Figure 1. Analyses of the interconnected furrow network that guide self-organization of P. aeruginosa interstitial biofilms. (A) High magnification 3D optical profilometery scan of the region where the time-lapse phase-contrast microscopy (Movie S1) was performed. Overlapping scan regions were manually stitched to provide a larger field of view. Scale bar indicates 100 μm. (B) Overlapping phase contrast microscopy images of the region observed in Movie S1 were captured after the time-series was completed and manually stitched to provide a larger field of view that captures the leading edge rafts that had migrated out of frame in Movie S1. Scale bar indicates 100 μm. Relevant morphological features of the interstitial biofilms are labeled. (C) Correlation of the biofilm cell trails to the underlying furrow network. The stitched image depicted in (B) was false colored green and overlaid onto the 3D optical profilometery image depicted in (A). The cellular network lays completely within the furrow network. Scale bar indicates 100 μm. (D) Analysis of furrow widths from the different areas within the interstitial biofilms indicated in (B). Analysis was performed using the high magnification profilometery scans with the number of measurements for each group as follows: raft head n = 21, raft trails n = 30, behind rafts n = 43, and lattice n = 77. Analyses are depicted as mean ± SD P value < 0.0001 determined using a one-way ANOVA test. (E) 3D optical profilometery image (500μm x 280μm) of the underlying semi-solid media where Movie S1 was performed using low magnification to obtain an overview of the furrow network relative to the virgin territories. Height scale is equivalent for (A), (C), and (E).