Figure8a-d_m.nd2

Figure 8. Co-culture of S. aureus MRSA and P. aeruginosa biofilms at 4 hours (A – D), 8 hours (E – H), and 24 hours (I – L). Wheat Germ Agglutinin Alexa Fluor 647 was used to identify S. aureus cells (A, E, I) while SYTO 84 was used to label both intracellular and extracellular DNA (B, F, J) including eDNA in the matrix of the biofilm. Panels C, G, and K show membrane depolarization using DiBAC4 voltage sensitive fluorophore. Panels D, H, and L are merged images of all fluorophores used showing overlap between S. aureus cells and cells that are depolarized which is further show throughout the entire biofilm (M – O). The number of S. aureus cells and corrected total cell flu-orescence was calculated to further shown the depolarization present (P – Q). Statistical analysis was performed by one-way ANOVA with Dunnett’s multiple comparison test with 95% (p<0.05) of significance. ****: p < 0.0001.