Figure 6. Cell death and lysis contributes to the Psl matrix cavity formation.

In panel A, Psl matrix was stained in green, whereas in panel B, the green fluorescent signal represents viable cells stained by SYTO9. In all panels, red fluorescence is due to propidium iodide (PI) staining of either eDNA (weak and diffuse red) or cells with a compromised cell membrane (dead cells, bright concentrated red). Bar, 5 µm for WFPA801 in panel A and 10 µm for the other images. (A) Images of 2-day-old WFPA801 and rpoN mutant biofilms stained by HHA-FITC (green) and PI (red). A top-down view of 3D reconstructed images is shown in left panel. Sets of images optically sectioned (horizontally) at the neck of the same microcolony are shown in the right panel. The corresponding merge image of Psl matrix and DNA matrix is shown in the middle panel (large square). Two corresponding vertical section images are also shown (rectangle). (B) LIVE/DEAD viability staining of PAO1 and WFPA801 biofilm microcolonies show that there are viable swimming cells in the lower center of the microcolony and dead cells/extracellular DNA that fill up all void spaces. Three microcolonies prior to dispersion and one microcolony (the upper right) post the seeding dispersal are shown. The arrow points out the swimming cells in the center of the microcolony.