The H5N1 highly pathogenic avian influenza (HPAI) outbreak and subsequent infection of humans that occurred in Hong Kong in 1997 kindled the recognition of influenza A viruses as potentially zoonotic pathogens. In order to resolve questions regarding the perpetuation and transmission of this virus, a repertoire of avian and mammalian species that could serve as suitable hosts for the A/chicken/Hong Kong/220/97 (H5N1) (chicken/HK) was determined. The results of this investigation confirm that this H5N1 virus can infect and replicate in multiple avian species following intranasal inoculation. However, the clinical disease, demonstration of viral antigen, and consistency of virus reisolation from swabs and tissues were variable among the species, even species within the same order. The greatest consistency was demonstrated among the gallinaceous birds, which were undoubtedly the most susceptible species in terms of the virulence and lethality of the chicken/HK virus. The chicken/HK virus also demonstrated high pathogenicity for Zebra finches (Taeniopygia guttata), House finches (Carpodacus mexicanus), and budgerigars (Melopsittacus undulatus), with each of these species suffering high morbidity and mortality relative to systemic infection. Despite a lack of mortality, the chicken/HK virus caused substantial morbidity relative to neurological dysfunction in emus (Dramaius novaehollandiae) and Embden geese (Anser anser). In contrast, several other avian species, including House sparrows (Passer domesticus), European starlings (Sturnus vulgaris), Pekin ducks (Anas platyrhynchos), pigeons (Columbia livia), and Laughing gulls (Larus atricilla) as well as two mammalian species, including rats and rabbits, were refractory to productive viral replication and associated clinical disease. In total, this investigation demonstrates that the H5N1 Hong Kong-origin virus can display a broad range of virulence from apathogenic to highly pathogenic among the susceptible avian species. Furthermore, profound differences were observed in the susceptibility of selected species from the same order, indicating that it may be difficult to predict the susceptibility of a particular avian species based on its phylogenetic classification.