Graphic representation of the proteins identified in ECVs and phagolysosomes containing latex beads.

<p>(A): Schematic representation of proteins present in ECVs and latex bead phagolysosomes; (B): Proteins shared by ECVs and latex bead-containing phagolysosomes were grouped according to their cellular locations; (C): Proteins detected in all latex bead-containing phagolysosomes but not detected in any ECVs were classified based on their cellular locations; (D): Proteins present only in ECVs were grouped according to their cellular locations.</p

LysoSensor Green DND-189 and DND-153 labeling of <i>E. chaffeensis</i>-infected DH82 cells.

<p>(A and D): The confocal images containing DAPI-stained nuclei and <i>E. chaffeensis</i> morulae (blue); (B): LysoSensor Green DND-153-labeled infected DH82 cells; (C): merged image of A and B; (E): LysoSensor Green DND-189-labeled infected DH82 cells (green); (F): merged image of D and E; (G): DAPI-stained uninfected DH82 cells; (H): LysoSensor probe-labeled uninfected DH82 cells; (I): merged image of G and H. Arrows indicate <i>E. chaffeensis</i> morulae. These results were confirmed in three independent labeling experiments.</p

Confocal microscopy images of selected cell markers in infected DH82 cells.

<p>When the infectivity reached 95%, the infected cells were fixed and permeabilized. The slides were incubated with the primary antibodies specific to the selected makers (anti-CD71, anti-EEA1, anti-Rab5, anti-LAMP II, anti-Rab7, anti-VATPase, anti-MAP LC3, or anti-BECN antibodies), and then incubated with corresponding secondary conjugated antibodies. In all images, the blue indicated DAPI-stained nuclei (large blue body) or <i>E. chaffeensis</i> morulae; the red indicated host cell proteins labeled with antibodies to CD71 (A), EEA1 (B), Rab5 (C), LAMP II (D), Rab 7 (E), cathepsin D (F), VATPase (G), MAP LC3 (H), and BECN (I). Uninfected DH82 cells incubated with primary antibodies and corresponding secondary antibodies (J) and infected DH82 cells incubated with secondary conjugated antibodies alone (K) served as negative controls. In each panel, whether the DAPI staining was co-stained with host cell proteins determined the co-localization of <i>E. chaffeensis</i> and the host cell proteins. These results were confirmed in three independent experiments.</p

Co-inoculation of DH82 cells with <i>E. chaffeensis</i> and formalin-fixed <i>E. coli</i>.

<p>Three days after <i>E. chaffeensis</i> infection, DH82 cells were inoculated with formalin-fixed <i>E. coli</i> and stained with DAPI and LysoTracker Red. (A): The confocal images containing DAPI-stained nuclei, <i>E. chaffeensis</i> morulae and formalin-fixed <i>E. coli</i> (green); (B): LysoTracker Red-stained infected DH82 cells; (C): merged image of A and B. When the DAPI and LysoTracker Red images were merged (C), co-localization was not apparent providing evidence that ECVs did not fuse with the cellular lysosomes. But, <i>E. coli</i> in the same cell near the nuclei were co-localized with Lysotracker Red (yellow in the merged panel).</p