Polymerase Chain Reaction (PCR) is a molecular biology technique used to amplify specific DNA sequences, allowing for detailed genetic analysis in horses. This method enables the detection and quantification of genetic material, facilitating research in areas such as genetic disorders, infectious diseases, and population genetics in equine species. PCR applications in horses include identifying pathogens, verifying parentage, and studying genetic variations. The technique's sensitivity and specificity make it a valuable tool in equine veterinary diagnostics and research. This page compiles peer-reviewed research studies and scholarly articles that explore the applications, methodologies, and advancements of PCR in equine science.
EHV1 and EHV4 are the most important herpesviruses in horses. Repeated cases of abortion in mares regularly vaccinated, prompted us to investigate the immune response after vaccination with the same inactivated vaccine, but with three different protocols. Eighteen mares were chosen and randomly divided in three study groups (G-G-G) and a control group (Ctrl). For serologic and PCR investigations nasal swabs, sera and blood were collected. The protocol used in G (4 doses) increased the titer recorded by ELISA and seroneutralization (SN). Poor agreement and no correlation were observed in titer ...
Akter R, Legione A, Sansom FM, El-Hage CM, Hartley CA, Gilkerson JR, Devlin JM.Equine abortion is a cause of severe economic loss to the equine industry. Equine herpesvirus 1 is considered a primary cause of infectious abortion in horses, however other infectious agents can also cause abortion. Abortions due to zoonotic pathogens have implications for both human and animal health. We determined the prevalence of Coxiella burnetii, Leptospira spp. and Toxoplasma gondii in 600 aborted equine foetal tissues that were submitted to our diagnostic laboratories at the University of Melbourne from 1994 to 2019. Using qPCR we found that the prevalence of C. burnetii was 4%. The h...
Onen EA.The aim of this study was to evaluate formalin-inactivated autovaccination to treat cutaneous papillomatosis and to perform molecular typing of the papillomavirus in four horses (two foals, one 3-year-old filly and a 5-year-old stallion). Methods: Histopathological slides of lesions were prepared and stained with haematoxylin and eosin (H&E) to establish a diagnosis that was based on observation koilocytosis, which is a pathognomonic cytopathic change that is associated with papillomatosis, using light microscopy. Polymerase chain reaction (PCR) and DNA sequencing were performed using the ...
Salco R, Bowers J, Hernandez V, Barnum S, Pusterla N.This study aimed to determine if the administration of a modified live equine influenza virus vaccine (FluAvert) to foals would positively impact their health and reduce colonization of their upper airways with equine herpesviruses (EHV) during the weaning period. A single dose of FluAvert was given to 20 healthy foals 7 days prior to being weaned; 20 healthy foals served as unvaccinated controls. Nasal secretions and blood were collected before vaccination, the day of weaning, and weekly thereafter for 3 weeks. Nasal secretions were tested by quantitative polymerase chain reaction (qPCR) for ...
Apprich V, Licka T, Freiler S, Gabriel C.Impaired keratinocyte differentiation has recently been suggested as a key event in equine hoof canker development. Koilocytotic appearance of keratinocytes, one of the most characteristic morphological alterations in hoof canker tissue, is also a common marker for papillomavirus (PV) infection, and bovine PV-1 and/or -2 (BPV-1/2) has previously been detected in equine canker patients. Therefore, the present study aimed to correlate the frequency and severity of koilocytotic keratinocytes with BPV detection in hoof canker samples. Hoof tissue of 5/18 canker-affected horses and 2/6 control hors...
Paulino PG, Almosny N, Oliveira R, Viscardi V, Müller A, Guimarães A, Baldani C, da Silva C, Peckle M, Massard C, Santos H.This study aims to report the presence of Neorickettsia risticii DNA in blood samples from naturally infected horses in Rio de Janeiro, provide clinicopathological findings related to the infection, and report the phylogenetic diversity of the 16S rDNA of N. risticii in order to evaluate its heterogeneity. Real-time quantitative polymerase chain reaction (qPCR) was performed to investigate the presence of N. risticii in samples collected from horses (n = 187). Five positive samples were found in the molecular screening. Hypoalbuminemia and high levels of creatine kinase and lactate dehydro...
Tartor YH, Hamad MH, Abouzeid NZ, El-Belkemy FA.Equine pythiosis is an emerging, devastating disease that is hard to treat. The tumour-like nodular skin masses grow rapidly and the outcome is generally fatal, and thus early diagnosis and intervention are important. Objective: (i) To highlight the clinical, histological and haematological findings in pythiosis, and (ii) to evaluate the efficacy of direct sample multiplex-PCR targeting the single nucleotide polymorphisms within the ribosomal DNA region for detection and genotyping of Pythium insidiosum. Methods: Two hundred and twenty horses including 204 Arabian and 16 draft horses were surv...
Lab on a chipApril 26, 2020
Volume 20, Issue 9 1621-1627 doi: 10.1039/d0lc00304b
Sun F, Ganguli A, Nguyen J, Brisbin R, Shanmugam K, Hirschberg DL, Wheeler MB, Bashir R, Nash DM, Cunningham BT.Rapid, sensitive and specific detection and reporting of infectious pathogens is important for patient management and epidemic surveillance. We demonstrated a point-of-care system integrated with a smartphone for detecting live virus from nasal swab media, using a panel of equine respiratory infectious diseases as a model system for corresponding human diseases such as COVID-19. Specific nucleic acid sequences of five pathogens were amplified by loop-mediated isothermal amplification on a microfluidic chip and detected at the end of reactions by the smartphone. Pathogen-spiked horse nasal swab...
Tozaki T, Ohnuma A, Kikuchi M, Ishige T, Kakoi H, Hirota KI, Kusano K, Nagata SI.Gene doping, an activity which abuses and misuses gene therapy, is a major concern in sports and horseracing industries. Effective methods capable of detecting and monitoring gene doping are urgently needed. Although several PCR-based methods that detect transgenes have been developed, many of them focus only on a single transgene. However, numerous genes associated with athletic ability may be potential gene-doping material. Here, we developed a detection method that targets multiple transgenes. We targeted 12 genes that may be associated with athletic performance and designed two TaqMan prob...
Santagostino M, Piras FM, Cappelletti E, Del Giudice S, Semino O, Nergadze SG, Giulotto E.Interstitial telomeric sequences (ITSs) are short stretches of telomeric-like repeats (TTAGGG)n at nonterminal chromosomal sites. We previously demonstrated that, in the genomes of primates and rodents, ITSs were inserted during the repair of DNA double-strand breaks. These conclusions were derived from sequence comparisons of ITS-containing loci and ITS-less orthologous loci in different species. To our knowledge, insertion polymorphism of ITSs, i.e., the presence of an ITS-containing allele and an ITS-less allele in the same species, has not been described. In this work, we carried out a gen...
Hedberg-Alm Y, Penell J, Riihimäki M, Osterman-Lind E, Nielsen MK, Tydén E.All grazing horses are exposed to intestinal parasites, which have the potential to cause gastrointestinal disease. In Sweden, there is a concern about an increase in parasite-related equine gastrointestinal disease, in particular , since the implementation of prescription-only anthelmintics approximately 10 years ago. In a prospective case-control study, parasitological status, using fecal analyses for strongyle egg counts, the presence of eggs and Polymerase chain reaction (PCR) as well as serology for , were compared between horses presenting with or without gastrointestinal disease at a ...
Lambertini C, Bombardi C, Zannoni A, Bernardini C, Dondi F, Morini M, Rinnovati R, Spadari A, Romagnoli N.Proteinase activated receptor 4 (PAR) in the gastrointestinal tract is involved in the regulation of inflammation and pain pathways. The aim of the present study was to evaluate the distribution and expression of PAR in the jejunum of healthy horses and in the pathologic tracts from horses undergoing surgery for herniation of the small intestine through the epiploic foramen. Eight healthy horses (Group H) and eight horses with epiploic hernia (Group EH) were included; the jejunum samples were collected at the slaughter or intraoperatively after enterectomy, respectively. To evaluate PAR expres...
Vengust M, Jager MC, Zalig V, Cociancich V, Laverack M, Renshaw RW, Dubovi E, Tomlinson JE, Van de Walle GR, Divers TJ.Equine parvovirus-hepatitis (EqPV-H) has been proposed as the aetiological cause of Theiler's disease, also known as serum hepatitis. EqPV-H-associated Theiler's disease has not been previously reported in Europe. Objective: To determine whether EqPV-H infection was associated with a 2018-2019 outbreak of Theiler's disease in four horses on a studfarm. Methods: Descriptive case series. Methods: The medical records of four horses from the same farm diagnosed with fatal Theiler's disease were examined retrospectively. Information collected included a clinical history, physical examination findin...
Benfodil K, Büscher P, Abdelli A, Van Reet N, Mohamed-Herif A, Ansel S, Fettata S, Dehou S, Bebronne N, Geerts M, Balharbi F, Ait-Oudhia K.Trypanosoma evansi (T. evansi) is a hemoflagellate parasite that affects a broad range of mammalian hosts and that causes a disease called surra. Diagnosis of surra based on clinical symptoms alone is inaccurate. Therefore, a variety of serological and molecular diagnostic tests are used to assist in the detection of T. evansi infections. The aim of this study was to compare the diagnostic performance of four serological tests (CATT/T.evansi, immune trypanolysis, ELISA with purified variant surface glycoprotein RoTat 1.2 and with whole cell lysate) and two molecular PCR tests targeting sequenc...
Avila VA, López-García Y, Hernández-Castro R, Salas-Garrido CG, Ramírez-Lezama J, Calderón-Villa R, Martínez-Chavarría LC.Aberrant nematode larval migration in the CNS of horses is rare but frequently fatal; one of the main etiological agents involved in this illness is Halicephalobus gingivalis. This soil nematode has been associated with several fatal equine meningoencephalitis reports worldwide; however, it had never been diagnosed in horses of Mexico. A 10 year-old Andalusian horse presented dysphagia, fever, weakness, prostration and ataxia; the patient expired during the medical attention. Post mortem examination was performed and no gross alterations were found. Histopathology revealed meningoencephalitis...
Alloway E, Linder K, May S, Rose T, DeLay J, Bender S, Tucker A, Luff J.Squamous cell carcinoma (SCC) is the most common neoplasm of the equine stomach. However, the mechanisms underlying malignant transformation are unknown. As papillomavirus-2 (EcPV-2) is a likely cause of some genital SCCs, we hypothesized that EcPV-2 is associated with a subset of equine gastric SCCs. To this aim, we performed polymerase chain reaction (PCR) and in situ hybridization (ISH) for EcPV-2 E6/ E7 oncogenes on 11 gastric SCCs and on gastric samples from 15 control horses with no SCC. PCR for EcPV-2 was positive in 7/11 (64%) gastric SCCs; non-SCC gastric samples were all negative. I...
Balasuriya UBR.The primary goals of this chapter are to discuss common viral RNA isolation and purification methods that are routinely used by various diagnostic laboratories and to highlight the advantages and drawbacks of each method and to identify the most suitable and reliable method to increase the sensitivity and specificity of RT-PCR assays for the detection of equine influenza virus (EIV) in clinical specimens. Our experiences and review of literature show that magnetic bead-based nucleic extraction methods (manual and automatic) work well for isolation and purification of EIV RNA from nasal swab sp...
Balasuriya UBR.Equine influenza (EI) is a highly contagious disease of horses caused by the equine influenza virus (EIV) H3N8 subtype. EI is the most important respiratory virus infection of horses and can disrupt major equestrian events and cause significant economic losses to the equine industry worldwide. Influenza H3N8 virus spreads rapidly in susceptible horses and can result in very high morbidity within 24-48 h after exposure to the virus. Therefore, rapid and accurate diagnosis of EI is critical for implementation of prevention and control measures to avoid the spread of EIV and to reduce the econom...
Noll LW, Stoy CPA, Wang Y, Porter EG, Lu N, Liu X, Burklund A, Peddireddi L, Hanzlicek G, Henningson J, Chengappa MM, Bai J.Streptococcus equi subsp. equi is a Gram positive bacterial pathogen commonly associated with strangles in horses, a respiratory disease characterized by abscessation of submandibular and retropharyngeal lymph nodes which can lead to obstruction of the airway. Several real-time PCR (qPCR) assays have been developed for detection of S. equi from horses with many targeting conserved regions of the S. equi cell wall-associated M-protein (SeM), a major virulence factor and immunogen of S. equi. Our objective was to develop a nested PCR (nPCR) targeting SeM and an 18S rRNA internal control gene for...
Lei R, Wang X, Zhang D, Liu Y, Chen Q, Jiang N.Equine piroplasmosis (EP) is a severe disease of horses caused by the tick-borne protozoa Theileria equi (T. equi) and Babesia caballi (B. caballi). Infectious carriers are not always symptomatic, meaning there is a risk to non-enzootic areas. Regulatory tests for EP include sero-epidemiological methods for equine babesiosis, but these lack specificity due to cross-reactivity with other Babesia species. In this study, we present a real-time quantitative recombinase polymerase amplification (qRPA) method for fast simultaneous detection of both T. equi and B. caballi. In this method, primers and...
Wernery U, Joseph S, Raghavan R, Dyer B, Spendrup S.Our investigation has shown that multiple vaccinations with inactivated African horse sickness (AHS) vaccines containing all 9 serotypes and produced at the Central Veterinary Research Laboratory in Dubai, UAE, protect horses from AHS. However, the immunization did not prevent African horse sickness fever (AHSF) in approximately 10% of the vaccinated horses despite high enzyme-linked immunosorbent assay and virus neutralizing antibodies. African horse sickness fever is a very mild form of AHS with similar clinical signs. From all 6 horses which had developed AHSF, no virus was isolated from ED...
Lim QL, Tan YL, Ng WL, Yong CSY, Ismail A, Rovie-Ryan JJ, Rosli N, Annavi G.A molecular sexing method by polymerase chain reaction (PCR) amplification of a portion of the sex-determining region Y (SRY) and the zinc finger (ZF) gene, as well as six equine Y-chromosome-specific microsatellite markers, were tested in the Malayan tapir (Tapirus indicus). While the microsatellite markers did not yield any male-specific amplicons for sex-typing, the SRY/ZF marker system produced reliable molecular sexing results by accurately sex-typing 31 reference Malayan tapirs, using whole blood, dried blood spot (DBS), or tissue samples as materials for DNA extraction. The marker syste...
Tirosh-Levy S, Steinman A, Einhorn A, Apanaskevich DA, Mumcuoglu KY, Gottlieb Y.Theileria equi Mehlhorn and Schein, 1998 (Piroplasmida: Babesiidae) is an important tick-borne pathogen of horses that is highly endemic in many parts of the world, including Israel. The present study evaluated the potential roles of five hard tick species [Hyalomma excavatum Koch, 1844; Hyalomma marginatum Koch, 1844; Rhipicephalus turanicus Pomerantsev 1936; Rhipicephalus annulatus Say, 1821; Haemaphysalis parva (Neumann, 1897) (all: Ixodida: Ixodidae)], previously found to infest horses in Israel, in acting as vectors for piroplasmosis. For this, DNA was extracted from whole ticks and, when...
Tirosh-Levy S, Gottlieb Y, Mimoun L, Mazuz ML, Steinman A.Although the main route of transmission of is through tick feeding, transplacental transmission is also possible and may lead to abortion, or to the birth of a sick or carrier foal. The aim of this study was to evaluate the role of as a cause of abortions in Israel and the risk of foals being infected at a young age. Eight aborting mares were serologically evaluated for exposure to via the immunofluorescence antibody test (IFAT) and their aborted fetuses were evaluated using PCR and qPCR. In addition, five mares and their foals (aged 4-6 months) from a highly endemic farm were tested for i...
Sunday Idoko I, Tirosh-Levy S, Leszkowicz Mazuz M, Mohammed Adam B, Sikiti Garba B, Wesley Nafarnda D, Steinman A.Equine piroplasmosis (EP) is a tick-borne disease of equids, caused by the two haemoprotozoal parasites: and Nigeria constitutes a major crossroads of animal transport in West Africa and may serve as a factor in EP dissemination in the region. The study aim was to characterize EP parasites in donkeys and horses in northern Nigeria using a molecular approach. Blood was collected from 57 donkeys and 47 horses. EP infection was detected and characterized by polymerase chain reaction (PCR). Twenty five donkeys (43.8%) were infected with , five (8.8%) with three (5.3%) with dual infections. Four...
Nyaoke AC, Navarro MA, Fresneda K, Diab SS, Moore J, Lyras D, Awad M, Uzal FA.Enteric disease in horses may be caused by a variety of microorganisms, including several clostridial species. (previously ) has been frequently associated with gas gangrene in humans and several animal species, including horses. However, its role in enteric diseases of animals has not been fully determined. We describe herein 7 cases of enteric disease in horses associated with infection. Grossly, the small and/or large intestines were necrotic, hemorrhagic, and edematous. Microscopically, there was severe mucosal necrosis and hemorrhage of the small and/or large intestine of all horses. w...
Szczerbal I, Nowacka-Woszuk J, Kopp-Kuhlman C, Mackowski M, Switonski M.X monosomy is the most common disorder of sex development in horses. Although cytogenetic analysis is still the gold standard in the diagnosis of equine X monosomy, novel molecular techniques are being sought to quickly and reliably detect this chromosome abnormality. Objective: The goal of this study was to evaluate the usefulness of a novel variant of the PCR technique-namely, droplet digital PCR (ddPCR)-in the detection of X monosomy in mares. Methods: A proof of concept of the usefulness of ddPCR in diagnosing an abnormal number of X chromosomes in mares. Methods: We examined an infertile ...
Elata A, Mossaad E, Satti R, Matar N, Ohari Y, Xuan X, Inoue N, Suganuma K.In Sudan, donkeys are important animals, providing transportation and income possibilities. However, the prevalence of parasites in donkeys in Sudan has not been thoroughly characterized. Accordingly, in this study, we aimed to detect selected hemoprotozoan parasites in donkeys in West Omdurman, Khartoum State, Sudan, wherein people depend mainly on donkeys for their daily life. In total, 198 blood samples collected from donkeys in a local market in West Omdurman, were screened using serological and molecular diagnostic techniques. Serologically, 52 (26.3%), 56 (28.3%), and 19 (9.6%) samples w...
Ereqat S, Nasereddin A, Al-Jawabreh A, Al-Jawabreh H, Al-Laham N, Abdeen Z.Trypanosoma evansi is the causative agent of surra, a disease that occurs in many animal species. The disease is responsible for substantial losses in global production and can be fatal if not diagnosed early. This study aims to determine the prevalence of T. evansi in livestock, equids and dromedary camels in Palestine. Methods: Blood samples were collected during 2015-2017 from domesticated animals (n = 259 animals; 77% females and 23% males) including camels (n = 87), horses (n = 46), donkeys (n = 28), mules (n = 2), sheep (n = 49) and goats (n = 48) from eight districts: Ariha (Jericho), N...
Andrade DGA, Basso RM, Castiglioni MCR, Silva JP, Machado VMV, Laufer-Amorim R, Borges AS, Oliveira-Filho JP.Four causative mutations (D1, D2, D3*, and D4) of chondrodysplastic dwarfism have been described in the equine () gene. Homozygotes for one of these mutations and heterozygotes for any combination of these mutations exhibit the disproportionate dwarfism phenotype. However, no case description of homozygotes for D4 (D4/D4) has been reported in the literature, to our knowledge. We report 2 Miniature horses with the genotype D4/D4 in the gene. Clinically, the 2 dwarfs had a domed head that was large compared to the rest of the body, mandibular prognathism, and short and bowed limbs, mainly in t...
Cursino AE, Vilela APP, Franco-Luiz APM, de Oliveira JG, Nogueira MF, Júnior JPA, de Aguiar DM, Kroon EG.Equine infectious anemia (EIA) has a worldwide distribution, and is widespread in Brazil. The Brazilian Pantanal presents with high prevalence comprising equine performance and indirectly the livestock industry, since the horses are used for cattle management. Although EIA is routinely diagnosed by the agar gel immunodiffusion test (AGID), this serological assay has some limitations, so PCR-based detection methods have the potential to overcome these limitations and act as complementary tests to those currently used. Considering the limited number of equine infectious anemia virus (EIAV) seque...
Boyle AG, Rankin SC, D￮ L, Boston RC, Wheeler-Aceto H.Bacterial culture and polymerase chain reaction (PCR) assays for the detection of Streptococcus equi in nasopharyngeal washes (NPW) and guttural pouch lavage (GPL) samples have low sensitivity. In human diagnostics, processing of samples with flocked swabs has improved recovery rates of bacterial agents because of improved surface area and elution factors. Objective: For S. equi subsp. equi (S. equi) detection in NPW and GPL samples we hypothesized that: direct-PCR would be more reliable than flocked swab culture (FS culture); flocked swab PCR (FS-PCR) would be equivalent to direct-PCR; and FS...
Howerth EW, Mead DG, Mueller PO, Duncan L, Murphy MD, Stallknecht DE.Horses were inoculated with Vesicular stomatitis New Jersey and Indiana viruses by routes simulating contact and vector transmission. Clinical signs, lesions, antibody development, viral shedding and persistence, and viremia were monitored. Horses were infected with both viruses by all routes as confirmed by seroconversion. Salivation, primary lesions at inoculation sites, and secondary oral lesions were the most common clinical findings. Viral shedding was most often from the oral cavity, followed by the nasal cavity; titers were highest from oral cavity samples. Virus was rarely isolated fro...
Cullinane A, Quinlivan M, Nelly M, Patterson H, Kenna R, Garvey M, Gildea S, Lyons P, Flynn M, Galvin P, Neylon M, Jankowska K.In 2006 there was an outbreak of equine infectious anaemia (EIA) in Ireland. This paper describes the use of the diagnosis of clinical and subclinical cases of the disease. In acute cases the ELISAs and the immunoblot were more sensitive than the AGID. In one mare, fluctuating antibody levels were observed in all the serological assays before it seroconverted by AGID. Viral RNA and DNA were detected by RT-PCR and PCR in all the tissues from the infected animals examined postmortem. The PCR detected viral DNA in plasma regardless of the stage of the disease. In contrast, the RT-PCR detected RNA...
Biswas B, Mukherjee D, Mattingly-Napier BL, Dutta SK.Genomic amplification by the polymerase chain reaction (PCR) was used to identify a unique genomic sequence of Ehrlichia risticii directly in DNA isolated from peripheral-blood buffy coat cells of E. risticii-infected horses (Potomac horse fever) and from infected cell cultures. A specific primer pair, selected from a cloned, species-specific, 1-kb DNA fragment of the E. risticii genome as a template, was used for the amplification of the target DNA of 247 bp. The optimal number of 40 PCR cycles, determined by analyzing an amplification profile obtained with a constant Taq polymerase concentra...
Coultous RM, Phipps P, Dalley C, Lewis J, Hammond TA, Shiels BR, Weir W, Sutton DGM.Equine piroplasmosis (EP) has historically been of minor concern to UK equine practitioners, primarily due to a lack of competent tick vectors. However, increased detection of EP tick vector species in the UK has been reported recently. EP screening is not currently required for equine importation, and when combined with recent relaxations in movement regulations, there is an increased risk regarding disease incursion and establishment into the UK. This study evaluated the prevalence of EP by both serology and PCR among 1242 UK equine samples submitted for EP screening between February and Dec...
Semevolos SA, Nixon AJ, Brower-Toland BD.To determine molecular changes in the expression of insulin-like growth factor-I (IGF-I) and transforming growth factor-beta1 (TGF-beta1) in horses with osteochondrosis, and to characterize expression of matrix aggrecan and collagen types I, II, and X in articular cartilage of affected joints. Methods: Articular cartilage from affected stifle or shoulder joints of 11 horses with naturally acquired osteochondrosis and corresponding joints of 11 clinically normal horses. Methods: Harvested specimens were snap frozen in liquid nitrogen, and total RNA was isolated. Specimens were fixed in 4% paraf...
Galuppi R, Piva S, Castagnetti C, Sarli G, Iacono E, Fioravanti ML, Caffara M.This paper describes the transmission of a zoonotic subtype of Cryptosporidium parvum between two foals hospitalized in an Equine Perinatology Unit (EPU) linked to an outbreak of cryptosporidiosis in veterinary students. Fecal specimens of 36 mares (105 samples) and 28 foals (122 samples) were subjected to Ziehl-Neelsen staining, nested PCR of 18S rDNA. Two foals tested positive for Cryptosporidium; PCR restriction fragment length polymorphism (PCR-RFLP) analysis and subtyping by nested PCR of the 60kDa glycoprotein (gp60) gene revealed C. parvum subtype IIdA23G1. The introduction of Cryptospo...
Pusterla N, Madigan JE, Leutenegger CM.The focus of rapid diagnosis of infectious disease of horses in the last decade has shifted from the conventional laboratory techniques of antigen detection, microscopy, and culture to molecular diagnosis of infectious agents. Equine practitioners must be able to interpret the use, limitations, and results of molecular diagnostic techniques, as they are increasingly integrated into routine microbiology laboratory protocols. Polymerase chain reaction (PCR) is the best-known and most successfully implemented diagnostic molecular technology to date. It can detect slow-growing, difficult-to-cultiv...
Evers F, Garcia JL, Navarro IT, Zulpo DL, Nino Bde S, Ewald MP, Pagliari S, Almeida JC, Freire RL.This study aimed to investigate anti-Toxoplasma gondii antibodies and to isolate the parasite from the brains of horses processed at slaughterhouses in Brazil. We collected brain and blood samples from 398 horses of various ages, from six Brazilian states. Serum samples were evaluated by indirect fluorescent antibody test (IFAT cut-off titre ≥ 1:64), and brains were submitted to mouse bioassay. Among the 398 horses, positivity for T. gondii was identified in 46 (11.6%) by IFAT and in 14 (3.5%) by mouse bioassay. In 12 of those 14 bioassays, mice were positive only by IFAT (cut-off titre ≥ ...
Alves Ada S, Melo AL, Amorim MV, Borges AM, Gaíva E Silva L, Martins TF, Labruna MB, Aguiar DM, Pacheco RC.The aim of the study was to evaluate exposure of equids to rickettsial agents (Rickettsia rickettsii, Rickettsia parkeri, 'Candidatus Rickettsia amblyommii', Rickettsia rhipicephali, and Rickettsia bellii) and rickettsial infection in ticks of a Pantanal region of Brazil. Sera of 547 equids (500 horses and 47 donkeys) were evaluated by indirect immunofluorescence assay. In total, 665 adults and 106 nymphal pools of Amblyomma cajennense F. sensu lato, 10 Dermacentor nitens Neumann ticks, and 88 larval pools of Amblyomma sp. were tested by polymerase chain reaction (PCR). Overall, 337 (61.6%) eq...
Abdel-Shafy S, Abdullah HHAM, Elbayoumy MK, Elsawy BSM, Hassan MR, Mahmoud MS, Hegazi AG, Abdel-Rahman EH.Piroplasmosis and anaplasmosis are serious tick-borne diseases (TBDs) that are concerning for the public and animal health. This study aimed to detect the molecular prevalence and epidemiological risk factors of Piroplasma and Anaplasma species in animal hosts and their associated ticks in Egypt. A total of 234 blood samples and 95 adult ticks were collected from animal hosts (112 cattle, 38 sheep, 28 goats, 26 buffaloes, 22 donkeys, and 8 horses) from six provinces of Egypt (AL-Faiyum, AL-Giza, Beni-Suef, Al-Minufia, Al-Beheira, and Matruh). Blood and tick samples were investigated by polymer...
Taktaz Hafshejani T, Nekoei S, Vazirian B, Doosti A, Khamesipour F, Anyanwu MU.This study was undertaken to investigate molecularly the occurrence of EHV-1 and EHV-4 infection among equine population in regions, Iran. Blood samples from 53 and 37 randomly selected horses settled in Isfahan and Shahrekord, Iran, respectively, were collected. Detection of EHV-1 and EHV-4 genes in the blood samples was done using polymerase chain reaction (PCR). Out of 53 and 37 samples from Isfahan and Shahrekord, 4 (18.18%) and 3 (8.10%) were positive for PCR of EHV-1, respectively. Nine (16.98%) and 6 (16.21%) were positive for PCR of EHV-4, while 6 (11.32%) and 3 (8.10%) were positive f...
Yeh JY, Lee JH, Seo HJ, Park JY, Moon JS, Cho IS, Lee JB, Park SY, Song CS, Choi IS.The aim of this study was to develop a highly sensitive and specific one-step duplex reverse transcriptase PCR (RT-PCR) assay for the simultaneous and differential detection of West Nile (WNV) and Japanese encephalitis (JEV) viruses. The bioinformatic analysis of published sequences of WNV and JEV revealed conserved regions not targeted by previously reported primers. A total of 13 primers were designed based on these regions to detect all of the WNV and JEV lineages and to discriminate between the two viruses by the generation of 482- and 241-bp cDNA products, respectively. The results indica...
Martens A, De Moor A, Demeulemeester J, Peelman L.To examine apparently normal skin around equine sarcoids for evidence of bovine papilloma virus (BPV) DNA, and to relate this finding to the observed recurrence after surgery. Methods: Prospective study. Methods: Forty-one equine sarcoids from 19 horses. Methods: The tumors were surgically excised at a measured distance of 8, 12, or 16 mm. Samples from the tumor and of the entire surrounding skin were taken at 4, 8, 12, and 16 mm from the tumor border and analyzed for the presence of BPV DNA using polymerase chain reaction (PCR) amplification. The samples were grouped per examined sarcoid, and...
Fortier LA, Balkman CE, Sandell LJ, Ratcliffe A, Nixon AJ.This study evaluated the constitutive insulin-like growth factor-I (IGF-I) gene expression pattern in spontaneously healing cartilage defects over the course of 16 weeks, and correlated the tissue morphology and matrix gene expression with IGF-I mRNA levels. Full-thickness 15 mm cartilage defects were debrided in the femoral trochlea of both femoropatellar joints of 8 horses and the healing defects examined 2, 4, 8, or 16 weeks after surgery. Samples were harvested for histologic assessment of tissue healing using H&E staining, toluidine blue histochemical reaction for proteoglycan deposition,...
Akter R, Legione A, Sansom FM, El-Hage CM, Hartley CA, Gilkerson JR, Devlin JM.Equine abortion is a cause of severe economic loss to the equine industry. Equine herpesvirus 1 is considered a primary cause of infectious abortion in horses, however other infectious agents can also cause abortion. Abortions due to zoonotic pathogens have implications for both human and animal health. We determined the prevalence of Coxiella burnetii, Leptospira spp. and Toxoplasma gondii in 600 aborted equine foetal tissues that were submitted to our diagnostic laboratories at the University of Melbourne from 1994 to 2019. Using qPCR we found that the prevalence of C. burnetii was 4%. The h...
Pringle J, Venner M, Tscheschlok L, Bächi L, Riihimäki M.After strangles outbreaks, Streptococcus equi ssp. equi (S. equi) can persist in clinically normal silent carriers for months to years. Two naturally occurring outbreaks of strangles with 53 and 100% morbidity, respectively, were followed longitudinally to assess occurrence of carrier state and optimal detection methods Outbreak A involved 98 yearling warmbloods, and outbreak B 38 mature Icelandic horses. Fully recovered horses were sampled at least 6 months after index cases using nasal swabs (one sampling occasion only) nasopharyngeal lavage and guttural pouch visualisation and lavages for c...
Asif M, Parveen A, Ashraf S, Hussain M, Aktas M, Ozubek S, Shaikh RS, Iqbal F.The present study was designed to check the molecular detection of Anaplasma marginale and Theileria annulata in blood samples of horses and donkeys collected from Dera Ghazi Khan District in Punjab and to document their phylogenetic origin and their association with studied epidemiological factors (sex and age) and complete blood count parameters, if any. Results: A total of 195 blood samples were collected from apparently healthy horses (N = 141) and donkeys (N = 54). A. marginale DNA was detected by PCR in 4.9% (7/141) horse and in 9.2% (5/54) of donkey blood samples. Prevalence of ...
Sanz MG, Kwon S, Pusterla N, Gold JR, Bain F, Evermann J.Currently, diagnosis of equine coronavirus (ECoV) relies on the exclusion of other infectious causes of enteric disease along with molecular detection of ECoV in feces or tissue. Although this approach is complete, it is costly and may not always be achievable. Objective: We hypothesized that the overall fecal shedding of ECoV in hospitalized horses is low. Our objective was to determine whether systemically healthy horses and horses with gastrointestinal disorders shed ECoV in their feces at the time of admission to a referral hospital and after 48 hours of stress associated with hospitaliz...
Borchers K, Ebert M, Fetsch A, Hammond T, Sterner-Kock A.Equine herpes virus 2 (EHV-2), a gamma(2)-herpesvirus, is common in horses of all ages. Its role as a primary pathogen is unclear but there is an association between EHV-2, respiratory disease and keratoconjunctivitis. The purpose of this study was to gain more information on the prevalence of EHV-2 DNA in conjunctival swabs from horses with and without ocular disease and to define the anatomical site and cell type harbouring viral genome or antigen. By polymerase chain reaction (PCR) 22 out of 77 (28.6%) ocular swabs of clinically healthy and only 4 out of 48 (8.3%) samples from diseased hors...
Newkirk KM, Hendrix DV, Anis EA, Rohrbach BW, Ehrhart EJ, Lyons JA, Kania SA.Squamous cell carcinoma (SCC) is the most common tumor arising in the periocular and penile areas of horses. Both ultraviolet radiation and papillomaviruses have been implicated in the pathogenesis of SCC in various species, including the horse. This retrospective study used polymerase chain reaction (PCR) to detect papillomavirus DNA in archival biopsy samples from equine periocular and penile SCC from 3 different geographic areas (northeast, southeast, and central United States). Forty-two periocular SCCs were tested; none contained papillomavirus DNA. Twenty-two penile SCCs were tested, and...
Nemoto M, Yamanaka T, Bannai H, Tsujimura K, Kondo T, Matsumura T.Reverse transcription loop-mediated isothermal amplification (RT-LAMP) was applied to the detection of equine influenza virus (EIV). Because equine influenza is caused currently by EIV of the H3H8 subtype, the RT-LAMP primer set was designed to target the hemagglutinin gene of this subtype. The detection limit of the RT-LAMP assay was a virus dilution of 10(-5); which was 10(3) times more sensitive than the Espline Influenza A&B-N test and 10 times more sensitive than a reverse transcription polymerase chain reaction (RT-PCR) assay. The specificity of the RT-LAMP assay was examined by usin...
Cordoni G, Williams A, Durham A, Florio D, Zanoni RG, La Ragione RM.Strangles is one of the most common equine infectious diseases with serious health, welfare and socio-economic impact. However, the detection of Streptococcus equi subspecies equi can be challenging and persistently infected carriers are common. Furthermore, the use of classical microbiology can result in an underestimation of the prevalence of the disease. The difficulties associated with the slow diagnosis of Strangles can result in rapid spread of the disease. Therefore, rapid and economical diagnostic tests are urgently required. Here, two multiplex assays, were developed and validated for...
Pitzer JB, Kaufman PE, Tenbroeck SH, Maruniak JE.A species-specific multiplex polymerase chain reaction targeting the cytochrome b gene of cattle, horses, humans, and dogs was developed to determine the blood meal sources of stable flies, Stomoxys calcitrans (L.), collected from Florida equine facilities. Of 595 presumptive blood-fed stable flies analyzed, successful host amplification was obtained in 350, for a field host-detection efficiency of 58.8%. The majority of analyzed stable flies had fed on cattle (64.6%), followed by horses (24.3%), humans (9.5%), and dogs (1.6%). A survey of animal-enclosed pastures occurring within 3 km of stab...
Tong P, Duan R, Palidan N, Deng H, Duan L, Ren M, Song X, Jia C, Tian S, Yang E, Kuang L, Xie J.EHV-1 is one of the most serious viral pathogens that frequently cause abortion in horses around the world. However, so far, relatively little information is available on EHV-1 infections as they occur in China. In January 2021, during an abortion storm which occurred in Yili horses at the Chinese State Studs of Zhaosu (North Xinjiang, China), 43 out of 800 pregnant mares aborted. Results: PCR detection revealed the presence of EHV-1 in all samples as the possible cause of all abortions, although EHV-4, EHV-2 and EHV-5 were also found to circulate in the aborted fetuses. Furthermore, the parti...
Nemoto M, Morita Y, Niwa H, Bannai H, Tsujimura K, Yamanaka T, Kondo T.A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the rapid detection of equine coronavirus (ECoV). This assay was conducted at 60 °C for 40 min. Specificity of the RT-LAMP assay was confirmed using several equine intestinal and respiratory pathogens in addition to ECoV. The novel assay failed to cross-react with the other pathogens tested, suggesting it is highly specific for ECoV. Using artificially synthesized ECoV RNA, the 50% detection limit of the RT-LAMP assay was 10(1.8)copies/reaction. This is a 50-fold greater sensitivity than conventio...
Hamond C, Pestana CP, Rocha-de-Souza CM, Cunha LE, Brandão FZ, Medeiros MA, Lilenbaum W.Leptospirosis is a zoonotic disease of global importance, and has a worldwide distribution. Equine leptospirosis is commonly manifested by recurrent uveitis, reproductive disorders, as abortions, embryonic absorption, stillbirth and the birth of weak foals. The aim of this study was to verify the presence of Leptospira sp or its DNA in genital tract of mares with reproductive problems. A total of 38 mares with reproductive problems were studied. All the mares were sampled for blood (for serology), urine (for culturing and qPCR), vaginal fluid-VF and endometrial biopsy-EB (for culturing, qPCR a...
Borchers K, Wolfinger U, Goltz M, Broll H, Ludwig H.Equine herpesvirus type 2 (EHV-2) is a slow-growing, cytopathogenic gammaherpesvirus, which is suggested to be ubiquitous in the equine population. However, its precise role as a pathogen and its tissue tropism remains uncertain. To estimate the prevalence of EHV-2 in Germany and to investigate the possible pathogenicity of the virus, peripheral blood leucocytes (PBL) from 172 horses were examined for EHV-2 DNA by a sensitive and specific nested PCR based on the EcoRI-N genomic fragment and by classical cocultivation. PBL samples from 51% of the horses were positive by PCR and virus was isolat...
Hodgkinson JE, Freeman KL, Lichtenfels JR, Palfreman S, Love S, Matthews JB.The efficacy of five daily fenbendazole (FBZ) treatments was tested against benzimidazole-resistant cyathostomins in naturally infected horses (n=13). Horses were treated with pyrantel embonate (PYR) to remove adult strongyles followed, 7 days later, by a 5-day course of FBZ. The PYR treatment produced an average faecal egg count reduction of 98%. All samples were negative by faecal egg count 7 days after the start of the FBZ treatment. Positive egg counts were observed from 28 days after the start of FBZ treatment and all horses displayed positive faecal egg counts by 77 days after treatment....
