Legionella pneumophila is a facultative intracellular parasite able to survive within both human monocytes and amoebae. We have demonstrated that processing of L. pneumophila by the free-living amoeba Acanthamoeba castellanii shows many similarities to the processing of L. pneumophila by monocytes. These similarities include uptake of L. pneumophila by coiling phagocytosis and the subsequent confinement of L. pneumophila in a ribosome-studded phagosome. In addition, as in monocytes, inhibition of lysosomal fusion with phagosomes containing L. pneumophila was detected in amoebae. With all clinical isolates, inhibition of phagosomes-lysosome fusion correlated with virulence. However, with one of the environmental isolates tested, no significant difference in phagosome-lysosome fusion was observed between the virulent and avirulent forms. These results indicate that the avirulent form of this isolate differed from the virulent form in some other respect critical to intracellular survival. Therefore, intracellular multiplication of L. pneumophila within A. castellanii may not be solely dependent upon the inhibition of lysosomal fusion.