Viability assessment of Y. pseudotuberculosis by vital staining. (A) Various combinations of live and isopropanol-killed Y. pseudotuberculosis were mixed, stained with SYTO 9 and propidium iodide, and analyzed by flow cytometry. Shown in the left column are the light-scattering plots of cultures containing either 100% live bacteria (top), 100% killed bacteria (bottom), or an equal mixture of live and dead bacteria (middle). The histograms shown to the right of each light-scattering plot are the corresponding signals in the FL1 (green channel) of the R1-gated events. The Medium fluorescence intensity of the events within the Ml region were 184 (top) 99 (middle) and 30 (bottom). (B) RAW cells were infected with either the wild-type (top) or yopB (bottom) strains at a MOI of 20 for 2 hours following which the bacteria were recovered, stained with SYTO 9/propidium iodide, and analyzed and presented as in (A). The percentages of the events falling in either the lower left (11) or upper right (ur) quadrants in the light-scattering plots were 36% (11) and 42% (ur) for the wild type and 43% (11) and 37% (ur) for yopB. In parenthesis is shown the medium fluorescence intensity of the events within the M1 region.