Distinct cellular and molecular responses to infection in three target cell types from horses, a species naturally susceptible to Ross River virus

Our current understanding of the pathogenesis of Ross River virus (RRV) infection has been derived from murine models, which do not reproduce clinical disease as experienced by infected humans and horses. This prompted us to establish more relevant host model systems to study host-virus interactions using ex vivo peripheral blood mononuclear cells (PBMCs) and in vitro primary synovial fibroblast and epidermal keratinocyte cultures. Transcriptomic analysis revealed that the expression of the transmembrane protein matrix remodelling associated 8 (mxra8), recently found to be responsible for RRV cell entry, was downregulated in all cell types when infected with RRV, compared to mock-infected controls. Potent antiviral and inflammatory responses were generated by both synovial fibroblasts and epidermal keratinocytes upon RRV infection. Upregulation of multiple genes, inducible by double-stranded RNA, together with upregulation of toll-like receptor (TLR) tlr-3, but not tlr-7, 8 and 9, suggests possible abortive replication of RRV in these cell types and potent antiviral mechanisms. This was corroborated by virus growth kinetic studies which indicated inefficient RRV replication in synovial fibroblasts and epidermal keratinocytes. Cellular metabolic flux studies on PBMCs and synovial fibroblasts showed that RRV infected cells had reduced mitochondrial function. In addition, compared to PBMCs of seronegative horses, an enhanced antiviral state and reduced inflammation related gene expression was seen in PBMCs of seropositive horses infected with RRV. Thus, despite potent antiviral and inflammatory responses via the interferon pathway exhibited in all cell types, restricting virus growth, mitochondria capacity and function of infected cells remained negatively impacted.