Recombinant reassortment technology was used to prepare H5N1 influenza vaccine strains containing a modified hemagglutinin (HA) gene and neuraminidase gene from the A/Hong Kong/156/97 and A/Hong Kong/483/97 isolates and the internal genes from the attenuated cold-adapted A/Ann Arbor/6/60 influenza virus strain. The HA cleavage site (HA1/HA2) of each H5N1 isolate was modified to resemble that of "low-pathogenic" avian strains. Five of 6 basic amino acids at the cleavage site were deleted, and a threonine was added upstream of the remaining arginine. The H5 HA cleavage site modification resulted in the expected trypsin-dependent phenotype without altering the antigenic character of the H5 HA molecule. The temperature-sensitive and cold-adapted phenotype of the attenuated parent virus was maintained in the recombinant strains, and they grew to 108.5-9.4 EID50/mL in eggs. Both H5N1 vaccine virus strains were safe and immunogenic in ferrets and protected chickens against wild-type H5N1 virus challenge.