SINGLE CELL ANALYSIS OF MAMMALIAN CELLS INFECTED WITH BACILLUS ANTHRACIS USING FLOW CYTOMETRY

Abstract

Host-pathogen interactions are regulated by the virulence factors and host components that can result either in host protection or promote pathogenesis. Historically, significant efforts have been focused on identifying microbe targets for the development of both therapeutic and preventive measures, while the host factors involved in microbial pathogenesis have not been fully explored. For a better understanding of microbial pathogenesis, novel assay techniques that are sensitive, rapid, and can monitor host-pathogen interactions in real time need to be developed. This study investigated the interaction of a gram positive, spore-forming bacteria, Bacillus anthracis, with its host target, a mouse leukemic monocyte macrophage cell line, RAW 264.7. The host-spore interaction was monitored at the single cell level using flow cytometry. Preliminary experiments focused on the use of flow cytometry to identify and characterize individual and mixed populations of spores, vegetative bacteria, and macrophages over time. Further experiments applied this information towards the investigation of early events of infection, such as phagocytosis, and then towards the late events such as B. anthracis induced cell death via flow cytometry. Understanding the interaction of B. anthracis with its host target is necessary for the development of rapid, high-throughput assays with potential application for screening therapeutic small molecules.