Influenza A viruses of the H7, H5, and H1 subtype used in this study are shown in Table 1 . Virus stocks were propagated in the allantoic fluid cavity of 10 day old embryonated hens' eggs as previously described [26] ; virus stocks were confirmed by sequencing to be free of mutations. The 50% egg infectious dose (EID 50 ) for each virus stock was calculated by the method of Reed and Muench [54] following serial titration in eggs. Fluorescenttagged virus (NL/219-FL) was generated using formalin-inactivated NL/219 virus and a SAIVI Antibody Alexa Fluor 680 Labeling kit (Invitrogen) per manufacturer's instructions as previously described [32] . All experiments with HPAI viruses were conducted under biosafety level 3 containment, including enhancements required by the U.S. Department of Agriculture and the Select Agent Program [55] .