The H5 hemagglutinin (HA) gene of a highly pathogenic avian influenza virus (AIV) isolate (A/chicken/Italy/8/98) was cloned and sequenced, and inserted at the non-essential UL50 (dUTPase) gene locus of a virulent strain of infectious laryngotracheitis virus (ILTV). Northern and Western blot analyses of the obtained ILTV recombinants demonstrated stable expression of the HA gene under control of the human cytomegalovirus immediate-early gene promoter. In vitro replication of the HA-expressing ILTV mutants was not affected, and infection of chickens revealed a reduced but still considerable virulence, similar to that of a UL50 gene deletion mutant without foreign gene insertion. The immunized animals produced specific antibodies against ILTV and AIV HA, and were protected against challenge infections with either virulent ILTV, or two different highly pathogenic AIV strains (A/chicken/Italy/8/98, A/chicken/Scotland/59). After challenge, no ILTV could be reisolated from protected animals, and shedding of AIV was considerably reduced. Thus, although attenuation remains to be improved, genetically engineered ILTV live-virus vaccines might be used as vectors to protect chickens also against other pathogens.