On the rules of co-existence between sheep, cattle, and OvHV-2

Abstract

Malignant catarrhal fever (MCF) is a lethal disease of ruminants and swine, characterized by vasculitis, necrosis, and accumulation of activated, dysregulated cytotoxic lymphocytes in various tissues. It is transmitted from so-called “reservoir” animal species to “indicator” animal species. Ovine gammaherpesvirus 2 (OvHV-2) is one of several causative agents of MCF, circulating among all sheep breeds worldwide, which represent the “reservoir” of this virus. Natural infection of sheep proceeds without clinical symptoms. The disease is characterized by proliferation of cytotoxic T-cells, which form perivascular infiltrates of lymphoid as well as non-lymphoid organs, leading to necrosis and tissue destruction. However, little is known about the immunopathogenic pathways leading to disease. Moreover, at the time when this work was initiated, only limited data were available for the situation in sheep. Therefore, we first analyzed the cell populations targeted by the virus during natural infection of lambs, since the range of targeted cells may be important for the pathogenesis, especially if it is different in sheep and indicator animals, respectively. In most sheep we found a first peak of viral DNA in CD4+ T-helpercells, followed by a second peak in CD8+ cytotoxic T-cells. Both, CD4+ as well as CD8+ cells are also found to harbor viral DNA in lymphocytes of cattle with MCF. From this finding we suggest that rather a species-specific fine-tuning of viral gene expression than the types of targeted cells may be accountable for the disease. The second question addressed in this work was whether production of viral particles leading to cell lysis is among the pathological factors of MCF. Diseases due to gammaherpesvirus infections are mostly associated to latent infection, during which only a limited number of viral proteins are expressed. These are important to tie the viral DNA to host chromosomes, ensuring synchronous replication of viral DNA with cellular DNA, to provide viral DNA to each daughter cell and likely render the host cell increased resistance against apoptosis. However, persistent infection with Human herpesvirus 8 (HHV-8) as an exception is depending on a mixture of latently and lytically infected cells. To address whether this may be the case during MCF as well, we produced antisera against structural viral proteins and tested several tissue samples of experimentally infected rabbits therewith. We detected viral proteins in epithelial and M-cells of the appendix of infected animals. Interestingly, in situ hybridization revealed viral RNA in the infected epithelial cells but not in M-cells. These data suggest that active OvHV-2 replication may play a role in the pathogenesis of the disease. Thirdly, we tested the gene expression patterns of OvHV-2 and the relative abundances of host cell transcripts in lymphocytes of diseased cattle to identify pathways possibly involved in the pathogenesis of MCF. To this, host and virus gene expression patterns were analyzed by microarrray. Only two regions of the viral genome were found to be transcriptionally active, one encoding a latency-associated nuclear antigen, which can be found during latency of other gammaherpesviruses as well. The other with no predicted open reading frame, which may represent micro RNA (miRNA). So far miRNA was not discussed as a pathological factor of MCF and opens completely new attempts. As could be expected, a large number of host genes related to inflammation, lymphocyte activation, cell proliferation and apoptosis were found to be at different abundances compared to healthy animals. One of these transcripts with decreased expression was IL-2. Since the phenotype of mice with IL-2 deficiency perfectly matches the clinical signs of MCF, we assume that IL-2 deficiency may play an important role in the development of disease.