Avian H7 influenza viruses from both the Eurasian and North American lineage have caused outbreaks in poultry since 2002, with confirmed human infection occurring during outbreaks in The Netherlands, British Columbia, and the United Kingdom. The majority of H7 infections have resulted in self-limiting conjunctivitis, whereas probable human-to-human transmission has been rare. Here, we used glycan microarray technology to determine the receptor-binding preference of Eurasian and North American lineage H7 influenza viruses and their transmissibility in the ferret model. We found that highly pathogenic H7N7 viruses from The Netherlands in 2003 maintained the classic avian-binding preference for α2-3-linked sialic acids (SA) and are not readily transmissible in ferrets, as observed previously for highly pathogenic H5N1 viruses. However, H7N3 viruses isolated from Canada in 2004 and H7N2 viruses from the northeastern United States isolated in 2002-2003 possessed an HA with increased affinity toward α2-6-linked SA, the linkage type found prominently on human tracheal epithelial cells. We identified a low pathogenic H7N2 virus isolated from a man in New York in 2003, A/NY/107/03, which replicated efficiently in the upper respiratory tract of ferrets and was capable of transmission in this species by direct contact. These results indicate that H7 influenza viruses from the North American lineage have acquired sialic acid-binding properties that more closely resemble those of human influenza viruses and have the potential to spread to naïve animals. © 2008 by The National Academy of Sciences of the USA.