Researchers generated recombinant pandemic influenza A/H1N1 viruses resistant to neuraminidase inhibitors (NAIs) through substitutions at positions E119 and I222 in the neuraminidase protein. Substitutions at E119 conferred multi-drug resistance, while I222V had a synergistic effect on viruses already resistant via the H274Y mutation. In vitro testing found that E119G/V mutations negatively impacted viral fitness, while I222V was associated with improved fitness. In vivo transmission experiments in ferrets found efficient contact and aerosol transmission for both wild-type and mutant viruses, with no differences in transmission rates.

1. Generation and Characterization of Recombinant Pandemic Influenza A/H1N1 Viruses Resistant to Neuraminidase Inhibitors Andrés Pizzorno, Xavier Bouhy, Yacine Abed, Guy Boivin Research Center in Infectious Diseases of the CHUL, Québec, QC, Canada PCIRN Annual Meeting April 20 - 21, 2011

2. Results – Resistance to NAIs S=sensitive: <5 IC50 fold increase, I=intermediate: 5 to 10 IC50 fold increase, R=resistant: >10 IC50 fold increase Substitutions at E119 confer multi-drug resistance Synergic effect of I222V on H274Y resistant phenotype (Mutations in N2 numbering)

3. Results – Replicative capacity in vitro E119G/V have a negative effect on viral fitness I222V can be associated with improved viral fitness * P<0.05, ** P<0.01

4. Results – In vivo transmission Contact Contact Inoculated Inoculated Aerosol Aerosol Efficient contact and aerosol transmission in the ferret model No differences in transmission rates among the WT and mutants