Identification of peripheral blood mononuclear cells targeted by Ovine herpesvirus-2 in sheep.

Sheep-associated malignant catarrhal fever (MCF), caused by Ovine herpesvirus 2 (OvHV-2), is usually a fatal disease of various ruminants and swine. In contrast, natural OvHV-2 infection in sheep, which are the main OvHV-2 reservoir, proceeds without any clinical symptoms. Since the range of targeted cells may be important for pathogenesis, we wanted to analyze the natural range of peripheral mononuclear blood cells (PBMC) targeted by OvHV-2. To this end, OvHV-2-free sheep were exposed to natural infection and blood samples were taken at intervals. Four different PBMC subpopulations were purified by fluorescence activated cell sorting (FACS) before being subjected to analysis for OvHV-2-DNA. After an incubation period of between 11 and 12 weeks, all exposed sheep became positive for OvHV-2. In most sheep, a first peak of OvHV-2-DNA was identified in the CD2 and CD4 double positive subpopulation. However, with time, the highest load of OvHV-2-DNA shifted to the CD2-positive and CD4-negative T-cells. Furthermore, low amounts of OvHV-2-DNA were occasionally detected also in the fractions that represented either CD14-positive monocytes or triple negative cells (CD2(-)/CD4(-)/CD14(-)). We conclude from these experiments that OvHV-2 has a similar host cellular range in sheep and cattle, respectively. Our results may be relevant in the context of comparative analysis of OvHV-2 pathogenesis in animal species that are susceptible to MCF.

Malignant catarrhal fever of cattle is associated with low abundance of IL-2 transcript and a predominantly latent profile of ovine herpesvirus 2 gene expression.

BACKGROUND: Malignant catarrhal fever (MCF) is a lethal disease of cattle, characterized by vasculitis, necrosis, and accumulation of activated, dysregulated cytotoxic lymphocytes in various tissues. Ovine gamma herpesvirus 2 (OvHV-2) is a causative agent of MCF, which may trigger the disease through immunopathogenic pathways. Lymphocytes are the main target of the virus. However, the pathogenic basis of the disease is still mysterious. METHODS/FINDINGS: We hypothesized that the gene expression patterns of OvHV-2 and the relative abundances of host cell transcripts in lymphnodes may be used to identify pathways that help to explain the pathogenesis of MCF. Therefore, viral and host cell gene expression patterns in lymph nodes of animals with MCF and healthy controls were analyzed by microarray. Two regions on the viral genome were transcriptionally active, one encoding an orthologue to the latency-associated nuclear antigen (ORF73) of other gamma herpesviruses, the other with no predicted open reading frame. A vast number of transcripts related to inflammatory processes, lymphocyte activation, cell proliferation and apoptosis were detected at different abundances. However, the IL-2 transcript was eminent among the transcripts, which were, compared to healthy controls, less abundant in animals with MCF. The ratio between CD4- and CD8-positive T-lymphocytes was decreased in the lymphnodes of animals with MCF compared to healthy controls. In contrast, the same ratio was stable, when peripheral blood lymphocytes were analyzed. CONCLUSIONS/SIGNIFICANCE: The phenotype of mice with a deficient IL-2-system almost perfectly matches the clinical signs observed in cattle with MCF, which feature a significantly decreased IL-2 transcript abundance, compared to healthy cattle. This supports the hypothesis that immunopathogenic events are linked to the pathogenesis of MCF. IL-2-deficiency may play an important role in the process. Therefore, this work opens new avenues for research on MCF.

Ovine herpesvirus 2 structural proteins in epithelial cells and M-cells of the appendix in rabbits with malignant catarrhal fever.

Sheep-associated malignant catarrhal fever (MCF), caused by Ovine herpesvirus 2 (OvHV-2), is a usually fatal disease of various ruminants and swine. A system for propagation of OvHV-2 in vitro has not yet been identified, although persistently infected cells have been derived from diseased animals and used to establish an animal model in rabbits. OvHV-2 structural proteins have not been detected in diseased animals and the pathogenesis of OvHV-2 infection is poorly understood. Recently, the genomic sequence of OvHV-2 has been determined, which allowed to predict the amino acid sequences of putative OvHV-2 structural proteins. Based on those predictions, we have generated antisera against two putative structural proteins (ORF43 and ORF63) of OvHV-2 in order to detect sites of active virus replication in experimentally OvHV-2-infected rabbits with signs of MCF. Although histological lesions typical of MCF were detected in multiple tissues, those sera detected viral capsid and tegument antigens exclusively in the appendix but not in other tissues of rabbits with MCF. More specifically, those viral proteins were detected in epithelial cells as well as in M-cells. However, in situ hybridization revealed that ORF63 mRNA was present in epithelial cells of infected rabbits but not in M-cells. Our data suggest that active OvHV-2 replication takes place in certain tissues of animals with MCF and that M-cells may play a role in the pathogenesis of MCF.