Figure 2.

Change of osmolarity on swarm plates measured with fixed liposome spots. (A) Fluorescence signals of sulforhodamine 101 (R) and calcein (G) recorded on a swarm. Time zero corresponds to the time when bacteria first invaded the liposome spot. (B) G/R ratio after correction for photobleaching. The gray dotted line is the reference baseline for osmolarity on the agar surface, which changed because of evaporation. The time required for the measurement, set by the dimensions of the swarm divided by its spreading rate, was relatively long. (C) The trace after the G/R ratio was converted to osmolarity and time was converted to distance (negative toward the swarm center). The gray dotted line is the reference baseline. It curves down because the evaporation rate was constant but the swarming speed slowed down. The initial swarming speed was 1.0 μm/s. The gray broken line indicates the osmolarity maximum and the double-headed arrow indicates the difference in osmolarity between the maximum and the baseline. The difference between the osmolarity trace and the baseline was maximal between the two gray down-pointing arrows. (D) Enlarged view of the boxed region in C with two other traces aligned on top of it (with swarm speeds in μm/s noted at the right ends of the traces). The small peak at time zero is indicated by gray arrows. The osmolarity maxima are highlighted by gray bars superimposed on the traces.