Biofilm Development is Governed by Cooperative Attachment and Adhesion Maintenance Proteins

The opportunistic fungal pathogen is capable of adhering to the oral mucosa despite forces created by salivary flow. Although many fungal adhesion proteins have been identified, less is known about the temporal development of cell adhesion and biofilm growth in a flow environment. In this study, we use a flow system with real-time imaging of cells as they adhere and grow. Rates of cell attachment and dispersion of knockout strains of putative adhesins, transcription factors, and deletions with a hyperfilamentous phenotype were quantified during 18 h of biofilm development. Cell adhesion under flow is a multi-phase process initiated with cell rolling, then an initial firm attachment to the substrate occurs. After attachment, cells enter a growth phase where cells either commit to adherence or disperse. Δ, Δ, Δ, and Δ cells had significantly reduced initial attachment and subsequent adhesion, while Δ/Δ had no change in initial attachment but reduced adhesion maintenance. WT cells had increased adhesion during the late growth phase when hyphae were more highly expressed. Hyperfilamentous strains had 10-fold higher total biofilm growth, a result of significantly reduced detachment rates, showing that hyphal morphogenesis is important for adhesion maintenance in the developing biofilm. The rate of biomass dispersion was most important for determining the density of the mature biomass. Adhesion maintenance was mediated in part by Ywp1, a protein previously thought to regulate dispersion, thus it functions as an adhesion maintenance protein in .