Figure 4: Morphology of biofilms grown under ALI (A-C) and LLI (D-F) interfaces: A) and D), representative 3D reconstruction of z-stack CLSM images B) and E) are 3D projected and orthoclase views of z-stacks obtained from the CLSM images. The live bacteria were stained with SYTO9 (green), and dead bacteria were stained red-fluorescent propidium iodide. C) and F) are representative SEM images of PAO1 biofilms showing the mushroom-shaped aggregates, the upper right corner is magnified ’mushroom-shape’ aggregates.
During biofilm formation, both live and dead bacteria are encased within the EPS matrix. The CLSM images (Fig.4) show a distinct qualitative difference between ALI and LLI biofilms in terms of the amount of live and dead cell. More specifically, biofilm grown on ALI had a higher ratio of living bacteria than the biofilm formed at LLI (Fig. 5). This result corroborates with our previous results in Fig. 3, showing that biofilms formed at LLI were more susceptible to the physical disturbance compared to ALI cultured biofilm, leaving more bacteria on the membrane for ALI model biofilm.