Correlating Staphylococcus epidermidis biofilm pH Gradients with Local Biofilm Mechanical Properties to Reveal Contributions of pH to Biofilm Disassembly and Dispersion

Bacterial biofilms are surface-adherent communities of microbial cells encased within a protective extracellular polymeric matrix that are frequently responsible for device-associated and chronic infections. The biofilm lifecycle consists of the following stages: initial bacterial surface adhesion, accumulation of bacteria and matrix materials on a surface, maturation into an organized community, and disassembly and dispersion of cells from the surface. The final stage of the biofilm lifecycle—disassembly and dispersion—contributes to the spread of infection within the body. The heterogeneity of the structural and mechanical properties of biofilms can result in the formation of pH gradients within biofilms due to the nonuniform distribution of nutrients or metabolic waste products. Bulk changes to the pH of staphylococcal biofilms above 7.3 have been shown to soften staphylococcal biofilm matrix materials¹. However, the extent to which biofilms can naturally control their mechanics through modulating pH is not well understood. Here we correlate microscale pH maps with biofilm mechanics to determine the role of natural pH gradients in controlling Staphylococcus epidermidis biofilm mechanics and disassembly. We use confocal laser scanning microscopy (CLSM) and image analysis techniques to quantitatively map pH values and determine the local mechanical behavior of biofilms using passive microrheology. We find that S. epidermidis biofilms grown for 24 hours contain more acidic pH regimes than those grown for 6, 12, and 18 hours. Spatially, we observe the most acidic microenvironments to be located close to the biofilm-substrate interface. Additionally, local S. epidermidis biofilm mechanics are found to vary spatially with pH, where regions of the biofilm with higher pH (pH > 6.9) are mechanically softer than regions of the biofilm with lower pH (pH < 6.9). This work advances the understanding of how natural biofilm pH gradients contribute to biofilm mechanics and reveals potential contributions of matrix phase stability to the disassembly and dispersion of staphylococcal biofilms.

¹Stewart, E.J. et al. Scientific Reports, (2015).