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Three-Dimensional Morphology Quantification of Biofilm Structures from Confocal Laser Scanning Microscopy Images

Staphylococcus aureus is an opportunistic human pathogen and a causative agent of a wide range of diseases. Its hallmark feature is a capability to form biological structures called biofilms, in which bacteria can embed themselves in a protective matrix. The study of biofilm structures and morphologies is central to understand the biological characteristics of the biofilm including its resistance to drugs. These studies, however, have focused on two-dimensional (2D) analysis of the biofilm images. In this paper, we show the importance of three-dimensional analysis of the biofilm morphology by taking into account the temporal information between the images. Specifically, we show that the morphological features computed from a simple gradient-based three-dimensional segmentation method outperforms not only its 2D homologue but also sophisticated two-dimensional segmentation methods like the watershed algorithm. Our analysis is applied to real biofilm confocal laser scanning microsopy images with a ground truth delineated manually by expert biologists at the University of Arkansas for Medical Sciences.