Report of the equine herpesvirus-1 Havermeyer Workshop, San Gimignano, Tuscany, June 2004.

Amongst the infectious diseases that threaten equine health, herpesviral infections remain a world wide cause of serious morbidity and mortality. Equine herpesvirus-1 infection is the most important pathogen, causing an array of disorders including epidemic respiratory disease abortion, neonatal foal death, myeloencephalopathy and chorioretinopathy. Despite intense scientific investigation, extensive use of vaccination, and established codes of practice for control of disease outbreaks, infection and disease remain common.

Time of transforming growth factor beta 1 inoculation alters the incubation of BSE in mice.

Groups of 20 C57BL/6J mice (10 males and 10 females) were given BSE strain 301C i.p. and subsequently given 2 microg recombinant human TGFbeta1 s.

Impaired motor coordination on static rods in BSE-infected mice.

Scrapie and bovine spongiform encephalopathy (BSE) are both progressive neurodegenerative diseases that are transmissible to mice. The onset of clinical symptoms is more subtle and variable in murine BSE than in murine scrapie. Assessment of behavioural changes that occur throughout disease would aid early diagnosis of disease so that more consistent end points could be made and potential therapies could be investigated.

Clusterin shortens the incubation and alters the histopathology of bovine spongiform encephalopathy in mice.

Clusterin accumulates in significant quantity in prion protein lesions associated with bovine spongiform encephalopathy (BSE) and we therefore sought to elucidate its ability to alter BSE pathogenesis and incubation time by comparison of wild type C57BL/6J mice and clusterin knock out (ko) mice. The ko mice had a 40 day increase in mean incubation time compared to wild type mice. PrP deposition in the medulla was less aggregated in clusterin knock out mice when compared to wild type BSE infected mice and a more marked astrocytosis, as determined by GFAP staining, was evident.

Equid herpesvirus 1 infection of endothelial cells requires activation of putative adhesion molecules: an in vitro model.

Antisera to activated equine endothelial cells, which detected surface molecules of 116 kD, 97 kD, 42 kD and 38 kD, were made to investigate the role of endothelial adhesion molecules in equid herpes virus 1 infection. These putative adhesion molecules could be induced by 17-beta oestradiol, chorionic gonadotrophin, or IL-2, as well as by LPS and PWM. In an in vitro flow system, using equine veins or arteries, equid herpesvirus 1 in leucocytes was only transferred to infect endothelial cells if both leucocytes and endothelial cells expressed these surface molecules.

Equid herpesvirus 1 is neurotropic in mice, but latency from which infectious virus can be reactivated does not occur.

Equid herpesvirus 1 (EHV-1) is the most common cause of virus-induced abortion in horses. After primary infection the virus becomes latent predominantly in the respiratory tract lymph nodes and the genome can also be detected in the peripheral nervous system. The role of mouse as a feasible model for the establishment of latency and reactivation of EHV-1 was investigated.

EHV-1 gene63 is not essential for in vivo replication in horses and mice, nor does it affect reactivation in the horse: short communication.

The aim of this study was to investigate the role of immediate early gene (gene63) in the pathogenesis of equine herpesvirus 1 (EHV-1) acute and latent infections in equine and murine models. EHV-1 gene63 mutant virus (g63mut) along with EHV-1 (Ab4) was used for intracerebral and intranasal infection of 3 and 17-day-old mice. Both viruses were recovered at the same frequency from tissues after infection.

Infection of endothelial cells with equine herpesvirus-1 (EHV-1) occurs where there is activation of putative adhesion molecules: a mechanism for transfer of virus.

Evidence is presented to show that activation of endothelial and leucoyte adhesion molecules is a key step in transferring virus from infected leucocytes; and determines the restricted tissue tropism. A range of tissues from 2 experimentally infected mares in late pregnancy at 4 and 8 days after infection with EHV-1 were compared with those from normal pregnant and nonpregnant mares. Rabbit antisera to equine activated endothelial cell molecules were used to identify which tissues expressed these molecules in normal nongravid and gravid mares, and to investigate whether the range of tissues was altered in pregnant mares as a consequence of infection.

Equid herpesvirus 1: platelets and alveolar macrophages are potential sources of activated TGF-B1 in the horse.

Cell mediated responses to Equid herpesvirus 1 (EHV-1) are of short duration in vivo and require considerable expansion to be detected in vitro. Raised serum levels of active transforming growth factor B (TGF-B1) have been shown to depress proliferative T cell responses in experimental infections with EHV-1 in ponies. The present work indicates that latent transforming growth factor B (TGF-B1) is present in circulating platelets, lymph node, bronchial epithelium and alveolar macrophages.

Clusterin in bovine spongiform encephalopathy (BSE).

Clusterin mRNA, detected in increased quantities in the cervical spinal cord of cattle with bovine spongiform encephalopathy (BSE), was localized mainly in the neuroglia (including astrocytes) of the lateral and ventral areas of white matter. Axonal degeneration was also observed in these areas. The dorsal horns of the spinal cord in which BSE prion protein (PrP(BSE)) was deposited did not exhibit strong clusterin "up-regulation" but showed increased clusterin immunolabelling with a punctate distribution in the neuropil.

Clusterin prevents aggregation of neuropeptide 106-126 in vitro.

The prion/amyloid neuropeptide 106-126 spontaneously aggregates to form fibrillar structures in vitro. The aggregation in vitro could be prevented in a dose-related manner by clusterin, and the specificity of this action was confirmed by reversal with antibody to clusterin. The relevance of these observations is discussed in relation to previous observations that clusterin and PrPBSE colocalise in naturally occurring cases of BSE.

Equine herpesvirus type 1 infects dendritic cells in vitro: stimulation of T lymphocyte proliferation and cytotoxicity by infected dendritic cells.

Equine herpesvirus type 1 (EHV-1) causes respiratory disease, abortion and myeloencephalopathy in horses. As with other herpesviruses, cell-mediated immunity is considered important for both recovery and protection. Although virus-specific T-cell proliferation and cytotoxicity can be detected following in vivo infection, little is known about the role of antigen presenting cells such as dendritic cells (DCs) in these processes.

In vitro reactivation of latent equid herpesvirus-1 from CD5+/CD8+ leukocytes indirectly by IL-2 or chorionic gonadotrophin.

IL-2 and equine chorionic gonadotrophin (eCG) initiated reactivation of equid herpesvirus-1 (EHV-1) from venous lymphocytes at a frequency of 1/10(-5). Indirect immunofluorescence showed that > 80% of virus-positive leukocytes were CD5+/CD8+ with the remaining 20% being CD5+/CD8-/CD4-. Cocultivation demonstrated that the reactivated virus was infectious.

The equid herpesvirus-1 gene 63 is expressed as a leaky late (gamma 1) transcript and is nonessential for replication in vitro.

The regulation of Equid herpesvirus-1 (EHV-1) gene 63 was investigated using molecular expression studies and its role in viral growth was identified by constructing a gene 63 mutant virus. Metabolic inhibitors were used to show that EHV-1 gene 63 is expressed as a leaky-late (gamma 1) transcript. Transient transfections and subsequent chloramphenicol acetyltransferase (CAT) reporter assays showed that gene 63 was transactivated by EHV-1 gene 64 (immediate early) protein.

Isolation and characterisation of equine dendritic cells.

Despite their important role in initiating T-cell responses in other species, dendritic cells have not been studied in the horse. A method for isolating blood dendritic cells by adherence and metrizamide gradients was adapted to equine cells. A number of monoclonal antibodies (mAbs), including some which label dendritic cells in other species, were tested for immunochemical reactivity with the isolated blood dendritic cells, and sections of lymph node and spleen.

Detection of latency-associated transcripts of equid herpesvirus 1 in equine leukocytes but not in trigeminal ganglia.

Results from Southern hybridization and PCR amplification experiments using a randomly synthesized reverse transcription-PCR product showed that peripheral blood leukocytes from horses showing no clinical signs of disease expressed a putative latency-associated transcript antisense to and overlapping the 3' end of the equid herpesvirus 1 (EHV-1) immediate-early gene (gene 64). A PCR product derived from this transcript has > or =96% identity with the published EHV-1 sequence. In situ hybridization studies of equine bronchial lymph nodes corroborated these findings and are consistent with reactivation data (D.

Transforming growth factor-beta induced by live or ultraviolet-inactivated equid herpes virus type-1 mediates immunosuppression in the horse.

Up to 21 days after exposure to live or ultraviolet-inactivated equid herpesvirus type-1 (EHV-1) autologous serum from ponies caused an immunosuppressive effect if incorporated into T-cell proliferation assays to EHV-1. The suppressive factor in the sera of ponies also inhibited T-cell response to phytohaemagglutinin. Increased levels of circulating activated transforming growth factor-beta 1 (TGF-beta 1) were detected, and the suppressive activity of the serum could be reversed by antibody to TGF-beta 1.

Nucleotide sequence of equine MxA cDNA.

A 2.6 kb cDNA species has been isolated from a cDNA library prepared from interferon-alpha stimulated equine peripheral blood leucocytes and the nucleotide sequence determined. The cDNA has a single open reading frame potentially encoding a 660 amino acid polypeptide showing a high degree of homology with known mammalian Mx proteins, including the possession of three consensus GTP-binding motifs.

Report of the First International Workshop on Equine Leucocyte Antigens, Cambridge, UK, July 1991.

The First International Workshop on Equine Leucocyte Antigens was organized and convened for the purposes of identifying immunologically relevant cell surface molecules of equine leucocytes and establishing a system of nomenclature for those molecules. Participating members of the workshop represented the majority of laboratories world-wide engaged in the tasks of production and characterization of equine leucocyte and lymphocyte markers using monoclonal antibodies. The workshop confirmed the identification of several equine CD molecules described previously by individual laboratories, and in addition recognized antibodies identifying new CD molecules.

The prevalence of latent Equid herpesviruses in the tissues of 40 abattoir horses.

Equid herpesviruses 1 or 4 (EHV-1 or -4) were isolated by cocultivation from 60% of 40 horses examined at slaughter. The lymph nodes draining the respiratory tract were the most common source of virus. EHV-1 or EHV-4 was never isolated from the trigeminal ganglia (SLG).

Equid herpesviruses 1 and 4 encode functional homologs of the herpes simplex virus type 1 virion transactivator protein, VP16.

The herpes simplex virus type 1 (HSV-1) tegument protein VP16 is a potent transcriptional inducer of immediate-early gene expression, comprising an N-terminal domain involved in binding DNA linked to an acidic transactivating C-terminal domain. The gene encoding the counterpart of this protein in equid herpesvirus 4 (EHV-4) was sequenced. Comparisons with VP16 and the homologous proteins of equine herpesvirus 1 (EHV-1) and varicella-zoster virus (VZV) showed that a region in the N-terminal domain involved in formation of a complex with cellular proteins is partially conserved in all four proteins.

The effect of EHV-1 infection upon circulating leucocyte populations in the natural equine host.

It has been suggested that EHV-1 infection may perturb immune responsiveness in the natural equine host. The mechanism underlying this phenomenon is not clear, but disturbances of circulating leucocyte populations could contribute. In order to objectively assess the nature of the haematological changes provoked by EHV-1 infection, two groups of conventionally-maintained Welsh mountain ponies were challenge-infected intra-nasally with the Ab4 isolate of EHV-1.