Legionella has been shown to not only survive but multiply intracellularly in 14 species of protozoa, such as Hartmannella, Acanthamoeba, Naegleria, and Tetrahymena. Often the contaminated water supply associated with Legionnaires' disease outbreaks have been found to contain amoebae and/or protozoa. Legionella survive in water and moist soil environments as intracellular parasites of free-living protozoa. While growth of legionellae in the absence of protozoa has been documented only on laboratory media.

Legionella survive within biofilms in building water systems. The bacteria are often more easily detected from biofilm swab samples than the free-flowing water. This suggests that the majority of Legionella are associated with biofilm. These biofilms provide shelter (and protection from biocides) and nutrients to the bacteria. A laboratory study using artificially constructed biofilm showed that Legionella will survive in the absence of amoeba, but will only multiply when protozoa are also present in the biofilm. This is not to say that it is impossible for Legionella to multiply in the absence of protozoa, as the diversity of biofilm makeup of nutrients and other bacteria is immense, it mealy suggests that it is possible that protozoa are required for multiplication.

It has been suggested that the infectious particle for Legionnaires' disease is not the bacteria alone but the bacteria within an amoeba. This has not been proven but  there are many observations which support the principle that protozoa play a major role in the infection of L. pneumophila. These are listed below

1. Many protozoa allow bacteria to multiply within the cell. This being proposed as the only means of replication  in the natural environment.
   
2. During Legionnaires' disease outbreaks investigations of the water implicated, amoebae and bacteria have often been isolated from the same source of infection. Further when the isolated amoebae has been cultured in the presence of the Legionella also isolated it has been shown that the bacteria will multiply within the amoeba cell.
   
3. It has been shown that once Legionella have multiplied within an amoeba, the bacteria shows a large increase in resistance to high temperature, acidity, and high osmolarity.
   
4. It has also been shown that Legionella isolated from amoeba/protozoa are more resistant to chemical disinfection and biocides than laboratory grown bacteria. These two points (3 and 4) suggests that this process helps the bacteria survive both in the natural environment and in artificial environments such as cooling towers.
   
5. Protozoa have been shown to release vesicles that contain numerous L. pneumophila bacteria, these vesicles are small enough to be inhaled to the lower lungs, and the bacteria within the vesicles are highly resistant to biocides. This means that while they are in the free-flowing water the bacteria are difficult to kill.
   
6. Once the bacteria have been released from the protozoan they show an increased ability to infect mammalian cells in laboratory studies.
   
7. Occasionally the bacterial counts made of water sources responsible for Legionnaires' disease are low or undetectable and it is possible that this may be due to the fact that the bacteria are more infective because they have multiplied in the protozoan host or that water tested by the laboratory has included intact vesicles that although they may contain many Legionella bacteria will only develop into one colony on culture plates and give falsely low count results.
   
8. There are known to be alive Legionella that can be grown in the laboratory but these bacteria have been shown to become revived by growing them within protozoa. This suggest that failure to isolate the bacteria from environmental sources of infection may be due to this dormant phase of the bacteria.
   
9. It has never been shown that Legionella can be spread from person-to-person. Despite other respiratory pathogens such as tuberculosis being able to be passed on by this route. The only known source of Legionella transmission is environmental aerosols from many human-made devices such as shower heads, water fountains, whirlpools, and cooling towers of air-conditioning systems. This suggests that it is the presence of protozoans either enhances the capacity for infection or is necessary for infection to occur.