Commonality of Virulence-Promoting Function in Rhodococcus equi Virulence Associated Proteins (Vaps)

Figure 6

Lysosomal acidity is reduced by Vaps A, B, K, and N. (a) Fluorescent images of macrophages infected with fluorescent-peptidoglycan R. equi strains 103+ or 103+ (ΔvapA). Peptidoglycan is in green and DNA is in blue. Scale bars 10 μm. (b) Quantitative pH measurements of R.equi-containing phagosomes using fluorescein-labelled bacteria. Lysosomal pH was measured using fluorescein-labelled dextran, and VapA-positive compartments were measured using VapA-pHluorin2. Fluorescein and pHluorin2 calibration curves are shown in Figure S3. Data are (). , . (c) Flow cytometry gating strategy used to select live cells based on DAPI exclusion (left plot; -axis = DAPI fluorescence; -axis = forward light scatter) and single cells (right plot; -axis is light scattering based on cell area; -axis is light scattering based on cell height). (d) Representative flow cytometry overlay histogram of LysoTracker (LT) fluorescence in cells treated with or without Vaps and bafilomycin-A₁ (Bafilomycin). 5,000 cells were counted for each condition. (e) Quantification of LysoTracker fluorescence in cells as shown in (d). Data are mean median fluorescence intensity (). , , .

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