Fast type I interferon response protects astrocytes from flavivirus infection and virus-induced cytopathic effects

Lindqvist, Richard; Mundt, Filip; Gilthorpe, Jonathan D.; Wölfel, Silke; Gekara, Nelson O.; Kröger, Andrea; Överby, Anna K.

VeroB4 cells were cultured in medium 199/EBSS (HyClone) containing 10 % FBS, 0.1 U/mL penicillin, and 0.1 μg/mL streptomycin (Gibco). Mouse embryonic fibroblasts (MEFs) were grown supplemented with 10 % FBS, 50 μM β-mercaptoethanol (AppliChem), and 2 μg/ mL tetracycline DMEM (Sigma). TBEV strains Hypr 71 (isolated in 1953 from blood of a patient in the Czech Republic), Aina (isolated in 1963 in Irkutsk from the blood of a patient), and Sofjin (isolated in 1937 from patient in Russia and showed 99 % sequence identity to strains Sofjin-Chumakov and SofjinKSY with BLAST) were a kind gift of G. Dobler (the Bundeswehr Institute of Microbiology, Munich, Germany). JEV (Nakayama strain) was a kind gift of S. Vene (Folkhälsoinstitutet, Stockholm, Sweden). WNV (isolated in 2003 in Israel WNV_0304h_ISR00, passage number 5) is a kind gift of S. Vene. Stocks were generated in VeroB4 cells. Vesicular stomatitis virus (VSV)-eGFP was propagated in VeroB4 cells. ZIKV MR 766 (isolated in 1947 in Zika Forest, Uganda) was propagated in VeroB4 cells and originally provided by Robert Shope (Yale Arbovirus Research Unit, New Haven, CT, USA) as a reference strain to Jürgen Pilaski (University of Düsseldorf, Germany), who kindly transferred the strain to the Bundeswehr Institute of Microbiology, Munich, Germany. TBEV strains, JEV, and ZIKV are cell culture-adapted reference strains with an unknown passage history. Cell monolayers were infected with virus for 1 h at 37°C. The virus inoculum was then removed and replaced with DMEM supplemented with 2 % FBS, 0.1 U/mL penicillin, and 0.1 μg/mL streptomycin. Viral titers were determined by focus forming assay as previously described [42] .