Immunofluorescence assay (IFA) is a laboratory technique used to detect and visualize specific antigens or antibodies in equine tissue samples or bodily fluids. This method employs fluorescent-labeled antibodies to bind target molecules, allowing for the observation of fluorescence under a microscope. In horses, IFA is employed in various research and diagnostic applications, including the study of infectious diseases, immune responses, and cellular localization of proteins. The technique provides valuable insights into the distribution and expression of specific proteins within equine cells and tissues. This page aggregates peer-reviewed research studies and scholarly articles that explore the methodology, applications, and advancements of immunofluorescence assay in equine research.
Carter RA, Engiles JB, Megee SO, Senoo M, Galantino-Homer HL.Abnormal epidermal stem cell regulation may contribute to the pathogenesis of equine chronic laminitis. Objective: To analyse the involvement of p63, a regulator of epidermal stem cell proliferative potential, in chronic laminitis. Methods: Epidermal tissues from skin, coronet and lamellae of the dorsal foot were harvested from 5 horses with chronic laminitis and 5 control horses. Tissues were analysed using histopathology, immunofluorescence microscopy and quantitative immunoblotting. Results: Hoof lamellae of laminitic horses had a lower frequency of p63 positive cells than control lamellae,...
Bedford-Guaus SJ, McPartlin LA, Xie J, Westmiller SL, Buffone MG, Roberson MS.Oocyte activation at fertilization is brought about by the testis-specific phospholipase C zeta (PLCZ), owing to its ability to induce oscillations in intracellular Ca(2+) concentration ([Ca(2+)](i)). Whereas this is a highly conserved mechanism among mammals, important species-specific differences in PLCZ sequence, activity, and expression have been reported. Thus, the objectives of this research were to clone and characterize the intracellular Ca(2+)-releasing activity and expression of equine PLCZ in sperm and testis. Molecular cloning of equine PLCZ yielded a 1914-bp sequence that translat...
Tulloch LK, Perkins JD, Piercy RJ.Skeletal muscle is composed of a heterogeneous mixture of several fibre types, each with specific physiological properties. In equine muscle, identification of these individual fibres (fibre typing) is important for both exercise physiology and pathological studies. Traditionally, fibre typing has been achieved by adenosine triphosphatase (ATPase) histochemistry or by immunoperoxidase labelling with antibodies directed at myosin heavy chain isoforms. ATPase histochemistry can be temperamental and lacks specificity, and both techniques require staining of serial cryosections to reveal the entir...
Deschene K, Céleste C, Boerboom D, Theoret CL.As a transient hypoxic state exists within skin wounds in horses and may be important for the healing process, this study sought to identify a molecular hypoxia response occurring in horse limb and body wounds healing by second intention. Hypoxia-inducible factor 1α (HIF1α) protein expression was studied throughout repair by Western blotting and immunofluorescence. Paradoxically, HIF1α was strongly expressed in intact skin and its expression decreased dramatically following wounding (p<0.01), despite the expected hypoxic state within the wounded tissue. HIF1α levels reincreased in parallel...
Oue Y, Ishihara R, Edamatsu H, Morita Y, Yoshida M, Yoshima M, Hatama S, Murakami K, Kanno T.A new equine coronavirus was isolated from the feces of adult horses with pyrogenic and enteric disease. The disease outbreak was mainly observed among 2- to 4-year-old horses living in stables of a draft-horse racetrack in Japan. On comparing the isolated virus (isolate Tokachi09) with the equine coronavirus NC99 strain, no significant differences were observed in several biological properties such as hemagglutinating activity, antigenicity (in indirect immunofluorescence and neutralization tests), and one-step growth (in cell culture). The sequences of the nucleocapsid and spike genes of iso...
Zoonoses and public healthJanuary 5, 2011
Volume 58, Issue 6 416-423 doi: 10.1111/j.1863-2378.2010.01382.x
Toledo RS, Tamekuni K, Filho MF, Haydu VB, Barbieri AR, Hiltel AC, Pacheco RC, Labruna MB, Dumler JS, Vidotto O.Spotted fever is a disease caused by bacteria from the genus Rickettsia of the spotted fever group (SFG). Rickettsia rickettsii is likely the main agent of Brazilian spotted fever (BSF). With the objective of gathering information on the circulation of SFG rickettsiae in Londrina, Parana state, ticks from dogs and horses and also blood from dogs, horses and humans were collected in a neighbourhood of the city which presented potential for circulation of rickettsiae between hosts and vectors. Amblyomma cajennense, Dermacentor nitens, and Rhipicephalus sanguineus ticks were subjected to Polymera...
Tamekuni K, Toledo Rdos S, Silva Filho Mde F, Haydu VB, Pacheco RC, Cavicchioli JH, Labruna MB, Dumler JS, Vidotto O.Brazilian spotted fever (BSF) is an emerging disease most likely caused by Rickettsia rickettsii. The objective of the present study was to estimate the seroprevalence of BSF rickettsia infections in equines from six horse farms located in Londrina County, Paraná, Southern Brazil. Six owners of horse farms situated in Cambé, Santa Fé, Guaraci and Londrina municipalities participated in the study. All farms were located in areas where BSF has not been reported. A total of 273 horses were sampled and their sera were tested by indirect immunofluorescence assay (IFA) using R. rickettsii and R. ...
Batista FG, Silva DM, Green KT, Tezza LB, Vasconcelos SP, Carvalho SG, Silveira I, Moraes-Filho J, Labruna MB, Fortes FS, Molento MB.Brazilian Spotted Fever (BSF) is a lethal rickettsiosis in humans caused by the bacteria Rickettsia rickettsii, and is endemic in some areas of Brazil. Horses and dogs are part of the disease's life cycle and they may also serve as sentinel animals in epidemiological studies. The first human BSF case in the State of Paraná was reported in 2005. The present study was conducted in the municipality of Almirante Tamandaré, where no previous case of BSF was reported. Serum samples were collected from 71 horses and 20 dogs from nine properties in the area. Ticks were also collected from these anim...
Brandt S, Tober R, Corteggio A, Burger S, Sabitzer S, Walter I, Kainzbauer C, Steinborn R, Nasir L, Borzacchiello G.In equids, bovine papillomaviruses of type 1 (BPV-1) and less frequently type 2 induce common, locally aggressive skin tumours termed sarcoids. Whereas BPV infection in cattle usually involves the epidermis and is productive in this skin layer, infection in equids is currently thought to be abortive, with virus solely residing as multiple episomes in dermal fibroblasts. Based on recent observations that do not agree with this assumption, we hypothesised that BPV also infects equid epidermis and is active in this skin layer. To test this hypothesis, we conducted a proof-of-principle study on ei...
Prause AS, Guionaud CT, Stoffel MH, Portier CJ, Mevissen M.To evaluate the expression of the 5-hydroxytryptamine 4 (5-HT4) receptor subtype and investigate the modulating function of those receptors on contractility in intestinal tissues obtained from horses without gastrointestinal tract disease. Methods: Smooth muscle preparations from the duodenum, ileum, and pelvic flexure collected immediately after slaughter of 24 horses with no history or signs of gastrointestinal tract disease. Methods: In isometric organ baths, the contractile activities of smooth muscle preparations in response to 5-hydroxytryptamine and electric field stimulation were asses...
Bermúdez CS, Zaldívar AY, Spolidorio MG, Moraes-Filho J, Miranda RJ, Caballero CM, Mendoza Y, Labruna MB.The present research evaluated the presence of Rickettsia spp. on ectoparasites of horses and dogs (using PCR techniques), and their sera (using immunofluorescence assay) in El Valle de Antón town in Panama. A total of 20 horses and 20 dogs were sampled, finding four species of ectoparasites on dogs (the ticks Rhipicephalus sanguineus, Amblyomma ovale, Amblyomma oblongoguttatum, and the flea Ctenocephalides felis), and two tick species on horses (Amblyomma cajennense and Dermacentor nitens). DNA of Rickettsia amblyommii was found in pools of A. cajennense, D. nitens, and R. sanguineus, while ...
Pacheco RC, Moraes-Filho J, Guedes E, Silveira I, Richtzenhain LJ, Leite RC, Labruna MB.The present study was performed in an area endemic for Brazilian spotted fever (BSF) in Juiz de Fora, state of Minas Gerais, Brazil, during the years 2007 and 2008, when fatal cases of BSF (caused by Rickettsia rickettsii) were reported. Adult ticks (Acari: Ixodidae) identified as Rhipicephalus sanguineus (Latreille) and Amblyomma cajennense (Fabricius) were collected from dogs and horses, respectively, and tested by polymerase chain reaction (PCR). Overall, 13.1% of the Rh. sanguineus ticks and none of the A. cajennense were found to be infected with R. rickettsii. Two isolates of R. ricketts...
Pickles KJ, Mair JA, Lopez-Villalobos N, Shaw DJ, Scott I, Pomroy W.Cyathostomins are the principal pathogenic nematode of equidae worldwide. In other species mast cell (MC) proteinases, in particular chymases, appear to have protective roles. Knowledge of the equine intestinal immune response to cyathostomins is limited. Objective: To investigate MC numbers and proteinase expression in equine cyathostomin-infected large intestine. Objective: MC populations in the large intestine are positively associated with cyathostomin burden and predominantly express chymase. Methods: The caecal cyathostomin burden of naturally infected horses (n = 25) was determined by l...
Burton AJ, Nydam DV, Dearen TK, Mitchell K, Bowman DD, Xiao L.To date, little is known about the prevalence, genotypes and zoonotic potential of Cryptosporidium spp. affecting horses, especially in North America. A cross-sectional study was conducted in New York, USA between February 25th and May 1st 2009. Fecal samples were collected from three hundred and forty nine 1-10-week-old foals and their dams on 14 different broodmare farms. All fecal samples were screened for Cryptosporidium spp. using a direct immunofluorescence assay (DFA). DNA extraction and PCR-RFLP analysis of the small-subunit (SSU) rRNA gene were performed on all the foal samples. PCR-p...
Dai X, Hilsen RE, Hu WG, Fulton RE.An electrochemiluminescence (ECL) immunoassay, incorporating chemically biotinylated and ruthenylated antibodies down-selected from a panel of monoclonal and polyclonal reagents, was developed to detect and identify Venezuelan equine encephalitis virus (VEEV). The limit of detection (LOD) of the optimized ECL assay was 10(3)pfu/ml VEEV TC-83 virus and 1 ng/ml recombinant (r) VEEV E2 protein. The LOD of the ECL assay was approximately one log unit lower than that of a sandwich enzyme-linked immunosorbent assay (ELISA) incorporating the same immunoreagents. Repetition of ECL assays over time and...
Aquino LP, Machado RZ, Lemos KR, Marques LC, Garcia MV, Borges GP.The present research investigated the presence of T. evansi antibodies in animals from the subregion of Nhecolandia, in the Pantanal Sul-mato-grossense, by means of an enzyme linked immunosorbent assay (ELISA) and indirect immunofluorescence antibody test (IFAT), and the pattern of polypeptide recognition by sera from experimentally and naturally infected hosts using Western blotting. Serum samples were obtained from bovines (n = 102), horses (n = 98), and dogs (n = 55), and from 32 free-ranging coatis (Nasua nasua). None of the bovines were found positive, while sera from 16 dogs (29%) and 23...
Freitas MC, Grycajuk M, Molento MB, Bonacin J, Labruna MB, Pacheco Rde C, Moraes-Filho J, Deconto I, Biondo AW.Brazilian Spotted Fever (BSF) is an often fatal zoonosis caused by the obligate intracellular bacterium Rickettsia rickettsii. The disease is generally transmitted to humans by Amblyomma spp. ticks. Serological evidence of past infection by R. rickettsii has been reported in horses, but the pathogenicity of R. rickettsii in horses remains unknown. Cart horses are still widely used in urban and urban fringe areas in Brazil, and these animals may constitute suitable sentinels for BSF human in these areas, for example, in Sao Jose dos Pinhais, where the first BSF human case in the state of Parana...
Carter RA, Shekk V, de Laat MA, Pollitt CC, Galantino-Homer HL.The dermo-epidermal interface that connects the equine distal phalanx to the cornified hoof wall withstands great biomechanical demands, but is also a region where structural failure often ensues as a result of laminitis. The cytoskeleton in this region maintains cell structure and facilitates intercellular adhesion, making it likely to be involved in laminitis pathogenesis, although it is poorly characterized in the equine hoof lamellae. The objective of the present study was to identify and quantify the cytoskeletal proteins present in the epidermal and dermal lamellae of the equine hoof by ...
Kurtz BM, Singletary LB, Kelly SD, Frampton AR.In this study, Equus caballus major histocompatibility complex class I (MHC-I) was identified as a cellular entry receptor for the alphaherpesvirus equine herpesvirus type 1 (EHV-1). This novel EHV-1 receptor was discovered using a cDNA library from equine macrophages. cDNAs from this EHV-1-susceptible cell type were inserted into EHV-1-resistant B78H1 murine melanoma cells, these cells were infected with an EHV-1 lacZ reporter virus, and cells that supported virus infection were identified by X-Gal (5-bromo-4-chloro-3-indolyl-beta-d-galactopyranoside) staining. Positive cells were subjected t...
Spolidorio MG, Labruna MB, Machado RZ, Moraes-Filho J, Zago AM, Donatele DM, Pinheiro SR, Silveira I, Caliari KM, Yoshinari NH.Blood samples collected from 201 humans, 92 dogs, and 27 horses in the state of Espirito Santo, Brazil, were tested by polymerase chain reaction, indirect immunofluorescence assays, and indirect enzyme-linked immunosorbent assay for tick-borne diseases (rickettsiosis, ehrlichiosis, anaplasmosis, borreliosis, babesiosis). Our results indicated that the surveyed counties are endemic for spotted fever group rickettsiosis because sera from 70 (34.8%) humans, 7 (7.6%) dogs, and 7 (25.9%) horses were reactive to at least one of the six Rickettsia species tested. Although there was evidence of ehrlic...
Loftus JP, Williams JM, Belknap JK, Black SJ.Laminitis is a crippling disease of horses characterized by an inflammatory response in the tissue that suspends the axial skeleton within the hoof. Pain is a common feature of laminitic pathology and its management is an important component of the treatment regime for this disease. Systemic lidocaine administration is commonly utilized to manage pain in equine laminitis; however, the potential anti-inflammatory effects of this drug during the treatment of equine laminitis have not been investigated. Here, we sought to determine if lidocaine concentrations achieved in the plasma (therapeutic c...
Albrizio M, Lacalandra GM, Micera E, Guaricci AC, Nicassio M, Zarrilli A.Opioid receptors and endogenous opioid peptides act not only in the control of nociceptive pathways, indeed several reports demonstrate the effects of opiates on sperm cell motility and morphology suggesting the importance of these receptors in the modulation of reproduction in mammals. In this study we investigated the expression of delta opioid receptors on equine spermatozoa by western blot/indirect immunofluorescence and its relationship with sperm cell physiology. Methods: We analyzed viability, motility, capacitation, acrosome reaction and mitochondrial activity in the presence of naltri...
Groppelli E, Tuthill TJ, Rowlands DJ.Equine rhinitis A virus (ERAV) is genetically closely related to foot-and-mouth disease virus (FMDV), and both are now classified within the genus Aphthovirus of the family Picornaviridae. For disease security reasons, FMDV can be handled only in high-containment facilities, but these constraints do not apply to ERAV, making it an attractive alternative for the study of aphthovirus biology. Here, we show, using immunofluorescence, pharmacological agents, and dominant negative inhibitors, that ERAV entry occurs (as for FMDV) via clathrin-mediated endocytosis and acidification of early endosomes...
Visser MB, Pollitt CC.Most research to date involving laminins and extracellular matrix protein function in both normal and pathological conditions involves in vitro culture of keratinocytes. Few methods are established to allow for prolonged propagation of keratinocytes from equine tissues, including the hoof lamellae. In this study we modified cell isolation and culture techniques to allow for proliferation and sub-culturing of equine lamellar keratinocytes. Additionally, the production and processing of extracellular matrix molecules by skin and lamellar keratinocytes were studied. Results: Physical and proteoly...
Zhang J, Stein DA, Timoney PJ, Balasuriya UB.A significant consequence of equine arteritis virus (EAV) infection of horses is persistence of the virus in a variable percentage of infected stallions. We recently established an in vitro model of EAV persistence in cell culture for the purpose of furthering our understanding of EAV biology in general and viral persistence in the stallion in particular. In this study we investigated whether persistently infected HeLa cells could be cured of EAV infection by treatment with an antisense peptide-conjugated phosphorodiamidate morpholino oligomer (PPMO) designed to target the 5'-terminal region o...
Sigg L, Gerber V, Gottstein B, Doherr MG, Frey CF.In Switzerland, the prevalence and incidence of equine piroplasma parasite (EPP) infections are unknown. In order to obtain a first insight into the prevalence, a representative sample of 689 sera of horses from Switzerland was serologically tested for the presence of antibodies directed against T. equi and B. caballi using the Indirect Fluorescence Antibody Test (IFAT). A total of 50 (7.3%) horses were seropositive for EPP: overall, the seroprevalence of T. equi was significantly higher than that of B. caballi (p=0.002). The seropositivities in indigenous horses (animals bred and raised in Sw...
Kalinová Z, Cisláková L, Halánová M.Ehrlichiosis and anaplasmosis are zoonoses caused by bacteria from the family Anaplasmataceae, including human and animal pathogens. The human pathogens are Ehrlichia chaffeensis, the causative agent of human monocytic ehrlichiosis (HME), Anaplasma phagocytophilum, the pathogen causing human granulocytic anaplasmosis (HGA), E. ewingii and Neorickettsia sennetsu, granulocytotropic and monocytotropic Ehrlichia species, respectively. Ehrlichia spp. are small, gram-negative, obligate intracellular bacteria. They replicate in the cytoplasmic vacuoles of host cells, especially granulocytes and monoc...
Breuil MF, Duquesne F, Sévin C, Laugier C, Petry S.Contagious equine metritis is a horse disease that causes endometrial inflammation due to Taylorella equigenitalis. Since Taylorella asinigenitalis was characterized, genital swab culture has proved to be an insufficient method for distinguishing between the two Taylorella species. Here, we developed an indirect immunofluorescence (IIF) test using polyclonal antibodies. Specificity, sensitivity, and detection limit were assessed using isolated bacteria (55 T. equigenitalis strains, 46 T. asinigenitalis strains and 18 other bacterial species), experimental and genital swabs in comparison to bac...
Jaffer O, Abdishakur F, Hakimuddin F, Riya A, Wernery U, Schuster RK.A total of 105 serum samples from endurance horses from different stables in Dubai were examined for the presence of antibodies against Theileria equi and Babesia caballi using immunofluorescence antibody test (IFAT) and competitive enzyme-linked immunosorbent assay (cELISA). A TaqMan real-time polymerase chain reaction (PCR) was used to detect DNA of piroplasms in specimens of clotted blood or EDTA blood samples of the same animals. Out of the 105 serum samples, the IFAT detected antibodies against T. equi in 35 (33.3%) cases while the cELISA gave 34 (32.4%) positive results. Eleven (10.5%) o...
de Mattos Carvalho A, Alves AL, Golim MA, Moroz A, Hussni CA, de Oliveira PG, Deffune E.The purpose of this work was to isolate and cultivate mesenchymal stem cells (MSC) derived from equine adipose tissue and conduct cellular characterization with the following markers: CD90, CD44 and CD13. Adipose tissue collection was performed at the base of the horses' tails, followed by immediate isolation and cultivation of the MSC and posterior characterization by flow cytometry for the interspecies reaction test using mouse anti-rat CD90 monoclonal antibody (mAb), fluorescein isothiocyanate (FITC), and tests with specific mAb mouse anti-horse CD13 and mouse anti-horse CD44. The technique...
House C, Mikiciuk PE, Berninger ML.Five serological methods of diagnosing African horse sickness were evaluated, using a battery of serum samples from experimental horses vaccinated and challenged with each serotype of African horse sickness virus (AHSV1 through AHSV9): agar gel immunodiffusion (AGID), indirect fluorescent antibody (IFA), complement fixation (CF), virus neutralization (VN), and enzyme-linked immunosorbent assay (ELISA). The 5 tests were also compared using a panel of field samples, convalescent equine sera with antibodies to domestic equine viral diseases, and sera from horses awaiting export. The ELISA describ...
Heras S, Smits K, De Schauwer C, Van Soom A.Global epigenetic reprogramming is considered to be essential during embryo development to establish totipotency. In the classic model first described in the mouse, the genome-wide DNA demethylation is asymmetric between the paternal and the maternal genome. The paternal genome undergoes ten-eleven translocation (TET)-mediated active DNA demethylation, which is completed before the end of the first cell cycle. Since TET enzymes oxidize 5-methylcytosine to 5-hydroxymethylcytosine, the latter is postulated to be an intermediate stage toward DNA demethylation. The maternal genome, on the other ha...
Sabeur K, Vo AT, Ball BA.Angiotensin II is a hormone with a wide array of physiological effects that exerts its effect via interaction with two major subtypes of receptor. The results of this study show that angiotensin II (from 1 to 100 nmol l(-1)) initiates acrosomal exocytosis in equine spermatozoa that have undergone capacitation in vitro in a TALP-TEST (Tyrode's albumin lactate pyruvate; 188.7 mmol TES l(-1), 84.8 mmol Tris l(-1)) buffer with cAMP. The acrosome reaction and sperm viability were assessed with fluorescein isothiocyanate-Pisum sativum agglutinin (FITC-PSA) and Hoechst 33258, respectively. The initia...
Lombardi F, Palumbo P, Augello FR, Giusti I, Dolo V, Guerrini L, Cifone MG, Giuliani M, Cinque B.The ability of a collagen-based matrix to support cell proliferation, migration, and infiltration has been reported; however, the direct effect of an aqueous collagen suspension on cell cultures has not been studied yet. In this work, the effects of a high-concentration aqueous suspension of a micronized type I equine collagen (EC-I) have been evaluated on a normal mouse fibroblast cell line. Immunofluorescence analysis showed the ability of EC-I to induce a significant increase of type I and III collagen levels, parallel with overexpression of crucial proteins in collagen biosynthesis, matura...
Choi YH, Ross P, Velez IC, Macías-García B, Riera FL, Hinrichs K.Equine embryos develop in vitro in the presence of high glucose concentrations, but little is known about their requirements for development. We evaluated the effect of glucose concentrations in medium on blastocyst development after ICSI. In experiment 1, there were no significant differences in rates of blastocyst formation among embryos cultured in our standard medium (DMEM/F-12), which contained >16 mM glucose, and those cultured in a minimal-glucose embryo culture medium (<1 mM; Global medium, GB), with either 0 added glucose for the first 5 days, then 20 mM (0-20) or 20 mM for th...
Sarkar S, Balasuriya UB, Horohov DW, Chambers TM.Equine herpesvirus-1 (EHV-1) infects equine endothelial cells (EECs) lining the small blood vessels in the central nervous system. However, the effect of type I IFN on EHV-1 replication in the EECs is not well studied. Thus, the primary objective of this study was to investigate the effect of type-I IFN on the replication of the neuropathogenic T953 strain of EHV-1 in vitro in EECs. The initial data showed that the EHV-1 was partly resistant to the biological effect of exogenously supplied recombinant equine IFN-α. Subsequent investigation into the mechanism of resistance showed that EHV-1 in...
Vieira TS, Vieira RF, Krawczak FS, Soares HS, Guimarães AM, Barros-Filho IR, Marcondes M, Labruna MB, Biondo AW, Vidotto O.Although well established in dogs, Ehrlichia sp. infection has been scarcely reported in horses. The aim was to perform a comprehensive serological and molecular survey for the detection of Ehrlichia spp. in carthorses from Southern Brazil. Blood samples from 190 carthorses from Paraná State were sampled. Horses were also tested for Borrelia burgdorferi and Anaplasma phagocytophilum. Anti-Ehrlichia sp. antibodies were detected by a commercial rapid ELISA, and immunofluorescence antibody assays (IFA) with E. chaffeensis and E. canis as crude antigens. The molecular and phylogenetic analysis of...
Batista FG, Silva DM, Green KT, Tezza LB, Vasconcelos SP, Carvalho SG, Silveira I, Moraes-Filho J, Labruna MB, Fortes FS, Molento MB.Brazilian Spotted Fever (BSF) is a lethal rickettsiosis in humans caused by the bacteria Rickettsia rickettsii, and is endemic in some areas of Brazil. Horses and dogs are part of the disease's life cycle and they may also serve as sentinel animals in epidemiological studies. The first human BSF case in the State of Paraná was reported in 2005. The present study was conducted in the municipality of Almirante Tamandaré, where no previous case of BSF was reported. Serum samples were collected from 71 horses and 20 dogs from nine properties in the area. Ticks were also collected from these anim...
Marsh AE, Hyun C, Barr BC, Tindall R, Lakritz J.Equine protozoal myeloencephalitis (EPM), caused by a protozoal parasite infection of the central nervous system, is the most commonly diagnosed neurologic disease of horses in North America. In specific regions of the United States approximately 50% of the horse population is seropositive to Sarcocystis neurona. However, not all seropositive horses develop clinical signs. Detailed clinical examination, along with cerebrospinal fluid antibody evaluation are often used to diagnose EPM. Postmortem evaluation of the brain stem and spinal cord for histopathologic lesions compatible with nonsuppura...
Cruz Villagrán C, Amelse L, Neilsen N, Dunlap J, Dhar M.Mesenchymal stromal cells (MSCs) are able to differentiate into extramesodermal lineages, including neurons. Positive outcomes were obtained after transplantation of neurally induced MSCs in laboratory animals after nerve injury, but this is unknown in horses. Our objectives were to test the ability of equine MSCs to differentiate into cells of neural lineage in vitro, to assess differences in morphology and lineage-specific protein expression, and to investigate if horse age and cell passage number affected the ability to achieve differentiation. Bone marrow-derived MSCs were obtained from yo...
Vernon SD, Webb PA.We developed an enzyme-linked immunosorbent assay (ELISA) that was capable of detecting immunoglobulin M (IgM) antibody to vesicular stomatitis virus (VSV) in the sera of experimentally and naturally infected cattle and horses. The detection of IgM in the sera of these animals permitted an estimate of the recency of infection by VSV serotype New Jersey. A VSV serotype New Jersey epizootic strain isolated from a horse and passed once in an Aedes albopictus cell line was used to infect a horse and a calf. Sera from these animals were used to standardize the ELISA. This assay was used to test ser...
Wu F, Zhang S, Zhang Y, Mo R, Yan F, Wang H, Wong G, Chi H, Wang T, Feng N, Gao Y, Xia X, Zhao Y, Yang S.Ebola virus infections lead to severe hemorrhagic fevers in humans and nonhuman primates; and human fatality rates are as high as 67%-90%. Since the Ebola virus was discovered in 1976, the only available treatments have been medical support or the emergency administration of experimental drugs. The absence of licensed vaccines and drugs against the Ebola virus impedes the prevention of viral infection. In this study, we generated recombinant baculoviruses (rBV) expressing the Sudan virus (SUDV) matrix structural protein (VP40) (rBV-VP40-VP40) or the SUDV glycoprotein (GP) (rBV-GP-GP), and SUDV...
Sponseller BT, Valberg SJ, Tennent-Brown BS, Foreman JH, Kumar P, Timoney JF.Four Quarter Horses (9 months to 7 years of age) with submandibular lymphadenopathy and firm muscles (palpation of which elicited signs of pain) were evaluated; in general, the horses had a stiff gait, and 3 horses became recumbent. Streptococcus equi was cultured from aspirates of lymph nodes or samples of purulent material collected from the auditory tube diverticula. Once the horses were recumbent, their condition deteriorated rapidly despite aggressive antimicrobial and antiinflammatory treatment, necessitating euthanasia within 24 to 48 hours. One horse did not become recumbent and recove...
de Castro RVG, Pieri NCG, Fantinato Neto P, Grizendi BM, Dória RGS, Meirelles FV, Smith LC, Garcia JM, Bressan FF.The cellular reprogramming into pluripotency is influenced by external and internal cellular factors, such as culture conditions (e.g., environmental oxygen concentration), and the aging process. Herein, we aimed to generate and maintain equine iPSCs (eiPSCs) derived from fibroblasts of a horse older than 20 years and to evaluate the effect of different levels of oxygen tension (atmospheric 20% O, 5% O, or 20% to 5% O) on these cells. Fibroblasts were reprogrammed, and putative eiPSCs were positive for positive alkaline phosphatase detection; they were positive for pluripotency-related genes ...
Levkutová M, Hípiková V, Faitelzon S, Benath G, Paulík S, Levkut M.Infection with the intracellular microsporidium Encephalitozoon cuniculi can cause a serious disease--encephalitozoonosis in various animals and people. Several species of mammals, including the horse, were seem to be potential sources of encephalitozoonosis for animal as well as human hosts. The disease diagnosis is based on clinical signs, pathological findings, and the detection of E. cuniculi or circulating antibodies directed against the parasite. This study investigates the seroconversion to E. cuniculi in horses admitted to the Veterinary Teaching Hospital of the Hebrew University of Je...
Knowles EJ, Moreton-Clack MC, Shaw S, Harris PA, Elliott J, Menzies-Gow NJ.Analysis of plasma adrenocorticotropic hormone concentration [ACTH] aids diagnosis of pituitary pars intermedia dysfunction (PPID). Comparisons of the validated chemiluminescent-immunoassay (CI) and immunofluorescent (IF) assays are limited. Objective: To compare the results of [ACTH] analysis by CI and IF methods of samples collected in autumn and spring and assess cross-reactivity. Methods: Method comparison. Methods: Plasma from nonlaminitic ponies was analysed concurrently using the IF and CI methods in autumn and the following spring. Diagnostic thresholds for the IF method were derived u...
Wang L, Pawlak E, Johnson PJ, Belknap JK, Alfandari D, Black SJ.To determine whether increased gene expression of a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4) in laminae of horses with starch gruel-induced laminitis was accompanied by increased enzyme activity and substrate degradation. Methods: Laminae from the forelimb hooves of 8 healthy horses and 17 horses with starch gruel-induced laminitis (6 at onset of fever, 6 at onset of Obel grade 1 lameness, and 5 at onset of Obel grade 3 lameness). Methods: Gene expression was determined by use of cDNA and real-time quantitative PCR assay. Protein expression and processing were ...
Xiao C, Liu Y, Jiang Y, Magoffin DE, Guo H, Xuan H, Wang G, Wang LF, Tu C.Four monoclonal antibodies (mAbs) were generated by immunizing BALB/C mice with recombinant nucleocapsid protein (N) of Nipah virus (NiV) and Hendra virus (HeV) expressed in E. coli. Two mAbs each were obtained for the HeV N and NiV N, respectively. All four mAbs displayed specific reactivity with the recombinant N proteins of both viruses by western blot, which was further confirmed by immunofluorescent antibody assay using fixed insect cells infected with recombinant baculoviruses expressing either the HeV or NiV N protein. Epitope mapping using a 12-mer random peptide phage display library ...
Słońska A, Cymerys J, Godlewski MM, Dzieciątkowski T, Tucholska A, Chmielewska A, Golke A, Bańbura MW.Equine herpesvirus type 1 (EHV-1) causes respiratory disease, abortion and neurological disorders in horses. In the present study, we investigated reorganization of the cytoskeleton in neurons infected with two EHV-1 strains: Jan-E (wild-type strain) and Rac-H (attenuated strain). The studies were performed on primary murine neurons, which are an excellent model for studying neurotropism and neurovirulence of EHV-1. We have demonstrated for the first time that EHV-1 infection causes rearrangements in the actin network of neurons that are dependent on the virus strain and its adaptation to cell...
Cole DJ, Snowden K, Cohen ND, Smith R.Feces collected from three asymptomatic horses and seeded with Cryptosporidium parvum oocysts (10(1) to 10(6)/g of feces) were evaluated by acid-fast staining (AF), an immunofluorescent antibody (IFA) technique, and flow cytometry. The thresholds of detection were 5 x 10(5) oocysts/g of feces for the IFA and AF techniques and 5 x 10(4) oocysts/g for flow cytometry.
Soule C, Dupouy-Camet J, Georges P, Ancelle T, Gillet JP, Vaissaire J, Delvigne A, Plateau E.Three groups of three horses each were, respectively, infected with 5000, 20,000 and 50,000 larvae of Trichinella spiralis. The strain used was isolated from a human biopsy during horsemeat-related outbreaks of trichinellosis in France. Transient muscular disorders were only observed in two of the horses infected with 50,000 larvae but none of the horses had fever. A significant increase in blood eosinophils was noticed in 5 horses. Serum LDH, aldolase and CPK peaked at the fifth week post-infection. Specific IgG assayed by indirect immunofluorescence and ELISA, appeared 2-5 weeks post-infecti...
Jakobsen JR, Schjerling P, Svensson RB, Buhl R, Carstensen H, Koch M, Krogsgaard MR, Kjær M, Mackey AL.The myotendinous junction (MTJ) is a specialized interface for transmitting high forces between the muscle and tendon and yet the MTJ is a common site of strain injury with a high recurrence rate. The aim of this study was to identify previously unknown MTJ components in mature animals and humans. Samples were obtained from the superficial digital flexor (SDF) muscle-tendon interface of 20 horses, and the tissue was separated through a sequential cryosectioning approach into muscle, MTJ (muscle tissue enriched in myofiber tips attached to the tendon), and tendon fractions. RT-PCR was performed...
Galvão CM, Rezende-Gondim MM, Chaves AC, Schares G, Ribas JR, Gondim LF.Donkeys (Equus asinus) are closely related to horses and are known to be infected by several equine pathogens. Neospora caninum and Neospora hughesi are protozoan parasites that infect horses, but they were not confirmed in donkeys up to this date. The aim of this study was to evaluate the exposure of donkeys (Equus asinus) to Neospora spp. using tachyzoites of N. caninum as antigen and employing two common serologic methods, IFAT and immunoblot. Sera from 500 donkeys were obtained from 30 municipalities in Bahia state and tested by IFAT. Two of 500 sera were positive for Neospora spp. by IFAT...
Echeverría MG, Pecoraro MR, Galosi CM, Etcheverrigaray ME, Nosetto EO.This paper describes the first isolation of equine arteritis virus (EAV) in Argentina. The virus was isolated from the semen of an imported seropositive stallion held in isolation at a breeding farm in Tandil in the Buenos Aires Province. In addition, viral nucleic acid was detected in seminal plasma using the reverse-transcription polymerase chain reaction. The isolated virus was propagated in cell cultures and confirmed as EAV by indirect immunofluorescence and virus neutralisation, using a serum specific for the reference Bucyrus strain of EAV. As far as the authors are aware, this is the f...
MacEachern KE, Smith GL, Nolan AM.Equine endothelial cells were isolated from the pulmonary artery by enzymatic digestion and grown to confluency. The cells were characterised by positive immunofluorescent staining for von Willebrand factor and NADPH-diaphorase staining for nitric oxide synthase. Measurements of endothelins indicated that there were significant release rates from the cells for up to six hours. Measurements of intracellular calcium concentration showed that the application of bradykinin caused a transient increase in calcium concentration with similar characteristics to those observed in other endothelial cell ...
Duarte PC, Daft BM, Conrad PA, Packham AE, Saville WJ, MacKay RJ, Barr BC, Wilson WD, Ng T, Reed SM, Gardner IA.The objectives of this study were to evaluate the accuracy of the indirect fluorescent antibody test (IFAT) using serum and cerebrospinal fluid (CSF) of horses naturally and experimentally infected with Sarcocystis neurona, to assess the correlation between serum and CSF titers, and to determine the effect of S. neurona vaccination on the diagnosis of infection. Using receiver-operating characteristic analysis, the areas under the curve for the IFAT were 0.97 (serum) and 0.99 (CSF). Sensitivity and specificity were 83.3 and 96.9% (serum, cutoff 80) and 100 and 99% (CSF, cutoff 5), respectively...
Vasić A, Răileanu C, Körsten C, Vojinović D, Manić M, Urošević A, Nikolić N, Dulović O, Tews BA, Petrović T, Silaghi C, Valčić M....West Nile virus (WNV) is a zoonotic mosquito-borne virus classified as family Flaviviridae and genus Flavivirus. The first WNV outbreak in humans in the Republic of Serbia was recorded in 2012. Equids and dogs can show clinical symptoms after WNV infection and are often used as sentinels. This study aimed to (i) give insight into seropositivity for WNV in clinically healthy dog and horse sera in different regions of Serbia and (ii) compare diagnostic value of 'in-house' and commercially available indirect immunofluorescence (IFA) and enzyme-linked immunoassay (ELISA) tests to 'gold standard' v...
Albrizio M, Lacalandra GM, Micera E, Guaricci AC, Nicassio M, Zarrilli A.Opioid receptors and endogenous opioid peptides act not only in the control of nociceptive pathways, indeed several reports demonstrate the effects of opiates on sperm cell motility and morphology suggesting the importance of these receptors in the modulation of reproduction in mammals. In this study we investigated the expression of delta opioid receptors on equine spermatozoa by western blot/indirect immunofluorescence and its relationship with sperm cell physiology. Methods: We analyzed viability, motility, capacitation, acrosome reaction and mitochondrial activity in the presence of naltri...
Kornicka K, Al Naem M, Röcken M, Zmiertka M, Marycz K.Osteochondritis dissecans (OCD) in equids, especially in sport horses, has become a growing issue as it contributes to the occurrence of lameness. Thus the aim of this study was to investigate the cytophysiological properties of OCD chondrocytes including expression of chondrogenic genes, apoptosis, mitochondria dynamics and autophagy. Horse chondrocytes were isolated from healthy (HE) and OCD cartilages. Properties of cells were evaluated using multiple assays e.g., polymerase chain reaction (PCR), immunofluorescence, Western blot. OCD chondrocytes were characterized by increased apoptosis an...
Albrizio M, Guaricci AC, Maritato F, Sciorsci RL, Mari G, Calamita G, Lacalandra GM, Aiudi GG, Minoia R, Dell'Aquila ME, Minoia P.The development of fertilizing ability in sperm cells is associated with changes in the plasma membrane. However, to date the exact nature of sequentially activated primary receptors and channels and the signal transduction pathways derived from these remains elusive. We analyzed the expression and localization of the mu-opioid receptor in equine spermatozoa. A transcript corresponding to the third extracellular loop that selectively binds mu agonists was amplified, sequenced and compared with the known sequences in humans, rats and cattle. The amplification product showed a high degree of nuc...
