An immunohistochemical investigation was performed to assess tissue tropism and viral replication of Italian H7N1 isolates belonging to different lineages in developing chicken, turkey, Muscovy duck, and mallard duck embryos. Low-pathogenic avian influenza (LPAI) isolates were selected on the basis of the location in the phylogenetic tree; a progenitor strain, A/turkey/Italy/977/V99 (exhibiting no additional glycosylation sites, nAGS), strain A/turkey/Italy/2379/V99 (AGS in position 123), and strain A/turkey/Italy/3675/V99 (AGS in position 149) were selected. The latter two strains belonged to distinct lineages originating from the pool of progenitor strains. The highly pathogenic avian influenza (HPAI) isolate A/turkey/Italy/4580/V99 was also included in the test. All the embryos tested supported the growth of HPAI. The LPAI isolates replicated readily in the allantoic layer of the chorioallantoic membrane of all the species tested and did not replicate to detectable levels in the developing chicken, turkey, and Muscovy duck embryos. In contrast, they replicated to different extents in the respiratory tract of the developing mallard embryo. The findings indicate that the pathogenesis of LPAI infections in mallard embryos is different to that observed in other species and should be investigated further.