Nanoscale characterization of Escherichia coli biofilm formed under laminar flow using atomic force microscopy (AFM) and scanning electron microscopy (SEM)

Biofilm contains heterogeneous three-dimensional structures composed of extracellular polymeric substance (EPS), which are greatly influenced by flow conditions. Here, we report a microfluidic platform highly suitable for nanoscale investigation of biofilms formed under laminar flows. This is possible because biofilms formed on glass beads having an average diameter of about 200 μm in the microfluidic device can be easily taken out and located for imaging under high resolution microscopes, such as atomic force microscope (AFM) and scanning electron microscope (SEM). Escherichia coli formed biofilms in the device at various flow conditions (0-50 μL min-1) for several days. SEM showed nanopores (14-100 nm) in biofilm at lower flow rates (0.5 and 5 μL min-1) only at within 3 days, while such small pores were not observed at higher flow rates (50 μL min-1) during the entire culture period (0-5 days). AFM results showed that the surface coverage and roughness of biofilm increased as the flow rate increased. These results suggest that like turbulent flow regime flow rates under laminar flow regime greatly influence on the morphology of biofilms in both micro- and nanoscales.