Smith-Fleming KM, Martins BDC.Matrix metalloproteinases (MMPs)-2 and -9 are present in corneal ulcers, and an imbalance between MMPs and tissue inhibitors of metalloproteinases (TIMPs) leads to further corneal degradation. Amniotic membrane homogenate (AMH) has proteolytic properties beneficial for corneal healing, but it is unknown whether AMH possesses TIMPs or effectively inhibits MMP-2 and MMP-9 activity. Objective: To determine if bovine and equine AMH reduce in vitro MMP-2 and MMP-9 activities associated with the presence of TIMPs. Methods: Undiluted and diluted twofold series (0-fold to 16-fold dilutions) of equine ...
Bae JH, Ahn K, Nam GH, Lee CE, Park KD, Lee HK, Cho BW, Kim HS.The horse BMAL1 gene encodes the brain and muscle Arnt-like protein 1, which is a key regulator of circadian rhythmic systems in most organs and cells. The first exon of the horse-specific BMAL1 gene is produced by an exonization event of LINE3 (CR1) and SINE (MIR) was detected by bioinformatic analysis. Alternative variants generated by cassette exon event in various horse tissues were also detected by RT-PCR amplification and sequencing. The cDNA sequences of the horse transcripts (BMAL1a, BMAL1b) contain additional 21 bp and 71 bp fragments relative to horse BMAL1. Quantitative real-time RT...
Vachon AM, Keeley FW, McIlwraith CW, Chapman P.Articular cartilage specimens from the distal articular surface of 32 radiocarpal bones from 24 2- to 5-year-old horses were analyzed. The total collagen content was determined on the basis of the 4-hydroxyproline content, using a colorimetric method. A method for estimating the proportions of types-I and -II collagen by measuring spectrophotometric densities of specific cyanogen bromide peptide bands from mixtures of types-I and -II collagen on sodium dodecyl sulfate-polyacrylamide gels was used. The cyanogen bromide peptides representative of each collagen types-I and -II were identified. Th...
De Martin I, Theoret CL.To describe immunolocalization of TGF-beta receptors (RI and RII) in normal equine skin and in thoracic or limb wounds, healing normally or with exuberant granulation tissue (EGT). Methods: Group A: six wounds on one metacarpus and one midthoracic area. Group B: six wounds on both metacarpi, one of which was bandaged to stimulate EGT. Immunohistochemistry was used to detect RI and RII expression in wound margins. Methods: Eight horses, randomly assigned to one of two study groups. Methods: Neutralizing polyclonal anti-rabbit RI and RII antibodies were used to detect spatial expression of RI an...
Liu Y, Uboh CE, Soma LR, Li X, Guan F, You Y, Rudy JA, Chen JW.In 2008, Pennsylvania (PA) became the first State in the USA to ban and enforce the ban on the use of anabolic and androgenic steroids (AAS) in equine athletes by using plasma for analysis. To enforce the ban, a rapid and high-throughput method for analysis of 60 AAS in equine plasma was developed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Analytes were recovered from plasma by liquid-liquid extraction (LLE) using methyl tert-butyl ether, separated on a reversed-phase C₁₈ column and analyzed by electrospray ionization mass spectrometry. Multiple-reaction monitoring (MRM)...
Stefańska I, Witkowski L, Rzewuska M, Dzieciątkowski T.Rhodococcus equi is the causative agent of rhodococcosis in horses, resulting in significant morbidity and mortality in foals. This bacterium has also been isolated from a variety of animals and is being increasingly reported as a cause of infection in humans, mainly in immunosuppressed individuals. Laboratory diagnostics of R. equi infections based only on conventional microbiological methods shows low accuracy and can lead to misidentification. The objective of the study was to develop and evaluate a real-time PCR assay for direct detection of R. equi in various clinical specimens, including...
Lunn DP, Schram BR, Vagnoni KE, Schobert CS, Horohov DW, Ginther OJ.Lymphokine activated killing (LAK) is an example of natural cytotoxicity, and as such is a critical means of defense against diseases such as viral infection and neoplasia. Despite this important role, the specific molecular interactions involved in LAK or other forms of natural cytotoxicity are only partially understood. In some species, cells capable of mediating natural cytotoxicity express the CD8 molecule, although no specific role has been demonstrated for CD8 in non-MHC restricted cytotoxicity. In this study the role of the EqCD8 equine homolog of CD8 in LAK cell activity was examined. ...
Hinson JA, Neal RA.The kinetics of the horse liver alcohol dehydrogenase (alcohol: NAD+ oxidoreductase EC 1.1.1.1) catalyzed metabolism of octanol and octanal to octanoic acid have been examined. On incubation of octanol with horse liver alcohol dehydrogenase in the presence of NAD+, NADH as well as octanal and octanoic acid were seen as the initial products. However, on continued incubation, the octanal concentration progressively decreased to where only negligible quantities were present in the incubation after 10 min. The production of NADH was biphasic. An initial phase was followed in about 2 min with a slo...
Uboh CE, Rudy JA, Railing FA, Enright JM, Shoemaker JM, Kahler MC, Shellenberger JM, Kemecsei Z, Das DN.Although urine is the sample of choice for drug tests in racehorses, it is rarely obtained following the sudden death of a racehorse on the track while racing. The purpose of this study was to demonstrate the significance of postmortem tissue samples as an alternative to urine and blood samples in equine drug analysis following the sudden death of a racehorse on the track while participating in a competitive race. Postmortem tissue samples were frozen (-80 degrees C) until analyzed. A 30-40-g portion of each organ was homogenized in a 0.1 M phosphate buffer (pH 7.4), deproteinized, hydrolyzed ...
Collinder E, Berge GN, Grønvold B, Lindholm A, Midtved T, Norin E.This study investigated the influence of zinc bacitracin on the intestinal flora of horses. The functionally active intestinal flora was examined in 6 horses during treatment with zinc bacitracin. Utilising gas chromatography, spectrophotometry, gel electrophoresis and paper chromatography, samples were analysed on biochemical markers reflecting the action of parts of the intestinal flora. The following 5 flora-related functions were studied in faecal samples and intestinal samples from different sections of the hindgut: conversion of cholesterol to coprostanol and of bilirubin to urobilinogen...
Recio P, López PG, Fernández JL, Garcia-Sacristán A.1. The presence and types of alpha and beta-adrenoceptors in the corpus cavernosum of the horse were studied in vitro by using selected ligands of adrenoceptors and isometric tension recording. 2. Noradrenaline and phenylephrine induced concentration-dependent contractions in corpus cavernosum preparations. B-HT 920 had no effect. 3. Phentolamine and prazosin produced a shift to the right of the dose-response curve of noradrenaline, while the alpha(2)-antagonist, rauwolscine had no effect on the response to noradrenaline. Phenylephrine-evoked contractions of corporal strips were significantly ...
Simpson KS, Adams MH, Behrendt-Adam CY, Baker CB, McDowell KJ.For development to proceed normally, the appropriate genes must be expressed in the correct tissues and in the correct time frame. Knowledge of gene expression during development provides information about the changes taking place within the conceptus as well as possible reasons for pregnancy failure. However, little is known about gene expression during development in the equine conceptus. In this study, we examined differences in gene expression between day 12 and day 15 equine conceptuses by suppression subtractive hybridization. This technique was used to isolate transcripts that are more ...
Giles C, Vanniasinkam T, Barton M, Mahony TJ.Equine adenovirus 2 (EAdV-2) is one of two serotypes of adenoviruses known to infect equines. Initial studies did not associate EAdV-2 infections with any specific clinical syndromes, although more recent evidence suggests that EAdV-2 may be associated with clinical and subclinical gastrointestinal infections of foals and adults respectively. In contrast, Equine adenovirus 1 is well recognised as a pathogen associated with upper respiratory tract infections of horses. In this study the complete genome sequence of EAdV-2 is reported. As expected, genes common to the adenoviruses were identified...
Black JA, Leaf G.1. The effects of cyanogen bromide on horse-heart cytochrome c and horse-heart myoglobin have been investigated. Cytochrome c yielded four fragments, of which two were haemopeptides. The two colourless peptides had amino acid compositions corresponding to those that are expected, on the basis of the sequence proposed for horse-heart cytochrome c by Margoliash, Smith, Kreil & Tuppy (1961), from cleavage at both methionine residues. Of the two haemopeptides, one was isolated and shown to be that derived from cleavage at only one methionine residue, that nearer to the C-terminus of the peptid...
Lapo RA, Gogny M, Chatagnon G, Lalanne V, Harfoush K, Assane M, Desfontis JC, Mallem MY.This work was designed to investigate (i) the effect of superoxide dismutase (SOD) inhibition on endothelial function and (ii) the free radical-induced endothelial dysfunction in equine digital veins (EDVs) and equine digital arteries (EDAs) isolated from healthy horses. EDV and EDA rings were suspended in a 5 ml organ bath containing Krebs solution. After a 60 min equilibration period, EDV and EDA rings were contracted with phenylephrine. Then, cumulative concentration-response curves (CCRCs) to acetylcholine were performed. In both EDVs and EDAs, acetylcholine (1 nM to 10 µM) produced conce...
Larrasa J, Garcia A, Ambrose NC, Alonso JM, Parra A, de Mendoza MH, Salazar J, Rey J, de Mendoza JH.Dermatophilus congolensis is the pathogenic actinomycete that causes dermatophilosis in cattle, lumpy wool in sheep and rain scald in horses. Phenotypic variation between isolates has previously been described, but its genetic basis, extent and importance have not been investigated. Standard DNA extraction methods are not always successful for D. congolensis due to its complex life cycle, one stage of which is encapsulated. Here we describe the development of rapid and reliable DNA extraction and random amplified polymorphic DNA (RAPD) methods that can be used for genotyping D. congolensis fie...
Stammers JP, Hull D, Silver M, Fowden AL, Ousey J, Rossdale PD.An in vitro incubation technique was used to examine release of lipids from the equine placenta. Placental tissue was obtained at term (n = 5, term = 320-365 days) and earlier in gestation (n = 8, mean = 266 days). Term placentae were incubated at two temperatures, 4 degrees C (control) and 37 degrees C for 2 h. Pre-term placentae were incubated at 37 degrees C with two different concentrations of fatty acid in the medium. Tissues and media were analysed for their lipid concentrations. Term and pre-term placentae released free fatty acid (FFA) and phospholipid into the incubation medium during...
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...
Weinstein WL, Moore PA, Sanchez S, Dietrich UM, Wooley RE, Ritchie BW.To determine whether a novel third-generation chelating agent (8 mM disodium EDTA dehydrate and 20 mM 2-amino-2-hydroxymethyl-1, 3-propanediol) would act as an antimicrobial potentiator to enhance in vitro activity of antifungal medications against fungal isolates obtained from horses with mycotic keratitis. Methods: Fungal isolates (3 Aspergillus isolates, 5 Fusarium isolates, 1 Penicillium isolate, 1 Cladosporium isolate, and 1 Curvularia isolate) obtained from horses with mycotic keratitis and 2 quality-control strains obtained from the American Type Culture Collection (ATCC; Candida albica...
Morris DD, Harmon BG, Moore JN.A study was performed to determine whether equine antiserum to core lipopolysaccharide (LPS) would enhance phagocytosis of smooth gram-negative (GN) organisms by equine macrophages. Five healthy adult horses (group A) were immunized with a bacterin prepared from the J-5 mutant of Escherichia coli 0111:B4 and Salmonella minnesota R595 to produce antibodies to core LPS. Five horses (group B) served as nonimmunized controls and were given physiologic saline solution instead of the rough mutant bacterin. Serum antibody titers to core LPS and to smooth E coli 0111:B4 were determined by indirect ELI...
Delbeke FT, Debackere M.The prototype of a commercial ELISA test kit designed for fentanyl determination in human urine has been evaluated for screening fentanyl in horse urine and plasma. The measurement of fentanyl after intravenous (2 mg) and intramuscular (0.25 mg) administration in undiluted plasma was not reproducible while accurate quantification of fentanyl in urine greatly depends on the composition of the horse urine. The ELISA assay, however, is simple and could be successfully used for quantitative measurements in diluted urine and for rapid qualitative screening for fentanyl in large numbers of urine sam...
