In this study a flow based biofilm model system was used to simulate the formation of Pseudomonas aeruginosa biofilms on a stainless steel surface. To investigate the complexity of biofilm-associated P. aeruginosa populations a combination of microscopic observations and flow cytometric analysis (FCM) was adopted. Biofilm-associated P. aeruginosa cells were evaluated (1) under optimal vs reduced nutrient-availability at the initial adhesion stage, and (2) irrespective of nutrient-availability within a mature biofilm. Microscopic estimation of the extent of attachment revealed more effective colonization upon optimal vs starvation conditions. FCM allowed an in situ evaluation of P. aeruginosa vitality, using cellular redox potential measurements to discriminate active, mid-active and non-active sub-populations. Samples from recently attached cells and mature biofilms showed significant differences in the percentages of bacterial cells from the defined sub-populations. The approach demonstrated that distribution of individual P. aeruginosa sub-populations was influenced by the stage of the biofilm life-cycle and nutrient availability.
Keywords: Biofilm; Pseudomonas aeruginosa; cell sorting; single-cell analysis; solid surfaces.