Fig. 4. PB2 A271T exhibits decreased polymerase activity in vitro and decreased mortality in ferrets relative to wild-type A/mink (H5N1).

Polymerase activity was determined by performing mini-genome assays in 293T cells at 33 and 37 °C. A Activity of the wild-type polymerase versus the polymerase with the mutation PB2 A271T at 24 h post-transfection. B Activity of the polymerase carrying the PB2 A271T mutation expressed as a percent of the activity of the wild-type polymerase. Data shown are mean ± SD from 1 of 3 representative independent experiments with n = 3 biologically independent samples/experiment. C–E display ferret studies. Four ferrets per group were inoculated with 10² TCID50/mL of either wild-type A/mink (H5N1) (i.e., PB2 271A) or A/mink (H5N1) with PB2 A271T mutation. Nasal wash samples were collected every other day, and weight loss and clinical signs were monitored daily for 14 days. C Displays viral titers as mean ± SD, D shows weight loss for individual animals, and E depicts survival (n = 4/group). Blue and green bars and lines represent wild-type PB2 and PB2 A271T, respectively. Dotted line denotes limit of detection. *p = 0.015 and **p = 0.018 using a two-tailed unpaired Student’s t test with Welch’s correction. Source data are provided as a Source Data file.