The quantification of the extent of engulfment was performed as follows: for each colony, to calculate the percentage or the disc circumference covered by the biofilm cells, we measured the intensity of brightfield image at seven points, evenly spaced around the disc. The points were consistent for all the colonies analyzed, with minimal manual adjustments to compensate for false positive results resulting from the slightly irregular shape of the disc, when needed. We also measured the intensity of the opposite side of the colony as a positive control and the intensity of the agar plate next to the disc as background. After subtraction of background, a threshold level was unbiasedly calculated from the positive control measurement. Each measurement was scored as either negative (below the threshold) or positive (above the threshold). Last, for each colony, the fraction of positive measurements out of all measurement was calculated. The results presented are the distribution of measurements for at least 12 colonies for each strain and time point, over three independent experiments. Data were analyzed using AxioVision suite software (Zeiss).

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