Alveolar epithelial cells, which constitute the majority of the alveolar surface, may represent a potential niche for intracellular replication ofLegionella pneumophilathat has been largely overlooked. We examined the phenotypes of a bank of 121 macrophage-defective mutants ofL. pneumophila(designated aspmiandmil) for their cytopathogenicity to and intracellular survival and replication within human alveolar epithelial cells. Our data showed that 91 of 121 mutants that were defective (modest–severe) in macrophages exhibited wild type-like phenotypes in human type I alveolar epithelial cells. In contrast, the other 30 mutants were defective in both macrophages and alveolar epithelial cells. Transmission electron microscopy of the intracellular infection by three mutants showed that the defect in intracellular replication in macrophages and epithelial cells was associated with a defect in recruitment of the RER around the phagosome. Differences in attachment to macrophages and epithelial cells were also exhibited by some of the mutants. Pulmonary infection studies of A/J mice showed that a mutant defective in macrophages but not in alveolar epithelial cells replicated like the wild type strain in the lungs of A/J mice. In contrast, a mutant defective in both macrophages and alveolar epithelial cells failed to replicate and was killed. We conclude that certain distinct genetic loci ofL. pneumophilaare uniquely required for intracellular survival and replication within phagocytic but not epithelial cells, which may be important in vivo.