The role of the genus Acanthamoeba in the process of maintaining, multiplying and transmitting human pathogens has not been well recognized. The aim of our study was to evaluate the association of potentially pathogenic Acanthamoeba spp. with bacteria in strains recently isolated from Lake Malta near the city of Poznan, Poland. Most of the isolated amoebae belonged to the species A. castellanii and A. rhysodes. The majority of the isolates were pathogenic for mice and amoebae could be recovered from their brains and lungs. In those cases when amoebae could not be recovered from the autopsy material histopathological analysis showed changes in tissues indicating bacterial infection. We found that approximately 50% of the isolates were associated with endosymbiotic or endocytobiotic bacteria, e.g., Proteus sp., Micrococcus sp., Chryseobacterium meningosepticum, Clostridium perfringens, Escherichia coli, Salmonella sp., Legionella pneumophila and other species. The presence of bacteria residing inside free-living amoebae possess a great challenge in terms of disease control and sanitation of contaminated water sources.