Influenza virus A/England/26/99 (H3N2) was isolated at the Health Protection Agency, Colindale, London, UK, during the routine surveillance program and has been minimally passaged in MDCK cells [51] . A/Dk/Singapore/97 (H5N3) and A/Dk/ England/62 (H4N6) are typical avian influenza strains that have been passaged in both embryonated chicken eggs and MDCK cells during laboratory handling. Highly pathogenic A/VN/1203/ 04 (H5N1) was biologically derived and minimally passaged in embryonated chicken eggs. A/Udorn/307/72 (H3N2) was passed in baby hamster kidney (BHK) cells and represents a clone expanded once in embryonated chicken eggs. Recombinant viruses, including wild-type A/Victoria/3/75 (H3N2) and mutants in either the A/Victoria/3/75 (H3N2) or A/PR/8/24 (H1N1) background, were generated from cloned cDNA in 293T and MDCK cell co-cultures as previously described [52, 53] . Mutant viruses were generated in either the A/Victoria/3/75 (H3N2) or A/PR/8/34 (H1N1) genetic background as follows: 1) Vic 627PB2; A/Victoria/3/75 containing a lysine to glutamic acid amino acid substitution at position 627; 2) Vic-226-228HA; A/ Victoria/3/75 containing two amino acid substitutions in the HA gene (L226Q, S228G) that confer an avian-like receptor binding preference [6, 40] ; 3) Vic+Chick N1; A/Victoria/3/75 in which segment 6 containing the endogenous N2 NA gene was exchanged for the N1 NA gene from avian isolate A/Chick/Italy/1347/99; 4) Vic-226-228HA+Chick N1; A/Victoria/3/75 containing both L226Q and S228G mutations and the avian N1; 5) PR8+Vic HA/ NA; A/PR/8/34 in which the endogenous H1 and N1 were replaced with the H3 and N2 from A/Victoria/3/75 and 6) PR8+Chick HA/NA (RD3); A/PR/8/34 in which the endogenous H1 and N1 were replaced with the H7 and N1 from A/ Chick/Italy/1347/99. (RD3 was previously described as a candidate vaccine strain [41] .) The last two reassortant viruses were generated by substituting segment 4 and segment 6 from PR8 with those from either A/Victoria/3/75 (H3N2) or A/Chick/ Italy/1347/99 (H7N1). The multi-basic cleavage site in the avian H7 HA gene used in these studies was removed prior to rescue of these recombinant viruses for safety. Available accession numbers (GenBank: http://www.ncbi.nlm.nih.gov.libproxy.lib.unc.edu) are V01086 for A/Victoria/3/75 HA and CAD37074 for A/Chick/ Italy/1347/99 HA.