FIG 2 .
Recombinant Ud viruses expressing a PB1 mRNA with CAI values similar to those of recent H3N2 viruses replicate better than the WT 1972 Ud virus in IFN-treated human cells. Cells were pretreated with or without 1,000 IU/ml IFN for 36 h. (a) Growth curves, carried out in triplicate, of virus construct 1 and WT Ud viruses in A549 cells infected at a MOI of 0.01. The means are reported here, with error bars indicating standard errors. (b) Growth curve of construct 2 and WT Ud viruses in A549 cells infected at a MOI of 0.01. (c) Viral protein production in A549 cells infected with a MOI of 5 PFU/cell of construct 1, construct 2, or WT Ud virus. At 9 h after infection, cell extracts were immunoblotted with appropriate antibodies to detect the indicated viral proteins or actin (see Text S1). (d) RT-PCR results for the mRNAs and vRNAs encoded by the (M1) NP and PB1 genes. RNA was isolated from IFN-treated and mock treatment cells at 3 h postinfection of 5 PFU/cell of construct 1, construct 2, or WT Ud virus, and the fold inhibition caused by IFN treatment was measured in triplicate by the threshold cycle (−ΔΔCT) method. Both constructs showed significantly reduced inhibition of viral RNA synthesis due to IFN treatment compared to the WT (P < 0.05 [two-tailed t test]) for all measured RNAs. Error bars (standard errors) are shown.