The NA gene of the new pandemic H1N1 virus was acquired from Eurasian swine influenza H1N1 virus, a lineage of virus that crossed from bird to pigs in the late 1970s. In this background, the small compromise in enzyme affinity for sialic acid substrate (observed by a 1.3-fold increase in the NA K m value) and the decrease in cell surface expression that results from the H275Y mutation, 14 had no effect on virus growth in MDCK cell culture. On the other hand, in vivo NA must cleave complex substrates to mediate virus release from an infected airway cell and gain access through a complex layer of mucins to the new target cell. Subtle decreases in NA activity or cell surface expression may have more profound consequences in the airway than in monoculture. To probe this, we tested the NA activity of the mutated virus in assays that presented large sialylated substrates: in a red cell elution assay we detected a 4-fold compromise in the ability of the virus with H275Y mutation to mediate desialylation of chicken erythrocytes, although the biological significance of this assay is not entirely clear. In differentiated cultures of human airway cells, the difference detected in replication of wild-type and mutant virus was not statistically significant on either of two separate occasions (Figure 1 and data not shown) . However, in competition assays in HAE cultures, the wild-type virus out-competed growth of the drug-resistant strain suggesting that, in the absence of drug, the 275Y motif carries a fitness cost in the environment of the human airway.