title: Microspectrometric insights on the uptake of antibiotics at the single bacterial cell level.
abstract: Bacterial multidrug resistance is a significant health issue. A key challenge, particularly in Gram-negative antibacterial research, is to better understand membrane permeation of antibiotics in clinically relevant bacterial pathogens. Passing through the membrane barrier to reach the required concentration inside the bacterium is a pivotal step for most antibacterials. Spectrometric methodology has been developed to detect drugs inside bacteria and recent studies have focused on bacterial cell imaging. Ultimately, we seek to use this method to identify pharmacophoric groups which improve penetration, and therefore accumulation, of small-molecule antibiotics inside bacteria. We developed a method to quantify the time scale of antibiotic accumulation in living bacterial cells. Tunable ultraviolet excitation provided by DISCO beamline (synchrotron Soleil) combined with microscopy allows spectroscopic analysis of the antibiotic signal in individual bacterial cells. Robust controls and measurement of the crosstalk between fluorescence channels can provide real time quantification of drug. This technique represents a new method to assay drug translocation inside the cell and therefore incorporate rational drug design to impact antibiotic uptake.
authors.affiliation: DISCO beamline, Synchrotron Soleil, Saint-Aubin, France.
authors.forename: Bertrand
authors.initials: B
authors.lastname: Cinquin
authors.affiliation: UMR-MD1, Aix-Marseille Université, IRBA, Marseille, France.
authors.forename: Laure
authors.initials: L
authors.lastname: Maigre
authors.affiliation: UMR-MD1, Aix-Marseille Université, IRBA, Marseille, France.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Pinet
authors.affiliation: UMR-MD1, Aix-Marseille Université, IRBA, Marseille, France.
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Chevalier
authors.affiliation: Antibacterial DPU, GlaxoSmithKline, Collegeville PA, USA.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Stavenger
authors.affiliation: Infection Bioscience, AstraZeneca R&D Boston, Waltham, Massachusetts, USA.
authors.forename: Scott
authors.initials: S
authors.lastname: Mills
authors.affiliation: DISCO beamline, Synchrotron Soleil, Saint-Aubin, France.
authors.forename: Matthieu
authors.initials: M
authors.lastname: Réfrégiers
authors.affiliation: UMR-MD1, Aix-Marseille Université, IRBA, Marseille, France.
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Pagès
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D001419:Bacteria / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D024881:Drug Resistance, Bacterial; D015591:Microspectrophotometry; D015394:Molecular Structure; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species; D059010:Single-Cell Analysis / Q000379:methods*
chemical_list: D000900:Anti-Bacterial Agents; D017382:Reactive Oxygen Species