FIG 5.

Flow cytometric analysis of A. castellanii amoebae infected with wild-type and T2SS mutant L. pneumophila. (A) A. castellanii amoebae were inoculated with either wild-type strain 130b (WT) expressing plasmid-encoded GFP or lspF mutant NU275 expressing GFP (lspF), as described in the legend of Fig. 4. Infected amoebae were incubated for an additional 12 h and collected for flow cytometry, with the results from wild-type infection shown on the left and those from mutant infection on the right. The GFP-positive interval gate was drawn such that it excluded signal from uninfected amoebae. The percentage of GFP-positive amoebae is indicated above each interval gate. (B) Histogram overlay of the GFP-positive populations from panel A to compare the GFP intensities of individual infected amoebae. The median fluorescence intensity (MFI) for each of the infections is indicated. The data presented are representative of three independent experiments.