Molecular prevalence of equine alphaherpesvirus-1 shedding in healthy broodmares in Ontario.

To localize the genes for the major glycoproteins of equine herpesvirus 1 (EHV-1), a library of the EHV-1 genome was constructed in the lambda gt11 expression vector. Recombinant bacteriophage expressing EHV-1 glycoprotein epitopes as fusion products with beta-galactosidase were detected by immunoscreening with monoclonal antibodies specific for each of six EHV-1 glycoproteins. Seventy-four recombinant lambda gt11 clones reactive with EHV-1 monoclonal antibodies were detected among 4 X 10(5) phage screened. Phage expressing determinants on each of the six EHV-1 glycoproteins were represented in the library. Herpesviral DNA sequences contained in lambda gt11 recombinants expressing epitopes of EHV-1 glycoproteins were used as hybridization probes for mapping insert sequences on the viral genome. Genes for five EHV-1 glycoproteins (gp2, gp10, gp13, gp14, and gp21/22a) mapped to the genome L component; only one EHV-1 glycoprotein (gp17/18) was expressed from the unique S region of the genome where genes of several major glycoproteins of other herpesviruses have been located. Two glycoproteins of EHV-1, gp13 and gp14, mapped to positions colinear with genes of major glycoproteins identified in several other alphaherpesviruses (gC- and gB-like glycoproteins, respectively). The genomic locations of other EHV-1 glycoproteins indicated the existence of major glycoproteins of EHV-1 (gp2, gp10, and gp21/22a) for which no genetic homologs have yet been detected in other herpesviruses. The results confirm the general utility of the lambda gt11 expression system for localizing herpesvirus genes and suggest that the genomic positioning of several high-abundance glycoproteins of EHV-1 may be different from that of the prototype alphaherpesvirus, herpes simplex virus.

Insertional mutagenesis was used to construct an equine herpesvirus 1 (EHV-1) mutant in which the open reading frame for glycoprotein D was replaced by a lacZ cassette. This gD deletion mutant (delta gD EHV-1) was unable to infect normally permissive RK cells in culture, but could be propagated in an EHV-1 gD-expressing cell line (RK/gD). Phenotypically complemented delta gD EHV-1 was able to infect RK cells, but did not spread to form syncytial plaques as seen with wild type EHV-1 or with delta gD EHV-1 infection of RK/gD cell cultures. Therefore EHV-1 gD is required for virus entry and for cell-cell fusion. The phenotypically complemented delta gD EHV-1 had very low pathogenicity in a mouse model of EHV-1 respiratory disease, compared to a fully replication-competent EHV-1 reporter virus (lacZ62/63 EHV-1). Intranasal or intramuscular inoculation of mice with delta gD EHV-1 induced protective immune responses that were similar to those elicited in mice inoculated with lacZ62/63 EHV-1 and greater than those following inoculation with UV-inactivated virus.

Expression of small regions of equine herpesvirus 1 glycoprotein C in Escherichia coli

Equine herpesvirus-1 infected peripheral blood mononuclear cell subpopulations during viremia

This study reports the first equine herpesvirus-1 (EHV-1) and equine herpesvirus-4 (EHV-4) seroprevalence investigation in horse populations of Morocco in 24 years. It also aims to determine antibody titers in horses vaccinated under field conditions with a monovalent EHV-1 vaccine. Blood samples were collected from 405 horses, including 163 unvaccinated and 242 vaccinated animals. They were tested using a commercial type-specific enzyme-linked immunosorbent assay (ELISA) and a virus neutralization test (VNT). Overall, 12.8% unvaccinated, and 21.8% vaccinated horses were positive for EHV-1. All samples were positive for EHV-4 when tested with the type-specific ELISA. In the vaccinated group, the VNT revealed a mean antibody titer of 1:49 for EHV-1 and 1:45 for EHV-4. The present study demonstrates that EHV-1 and EHV-4 are endemic in the horse populations in the north of Morocco, with prevalence differences between regions. Furthermore, horses vaccinated with a monovalent EHV-1 vaccine had low antibodies titers. This study highlights the necessity to establish and/or support efficient biosecurity strategies based on sound management of horses and characterize further and potentially improve the efficiency of the EHV vaccines and vaccination protocol used in the field.

Equid herpesviruses (EHVs) threaten equine health and can cause significant economic losses to the equine industry worldwide. Different equid herpesviruses, EHV‐1, EHV‐2, EHV‐4 and EHV5 are regularly detected among horse populations. In Egypt, monitoring is sporadic but EHV‐1 or EHV‐4 have been reported to circulate in the horse population. However, there is a lack of reports related to infection and health status of horses, likely due to the absence of regular diagnostic procedures. In the current study, the circulation of four infectious equid herpesviruses (EHV‐1, EHV‐2, EHV‐4 and EHV‐5) among different Arabian horse populations and donkeys residing the same farm was monitored. Different samples were collected and DNA was extracted and subjected to quantitative (q)‐PCR to detect the four equid herpesviruses using specific primers and probes. Antibody titres against EHV‐1 and EHV‐4 were tested using virus neutralization test and type‐specific ELISA. The results showed that EHV‐1, EHV‐2, EHV‐4 and EHV‐5 are endemic and can be a continuous threat for horses in the absence of vaccination programs and frequent virus reactivation. There is an urgent need for introduction of active regular surveillance measures to investigate the presence of different equid herpesviruses, and other equine viral pathogens, in various horse populations around Egypt and to establish a standardized cataloguing of equine health status.

The development of a competitive PCR–ELISA for the detection of equine herpesvirus-1

Protective effects of equine herpesvirus-1 (EHV-1) glycoprotein B in a murine model of EHV-1-induced abortion

Background. Equine herpesvirus type 1 (EHV-1) is responsible for respiratory disease in young animals, abortion in pregnant mares and neurological disease, whilst equine herpesvirus 4 (EHV-4) is mainly the causative agent of respiratory disorders and rarely causes abortion. These viruses are considered as one of the most clinically and economically important pathogens of horses and can be detected in a range of tissues. Scope and Approach. Serological methods are used to detect the presence and titre of specific antibodies to EHV-1 and EHV-4 in the sera of examined horses and are useful in epizootiological studies. Commercially available ELISA kits are able to differentiate specific EHV-1 and EHV-4 antibodies. EHV-1 and EHV-4 can both be isolated using susceptible cells such as primary horse cell cultures and other non-equine cells with visible cytopathic effect. Since standard diagnostic methods can be time consuming and arduous, the scope of many studies has been to develop and confirm the sensitivity and specificity of molecular diagnostic methods. Key Findings and Conclusions. Polymerase chain reaction (PCR) has proved to be a good screening method for the presence of latent infections of horses caused by these viruses, also making possible the rapid identification and differentiation of EHV-1 and- EHV-4 in the examined samples. Real-time PCR is a sensitive, specific and quantitative method that enables the determination of viral kinetics in infected horses. Genome sequencing can be used to discover mutations in the genomes of EHV-1 and EHV-4 as well as to track the spread of their different strains globally.

Epidemiology of Chlamydia psittaci infections in pregnant Thoroughbred mares and foals

Pre-infection frequencies of equine herpesvirus-1 specific, cytotoxic T lymphocytes correlate with protection against abortion following experimental infection of pregnant mares

Frequency and phenotype of EHV-1 specific, IFN- γ synthesising lymphocytes in ponies: The effects of age, pregnancy and infection

Pre-infection frequencies of equine herpesvirus-1 specific, cytotoxic T lymphocytes correlate with protection against abortion following experimental infection of pregnant mares

The Egyptian Arabian horse is one of the oldest and most popular horse breeds in the world. No previous efforts have been made for investigating the existence and prevalence of equine herpes viruses (EHVs) in this precious horse breed. In this report, ninety three clinical samples were collected from a cohort of Arabian horses located in Cairo, Egypt. Screening of the clinical samples for the presence of EHV antigens by cell-ELISA utilizing a polyclonal antibody pool against EHV-1, 2, and 4 identified 34 (36.56%) positive samples. Virus-specific semi-nested PCR assays were used for typing the positive samples. Three samples were found positive for EHV-1, seventeen for EHV-2, seven for EHV-4, one was a mixed infection of EHV-1 and EHV-4, and six did not produce any amplification signal with all assays. Sequence analysis of the amplified semi-nested PCR products of representative EHV strains further confirmed the virus identity. This study is the first that outlines the involvement of EHV infections in Arabian horse diseases worldwide. Besides, it presents new data about the prevalence of EHVs in the Egyptian horse population. Key words: Arabian horses, Egypt, equine herpes viruses, prevalence.

Equine herpesvirus 1 (EHV-1) causes considerable economic loss to the equine industry and is spread among susceptible animals during the cycles of latency and reactivation, causing rhinopneumonitis, abortion, and neurological disease. Nucleotide polymorphisms within ORF30 and ORF68 sequences of the viral genome are associated with strain neuropathogenicity and geographical origin. A total of 142 tissue and nasal swab samples from apparently healthy unvaccinated horses were examined to ascertain EHV-1 distribution, diversity, and clinical significance considering the results of virus isolation, sequence analysis, and anamnestic data. The ORF30 and ORF68 molecular study of these circulating strains and archival isolates from abortion storms aimed to contribute to the perception of strain pathogenicity and origin. EHV-1 was detected by PCR and virus isolation in 81 and 45.1% of the analyzed samples, respectively, and 82.1% of the representative samples were neuropathogenic strains. The ORF68-based grouping was restricted by the pronounced polymorphism of Balkan EHV-1 strains, and only two isolates were assigned to group 4. The cases of abortion were caused by neuropathogenic strains that also circulate within the horse population with no documented outbreaks of disease. It was evident that strain virulence is not solely accountable for the development of clinical symptoms in affected animals. Neural tissue is significant for virus latency and reactivation, considering the number of EHV-1 isolates from apparently healthy stressed horses. Special care must be taken when accommodating together immunologically naive and latently infected horses since asymptomatic carriers silently shed EHV-1.