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Topic:Equine Viral Arteritis
Equine Viral Arteritis (EVA) is a contagious viral disease affecting horses, caused by the equine arteritis virus (EAV). The virus primarily spreads through respiratory secretions and venereal transmission, impacting both the respiratory and reproductive systems of horses. Clinical signs of EVA can vary widely, from subclinical infections to more severe symptoms such as fever, nasal discharge, conjunctivitis, and swelling of limbs and genitalia. In pregnant mares, the virus can lead to abortion. EVA can be diagnosed through serological tests, virus isolation, and molecular techniques such as PCR. This page compiles peer-reviewed research studies and scholarly articles that explore the epidemiology, pathogenesis, diagnosis, and control measures of Equine Viral Arteritis in equine populations.
Extended phylogeny of the equine arteritis virus sequences including South American sequences. To perform genetic analysis of the ORF5 of equine arteritis virus (EAV) may provide new insights into the genetic evolution and origin of the Argentinean EAV sequences. Methods: A total of 76 sequences were analyzed by neighbor joining (NJ), maximum parsimony and maximum likelihood algorithms. The analysis of the selective pressures was performed using the Tajima's test. Results: The trees showed similar topologies. Two clades were identified: the first clade was formed by strains isolated mainly from a donkey, whereas the second clade presented four large groups from different geographic regi...
Evaluation of neutralization patterns of the five unique Argentine equine arteritis virus field strains reported. Equine viral arteritis (EVA) is a contagious viral disease that frequently causes mild or subclinical infections in adult horses. Only one EAV serotype has been described. However, there are differences in antigenicity, pathogenicity and neutralization characteristics of virus field strains. The interaction of two viral proteins, GP5 and M, is critical for infectivity and amino acid changes in the GP5 sequences have an effect on the neutralizing phenotype, regardless the effects of other viral proteins. The objective of the present study was to evaluate the neutralization phenotypes of the 5 u...
Molecular epidemiology and genetic characterization of equine arteritis virus isolates associated with the 2006-2007 multi-state disease occurrence in the USA. In 2006-2007, equine viral arteritis (EVA) was confirmed for the first time in Quarter Horses in multiple states in the USA. The entire genome of an equine arteritis virus (EAV) isolate from the index premises in New Mexico was 12 731 nt in length and possessed a previously unrecorded unique 15 nt insertion in the nsp2-coding region in ORF1a and a 12 nt insertion in ORF3. Sequence analysis of additional isolates made during this disease occurrence revealed that all isolates from New Mexico, Utah, Kansas, Oklahoma and Idaho had 98.6-100.0 % (nsp2) and 97.8-100 % (ORF3) nucleotide identity and c...
Complex interactions between the major and minor envelope proteins of equine arteritis virus determine its tropism for equine CD3+ T lymphocytes and CD14+ monocytes. Extensive cell culture passage of the virulent Bucyrus (VB) strain of equine arteritis virus (EAV) to produce the modified live virus (MLV) vaccine strain has altered its tropism for equine CD3(+) T lymphocytes and CD14(+) monocytes. The VB strain primarily infects CD14(+) monocytes and a small subpopulation of CD3(+) T lymphocytes (predominantly CD4(+) T lymphocytes), as determined by dual-color flow cytometry. In contrast, the MLV vaccine strain has a significantly reduced ability to infect CD14(+) monocytes and has lost its capability to infect CD3(+) T lymphocytes. Using a panel of five re...
Curing of HeLa cells persistently infected with equine arteritis virus by a peptide-conjugated morpholino oligomer. 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...
Antiviral effect of recombinant equine interferon-gamma on several equine viruses. Recombinant equine interferon-gamma (reIFN-gamma) was prepared using a baculovirus expression system and its antiviral activity was investigated using several equine viruses. The reIFN-gamma suppressed the replication of all equine viruses used in the present experiment in horse cell cultures, but did not affect the growth of host cells at concentrations of less than 1000 u/ml. A strong antiviral effect was observed, especially against RNA viruses. Equine picornavirus, equine rhinovirus and equine arteritis virus could not be propagated at all in 100 u/ml reIFN-gamma when 100 TCID(50) of infec...
RNA interference protects horse cells in vitro from infection with Equine Arteritis Virus. Equine Arteritis Virus (EAV) belongs to the Arteriviridae and causes viral arteritis in horses. In an attempt to develop novel and save therapies against the infection it was tested whether EAV is susceptible to RNA interference (RNAi) in an equine in vitro system. Horse cells were transfected with chemically synthesized small interfering RNA oligonucleotides (siRNAs) and challenged with EAV. Application of these siRNAs led to a significant protection of the cells, and virus titers decreased drastically. siRNAs derived from DNA plasmids expressing small hairpin RNAs (shRNAs) were also effectiv...
Equine arteritis virus: a new isolate from the presumable first carrier stallion in Argentina and its genetic relationships among the four reported unique Argentinean strains. Equine arteritis virus (EAV) was isolated from a testicle of the presumable first stallion infected with EAV in Argentina. This virus isolate (named LT-LP-ARG) was confirmed by GP5-specific PCR and indirect immunofluorescence assays. The PCR product was sequenced, and the phylogenetic analysis revealed that the LT-LP-ARG strain of EAV forms a monophyletic group, together with other strains previously isolated in our laboratory (LP02 group). However, all Argentinean EAV strains belong to a polyphyletic group. We believe that the virus isolate presented in this report could be the origin of EAV ...
Amino acid substitutions in the structural or nonstructural proteins of a vaccine strain of equine arteritis virus are associated with its attenuation. Comparative sequence analysis of a series of strains of equine arteritis virus (EAV) of defined virulence for horses, ranging from the horse-adapted virulent Bucyrus (VB) strain to a fully attenuated vaccine strain derived from it, identified 13 amino acid substitutions associated with attenuation. These include 4 substitutions in the replicase proteins and 9 in the structural proteins. Using reverse genetic techniques, these amino acid substitutions were introduced into a virulent infectious cDNA clone pEAVrVBS derived from the VB strain of EAV. Inoculation of horses with the recombinant viru...
Equine arteritis virus is delivered to an acidic compartment of host cells via clathrin-dependent endocytosis. Equine arteritis virus (EAV) is an enveloped, positive-stranded RNA virus belonging to the family Arteriviridae. Infection by EAV requires the release of the viral genome by fusion with the respective target membrane of the host cell. We have investigated the entry pathway of EAV into Baby Hamster Kidney cells (BHK). Infection of cells assessed by the plaque reduction assay was strongly inhibited by substances which interfere with clathrin-dependent endocytosis and by lysosomotropic compounds. Furthermore, infection of BHK cells was suppressed when clathrin-dependent endocytosis was inhibited ...
Equine viral arteritis: current status and prevention. Recently, there has been increased interest in equine viral arteritis (EVA) among veterinarians and horse owners. Outbreaks of the disease were identified initially in New Mexico, USA in 2006, and in the Normandy region of France in the summer of 2007. Both occurrences were associated with AI of cool-shipped semen. Each was linked to respiratory illness, neonatal death, abortion, development of carrier stallions, and cancellation of equestrian events. In light of the increased interest, this paper will present a brief case history, followed by a review addressing common concerns regarding EVA,...
Comparison of two real-time reverse transcription polymerase chain reaction assays for the detection of Equine arteritis virus nucleic acid in equine semen and tissue culture fluid. Two previously developed TaqMan fluorogenic probe-based 1-tube real-time reverse transcription polymerase chain reaction (real-time RT-PCR) assays (T1 and T2) were compared and validated for the detection of Equine arteritis virus (EAV) nucleic acid in equine semen and tissue culture fluid (TCF). The specificity and sensitivity of these 2 molecular-based assays were compared to traditional virus isolation (VI) in cell culture. The T1 real-time RT-PCR had a higher sensitivity (93.4%) than the T2 real-time RT-PCR (42.6%) for detection of EAV RNA in semen. However, the T1 real-time RT-PCR was les...
Formation of the arterivirus replication/transcription complex: a key role for nonstructural protein 3 in the remodeling of intracellular membranes. The replication/transcription complex of the arterivirus equine arteritis virus (EAV) is associated with paired membranes and/or double-membrane vesicles (DMVs) that are thought to originate from the endoplasmic reticulum. Previously, coexpression of two putative transmembrane nonstructural proteins (nsp2 and nsp3) was found to suffice to induce these remarkable membrane structures, which are typical of arterivirus infection. Here, site-directed mutagenesis was used to investigate the role of nsp3 in more detail. Liberation of the hydrophobic N terminus of nsp3, which is normally achieved by c...
The efficacy of a commercial ELISA as an alternative to virus neutralisation test for the detection of antibodies to EAV. Infection with equine arteritis virus is a notifiable disease with sporadic occurrence in the UK. As stallions may harbour the virus after infection, horses are screened for exposure by serological testing prior to breeding. The virus neutralisation test is considered the 'gold standard' serological screening test, but it is time-consuming and labour intensive; consequently there is a move towards more rapid screening methodology. In this study, a commercially available EVA antibody ELISA is assessed. The ELISA performed poorly with a specificity [corrected] of 26% and a sensitivity [corrected...
Characterization of equine arteritis virus particles and demonstration of their hemolytic activity. Equine arteritis virus (EAV), a member of the newly established family Arteriviridae, is a small, positive-stranded RNA virus. It carries two protein complexes in its envelope, gp5/M and the recently described gp2b/gp3/gp4 complex. We report here on several basic features of EAV replication in cell culture and on the protein composition of virus particles. We have also characterized gp2b, gp3, and gp4 expressed using a baculovirus system in insect cells. Finally, we provide evidence that EAV possess hemagglutinating and hemolytic activity. The hemolysis assay might be useful for determining wh...
Analysis of ORFs 2b, 3, 4, and partial ORF5 of sequential isolates of equine arteritis virus shows genetic variation following experimental infection of horses. Samples from horses experimentally infected with the "large plaque variant (LP3A+)" of equine arteritis virus were analysed. These included 182 nasal swabs collected from day 1 to 14 post-infection (p.i.), and 21 virus isolates obtained from white blood cells of animals that showed a prolonged viraemia between days 30 to 72 p.i. In order to determine the genetic stability of the virus and particularly to characterise the genetic variants found during the prolonged viraemia, partial sequences of open reading frame 5 (ORF5) encoding glycoprotein 5 (GP5) were generated. Viruses with amino acid su...
Development of a fluorescent-microsphere immunoassay for detection of antibodies specific to equine arteritis virus and comparison with the virus neutralization test. The development and validation of a microsphere immunoassay (MIA) to detect equine antibodies to the major structural proteins of equine arteritis virus (EAV) are described. The assay development process was based on the cloning and expression of genes for full-length individual major structural proteins (GP5 amino acids 1 to 255 [GP5(1-255)], M(1-162), and N(1-110)), as well as partial sequences of these structural proteins (GP5(1-116), GP5(75-112), GP5(55-98), M(88-162), and N(1-69)) that constituted putative antigenic regions. Purified recombinant viral proteins expressed in Escherichia col...
Comparison of different molecular methods for assessment of equine arteritis virus (EAV) infection: a novel one-step MGB real-time RT-PCR assay, PCR-ELISA and classical RT-PCR for detection of highly diverse sequences of Slovenian EAV variants. In the present study, a new one-step real-time reverse transcription-polymerase chain reaction (RT-PCR) strategy with minor-groove-binder (MGB) technology for the detection of EAV from 40 semen samples of Slovenian carrier stallions was tested. A novel MGB probe (EAVMGBpr) and a reverse primer (EAV-R) based on the multiple sequence alignment of 49 different EAV strain sequences of the highly conserved ORF7 (nucleocapsid gene) were designed. The performance of the assay was compared with different molecular detection methods. Three different primer pairs targeting the ORF1b and ORF7 were used, ...
Genetic variation and phylogenetic analysis of 22 French isolates of equine arteritis virus. Genetic variation and phylogenetic relationships among 22 French isolates of equine arteritis virus (EAV) obtained over four breeding seasons (2001-2004) were determined by sequencing open reading frames (ORFs) 2a-7. The ORFs 2a-7 of 22 isolates differed from the prototype virulent Bucyrus strain of EAV by between 14 (99.5% identity) and 328 (88.7% identity) nucleotides, and differed from each other by between 0 (100% identity) and 346 (88.1% identity) nucleotides, confirming genetic diversity among EAV strains circulating in France. Phylogenetic analysis based on the partial ORF5 sequences (n...
Development and characterization of an infectious cDNA clone of the virulent Bucyrus strain of Equine arteritis virus. Strains of Equine arteritis virus (EAV) differ in the severity of the disease that they induce in horses. Infectious cDNA clones are potentially useful for identification of genetic determinants of EAV virulence; to date, two clones have been derived from a cell culture-adapted variant of the original (Bucyrus) isolate of EAV, and it has previously been shown that recombinant virus derived from one of these (rEAV030) is attenuated in horses. A complete cDNA copy of the genome of the virulent Bucyrus strain of EAV has now been assembled into a plasmid vector. In contrast to rEAV030, recombinant...
Genetic typing of equine arteritis virus isolates from Argentina. We report the nucleotide sequence and genetic diversity of four Equine Arteritis Virus (EAV) ORF 5 and 6 from Argentina isolates, obtained from asymptomatic virus-shedding stallions. Nucleic acid recovered from the isolates were amplified by RT-PCR and sequenced. Nucleotide and deduced amino acid sequences from the Argentine isolates were compared with 17 sequences available from the GenBank. Phylogenetic analysis revealed that the Argentine isolates grouped together in a definite cluster near European strains. Despite the greater genetic variability among ORF 5 from different isolates and str...
Experiences with new generation vaccines against equine viral arteritis, West Nile disease and African horse sickness. Viral diseases constitute an ever growing threat to the horse industry worldwide because of the rapid movement of large numbers of horses for competition and breeding. A number of different types of vaccines are available for protective immunization of horses against viral diseases. Traditional inactivated and live-attenuated (modified live virus, MLV) virus vaccines remain popular and efficacious but recombinant vaccines are increasingly being developed and used, in part because of the perceived deficiencies of some existing products. New generation vaccines include MLVs with deletions and/or...
Proteolytic maturation of replicase polyprotein pp1a by the nsp4 main proteinase is essential for equine arteritis virus replication and includes internal cleavage of nsp7. The positive-stranded RNA genome of the arterivirus Equine arteritis virus (order Nidovirales) encodes the partially overlapping replicase polyproteins pp1a (1727 aa) and pp1ab (3175 aa). Previously, three viral proteinases were reported to cleave these large polyproteins into 12 non-structural proteins (nsps). The chymotrypsin-like viral main proteinase residing in nsp4 is responsible for eight of these cleavages. Processing of the C-terminal half of pp1a (the nsp3-8 region) was postulated to occur following either of two alternative proteolytic pathways (the 'major' and 'minor' pathways). He...
[Prokaryotic expression of the major antigenic domain of equine arteritis virus GL protein and the establishment of putative indirect ELISA assay]. According to the antigenic analysis of equine arteritis virus (EAV) GL protein, one pair of primers were designed, with which the gene fragment coding the high antigenic domain of EAV GL protein was amplified from the EAV genome. The cloned gene was digested with BamH I and Xho I and then inserted into pET-32a and resulted pET-GL1. The pET-GL1 was transformed into the host cell BL21(DE3) and the expression was optimized including cultivation temperature and concentration of IPTG. The aim protein was highly expressed and the obtained recombinant protein manifested well reactiongenicity as was c...
Application of polymerase chain reaction and virus isolation techniques for the detection of viruses in aborted and newborn foals. The occurrence of two important pathogens, equine herpesvirus 1 (EHV1) and equine arteritis virus (EAV) causing abortions, perinatal foal mortality and respiratory disease, was investigated by polymerase chain reaction (PCR) and virus isolation to demonstrate the presence of abortigenic viruses in samples from 248 horse fetuses in Hungary. We found 26 EHV1- and 4 EAV-positive aborted or prematurely born foals from 16 and 4 outbreaks, respectively, proving that despite the widely applied vaccination, EHV1 is a far more important cause of abortions in the studs than EAV. We compared the virus co...
Disease transmission in horses. Bacterial, viral and protozoal infections may cause severe reproductive losses. The present paper reviews the risk factors, clinical signs and preventive measures for the most important venereal or potential sexually transmitted diseases in horses. The stallion and use of semen for artificial insemination represent major risk factors for the transmission of bacterial contaminants of the penis, including Streptococcus equi subspecies zooepidemicus, Pseudomonas aeruginosa and Klebsiella pneumoniae, known to cause endometritis and infertility in the mare. The role of the stallion in disease trans...
Effective removal of equine arteritis virus from stallion semen. A method of removing equine arteritis virus (EAV) from equine semen used for artificial insemination is urgently needed. Recent medical studies suggest that a double semen processing technique of density gradient centrifugation followed by a 'swim-up' can provide virus-free sperm preparations for assisted reproduction. Objective: To investigate the use of the double semen processing technique to obtain virus-free sperm preparations from stallion semen containing EAV. Methods: Aliquots of an ejaculate from an uninfected stallion were spiked with virus and processed by the double processing tech...
