Quantifying the fractal complexity of nutrient transport channels in biofilms under varying cell shape and growth environment.

Recent mesoscopic characterization of nutrient-transporting channels in has allowed the identification and measurement of individual channels in whole mature colony biofilms. However, their complexity under different physiological and environmental conditions remains unknown. Analysis of confocal micrographs of colony biofilms formed by cell shape mutants of shows that channels have high fractal complexity, regardless of cell phenotype or growth medium. In particular, colony biofilms formed by the mutant strain Δ, which has a wide-cell phenotype, have a higher fractal dimension when grown on rich medium than when grown on minimal medium, with channel complexity affected by glucose and agar concentrations in the medium. Osmotic stress leads to a dramatic reduction in the Δ cell size but has a limited effect on channel morphology. This work shows that fractal image analysis is a powerful tool to quantify the effect of phenotypic mutations and growth environment on the morphological complexity of internal biofilm structures. If applied to a wider range of mutant strains, this approach could help elucidate the genetic determinants of channel formation in colony biofilms.