Revealing Phenotypic Differences in Photodynamic Inactivation. A Fluorescence Microscopy Approach at the Single Bacterium Level

ANDRES M. DURANTINI

Conferencia; CUNY, Brooklyn College chemistry seminar series; 2019

Phenotypic characteristics in bacteria can determinatethe levels of endogenous reactive oxygen species (ROS) being produced as aconsequence of oxidative stress. It can also set the survival pace for eachbacterium against exogenous sources of ROS since these characteristics are frequentlyexpressed in the cell wall composition. Inthe firspart of the presentation, a simple methodology that combines fluorescencemicroscopy with a cell death marker to study the survival times of bacteriaexposed to different photosensitizers will be described. Here, the phenotypicheterogeneity allowing bacteria to persist during photodynamic inactivation (PDI)treatments was evaluated in methicillin resistant Staphylococcus aureus (MRSA) and Escherichia coli as Gram-positive and Gram-negative models,respectively. By means of propidium iodide (PI), we monitored withspatiotemporal resolution cell viability at the single bacterium level.Thorough data analysis of individual cell survival times revealed clearphenotypic variation expressed in the cell wall that affected PI permeation andthus its intercalation with DNA. For the same bacterial sample, death times mayvary from seconds to hours. The second part of the presentation will be focused onthe ongoing work with Prof. Greer during my stay at CUNY, Brooklyn Collegethanks to the Fulbright Program. A new approach to generate airborne singletoxygen in heterogeneous systems with potentialapplication in healthcare willbe discuss.