Strains and inoculation procedure

We used PAO1-gfp (ATCC 7700) with improved genes which expresses green fluorescent protein (GFP) in these studies (Liu et al. 2009; Zhang et al. 2011). In the natural environment, P. aeruginosa is a normal biofilm-forming organism which was studied as a model organism for many biofilm studies (Mann and Wozniak 2011), it is also an opportunistic pathogen (Goeres et al. 2009) which has many advantage with fine properties for experimentation. The colony was moved to 3 ml of sterile tryptic soy broth (TSB) in a culture tube and through an overnight shaking at 225 rpm and 37 °C. For the mid-log phase culture, fifty microliters of this culture was transferred to 3 ml of sterile tryptic soy broth (TSB), at 225 rpm and 37 °C, and then diluted with sterile 1% TSB to OD₆₀₀ = 0.01 for inoculation into the flow cells. To quantify the turbidity of liquid cultures was used optical density (OD) measured at 600 nm, which was used as a corresponding measure of cell density. Under no flow (stagnant) condition, inoculated bacteria were injected into flow cell and attach to the cover slip for 1 h, and then continuously pumped into nutrient with 1% TSB medium. To avoid the disturbance by the medium, the biofilms were inoculated and cultured for 3 days to grow mature at a flow rate of 10 ml h⁻¹.