Differential Production of Psl in Planktonic Cells Leads to Two Distinctive Attachment Phenotypes in Pseudomonas aeruginosa

The attachment of planktonic cells to surfaces is the first and most crucial step in biofilm formation. In this paper, we show that planktonic cells of Pseudomonas aeruginosa differently attach to surfaces. Typically, in the later exponential phase, approximately 80% of the cells can quickly attach to surfaces within 15 min, whereas approximately 20% of the cells slowly attach to surfaces, which greatly affects the initial stage of biofilm formation in the presence of flows. This is because fast-attaching cells are more likely to attach on surfaces to form microcolonies, whereas slow-attaching cells are more likely to remain in the mobile phase. This scenario is different from the previous understanding of biofilm formation in the initial stage, in which planktonic cells were thought to uniformly attach to surfaces. Most notably, the results of this study show that the different attachment manner of planktonic cells to surfaces affects the subsequent stages of biofilm formation. This research highlights that the phenotypic variations in planktonic cells plays significant roles in various stages of biofilm formation. ABSTRACT Exponentially growing bacteria in a well-mixed planktonic culture are generally assumed to be physiologically and phenotypically uniform and distinct from their genetically identical counterparts living in biofilms. Using a combination of high spatiotemporal microscopy and a bacterial tracking algorithm, in this study, we showed that planktonic cells of Pseudomonas aeruginosa differently attached to surfaces even when they remained in the exponential phase. We consistently observed that fast- and slow-attaching phenotypes coexist in planktonic cells, regardless of their growth phase. Furthermore, we found that (i) the distinct attaching phenotypes of planktonic cells resulted from the differential production of Psl and (ii) the RsmYZ/RsmA signaling pathway mainly regulated the differential production of Psl. Our results indicate that the differential production of Psl in P. aeruginosa plays a significant role in biofilm development and formation. IMPORTANCE The attachment of planktonic cells to surfaces is the first and most crucial step in biofilm formation. In this paper, we show that planktonic cells of Pseudomonas aeruginosa differently attach to surfaces. Typically, in the later exponential phase, approximately 80% of the cells can quickly attach to surfaces within 15 min, whereas approximately 20% of the cells slowly attach to surfaces, which greatly affects the initial stage of biofilm formation in the presence of flows. This is because fast-attaching cells are more likely to attach on surfaces to form microcolonies, whereas slow-attaching cells are more likely to remain in the mobile phase. This scenario is different from the previous understanding of biofilm formation in the initial stage, in which planktonic cells were thought to uniformly attach to surfaces. Most notably, the results of this study show that the different attachment manner of planktonic cells to surfaces affects the subsequent stages of biofilm formation. This research highlights that the phenotypic variations in planktonic cells plays significant roles in various stages of biofilm formation.