An image processing-based quantification of gram variability in Acinetobacter baumannii

: Gram staining differentiates bacteria as gram-positive and gram-negative, depending on their cell wall constituents, and coloring cells in violet and pink, respectively. Sometimes, a subpopulation of the same bacterial species assumes different colors, ranging from pink to violet, for reasons that are not completely understood yet. We analyze conventional brightfield images and use an automated pipeline to count pink and violet cells. The ImageJ-based processing algorithm quantifies the gram variability in Acinetobacter baumannii ACICU in the stationary phase of growth with a percentage of 66% pink cells. Different bacterial strains and cell growth stages have been considered. RESEARCH HIGHLIGHTS: Gram staining differentiates bacteria into gram-positive (violet) and gram-negative (pink). Gram variability represents an inhomogeneous distribution of pink and violet cells within the same species. We developed an ImageJ-based pipeline for the quantification of Gram variability in Acinetobacter baumannii.