IADR Abstract Archives
Bacillus subtilis Biofilm Development – Computerized Study of Morphology Across Phenotypes
Objectives: Biofilms are commonly defined as accumulations of microbes, embedded in self-secreted extra-cellular matrix, on solid surfaces or liquid interfaces. Bacillus subtilis is widely used in research and is considered to be a facile model organism for the study of biofilms, particularly due to its ability to form distinctly segmented three-dimensional colony biofilms. This study investigates several aspects of B. subtilis biofilm formation using tools from the field of image processing. Specifically, a computerized approach is employed to characterize morphological features of B. subtilis colony type biofilms across multiple phenotypes.
Methods: GFP-labelled B. subtilis cells were cultured on Lysogeny broth agars, with or without supplementation with glycerol and manganese (biofilm-promoting combination). Microscopic images of developing colonies were analyzed using confocal microscopy with a series of staining methods. An algorithmic approach was then utilized to identify and investigate two distinct regions within the biofilm, a "core" region at the colony center and an "expanding" region on the periphery. We developed a methodology by which we convert color images of B. subtilis core regions into one-dimensional signals which are then used to mathematically describe the various phenotypes of the macrocolonies.
Results: Our results demonstrate that mature biofilm of B. subtilis grown on biofilm-promoting medium, unlike that grown on un-supplemented Lysogeny broth agar, is characterized by a uniquely developing complex structure observed at its core. As the biofilm develops, the core region undergoes specific patterns of cellular differentiation that separate it from the expanding region that is located further from the colony center, the colony “periphery”. By applying our methodology, we are able to map the phenotypic variances of B. subtilis biofilms cultured in different growth conditions.
Conclusions: We propose a new non-disruptive computational approach for image-based analysis of B. subtilis biofilms - such an approach represents a unique classification method of various colony type biofilm phenotypes.
Methods: GFP-labelled B. subtilis cells were cultured on Lysogeny broth agars, with or without supplementation with glycerol and manganese (biofilm-promoting combination). Microscopic images of developing colonies were analyzed using confocal microscopy with a series of staining methods. An algorithmic approach was then utilized to identify and investigate two distinct regions within the biofilm, a "core" region at the colony center and an "expanding" region on the periphery. We developed a methodology by which we convert color images of B. subtilis core regions into one-dimensional signals which are then used to mathematically describe the various phenotypes of the macrocolonies.
Results: Our results demonstrate that mature biofilm of B. subtilis grown on biofilm-promoting medium, unlike that grown on un-supplemented Lysogeny broth agar, is characterized by a uniquely developing complex structure observed at its core. As the biofilm develops, the core region undergoes specific patterns of cellular differentiation that separate it from the expanding region that is located further from the colony center, the colony “periphery”. By applying our methodology, we are able to map the phenotypic variances of B. subtilis biofilms cultured in different growth conditions.
Conclusions: We propose a new non-disruptive computational approach for image-based analysis of B. subtilis biofilms - such an approach represents a unique classification method of various colony type biofilm phenotypes.