Single-cell analysis of the disruption of bacteria with a high-pressure jet device: An application of atomic force microscopy

Abstract

The goal of this study was to reveal the mechanism of bacterial cell disruption by a high-pressure jet device (HPJD) for the reduction of excess sludge produced during wastewater treatment. Escherichia coli and Bacillus subtilis, representative Gram-negative and Gram-positive bacteria respectively, were subjected to HPJD treatment followed by observation of the damaged cells with epifluorescence microscopy and atomic force microscopy (AFM), as well as spectrophotometric quantification of the released intracellular polymeric compounds. The AFM observations demonstrated that the types of cell damage incurred by E. coli and B. subtilis were distinct. B. subtilis showed modest cell disruption comprising partial damage, i.e., the formation of pores and cracks on the cell envelope. In contrast, E. coli showed more extensive damage, i.e., cell cleavage and lysis, in addition to the partial damages observed for B. subtilis. The damaged cells of both bacterial species also exhibited leakage of intercellular compounds such as DNA, proteins and polysaccharides. This loss of intracellular macromolecules resulted in increased cell surface roughness and decreased cell diameter and elasticity, irrespective of the bacterial species tested. The results demonstrated that the degree of cell damage inflicted by HPJD treatment is dependent on the cell wall and membrane structure of the bacterial species. In summary, we propose multiple possible mechanisms of cell disruption resulting from HPJD treatment.