FIG 3.
Replication kinetics of MERS-CoV on hamster and ferret cell lines expressing human, hamster, or ferret DPP4. (A) Human DPP4 (red), hamster DPP4 (blue), ferret DPP4 (blue), and GFP (green) were expressed in BHK (circles) or primary ferret (squares) cells. Twenty-four hours posttransfection, cells were inoculated with MERS-CoV using an MOI of 1 TCID50/cell. Supernatants were harvested at 0, 24, 48, and 72 hpi, and viral titers were determined by endpoint titration in quadruplicate in VeroE6 cells. Mean titers were calculated from three independent experiments. Error bars indicate standard deviations. (B) BHK or primary ferret cells were left untransfected (red) or transfected with DPP4 (blue) and stained 24 h posttransfection using anti-DPP4 (R&D) and an FITC-conjugated secondary antibody (Life Technologies). Samples were collected using an LSRII flow cytometer (BD Biosciences) and analyzed using FlowJo software. (C) Expression of DPP4 mRNA was measured via qRT-PCR. Relative fold increase was calculated by the comparative CT method (35), where DPP4 expression is normalized to HPRT.