Fig. 5.
Reversion of R65-PB2K627E in mice but not in chickens. (A) Mouse passages of R65-PB2K627E. Lungs and brains were taken on days 1, 3, 5, or 9 from mice infected intranasally with 105 (1, 5, or 9 days p. i.) or 104 (3 days p. i.) PFU virus. From organ homogenates, viral RNA was isolated, and the PB2 segment was amplified by RT-PCR and sequenced to monitor the presence of PB2-627E. Red arrows on electropherograms indicate emergence of revertants carrying PB2-G27K. (B) Chicken passage of R65-PB2K627E. Brains, lungs, pancreas, and spleens from two R65-PB2K627E-infected and one R65-infected animal (104 PFU oculonasally) were homogenized, and the virus was propagated in avian DF-1 cells. From supernatants, viral RNA was isolated, and the PB2 segment was amplified by RT-PCR and sequenced to monitor the presence of PB2-627E.