Investigating the interferon antagonistic abilities of louping ill virus, a neglected animal pathogen endemic to the UK

Louping ill virus (LIV; Flavivirus, Flaviviridae) is an important—but poorly characterized—animal pathogen of significant economic concern within the UK. Transmitted by ticks, LIV predominantly causes disease in ruminants and grouse, resulting in heavy losses. LIV is closely related to another flavivirus—tick-borne encephalitis virus (TBEV) which, unlike LIV, is a significant human pathogen. The molecular mechanisms that underpin host restriction in these viruses are poorly understood however, previously it has been shown that the TBEV non-structural (NS) proteins do not act as type-I interferon (IFN) antagonists, unlike many other Flavivirus NS proteins. Therefore, to facilitate comparison with LIV we investigated the possible antagonistic actions of the LIV NS proteins using a luciferase-based IFN reporter assay. Utilising this assay we identified six LIV NS proteins that function as antagonists throughout the IFN induction cascade. We also identified and modelled a subgenomic flavivirus RNA (sfRNA) that is produced during LIV infection and is similar in structure to TBEV sfRNA. We found that the LIV and TBEV sfRNAs antagonise RIG-I, indicating that the IFN antagonistic ability of sfRNA is not limited to mosquito-borne flaviviruses. Finally, we established the first LIV reverse genetics system using circular polymerase extension reaction (CPER). This powerful tool can be used to produce chimeric viruses which will allow further investigation into the factors governing host restriction and virulence in tick-borne flaviviruses. Investigating the mechanisms that underlie LIV infection aids our understanding of interferon antagonism in flaviviruses and the molecular determinants of host restriction.