Antibiotic transport in resistant bacteria: synchrotron UV fluorescence microscopy to determine antibiotic accumulation with single cell resolution.
A molecular definition of the mechanism conferring bacterial multidrug resistance is clinically crucial and today methods for quantitative determination of the uptake of antimicrobial agents with single cell resolution are missing. Using the naturally occurring fluorescence of antibacterial agents a...
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2012-01-01
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doaj-6c7260a8978e4a01b3d3dfb39b8398092021-03-03T20:28:47ZengPublic Library of Science (PLoS)PLoS ONE1932-62032012-01-0176e3862410.1371/journal.pone.0038624Antibiotic transport in resistant bacteria: synchrotron UV fluorescence microscopy to determine antibiotic accumulation with single cell resolution.Slávka KaščákováLaure MaigreJacqueline ChevalierMatthieu RéfrégiersJean-Marie PagèsA molecular definition of the mechanism conferring bacterial multidrug resistance is clinically crucial and today methods for quantitative determination of the uptake of antimicrobial agents with single cell resolution are missing. Using the naturally occurring fluorescence of antibacterial agents after deep ultraviolet (DUV) excitation, we developed a method to non-invasively monitor the quinolones uptake in single bacteria. Our approach is based on a DUV fluorescence microscope coupled to a synchrotron beamline providing tuneable excitation from 200 to 600 nm. A full spectrum was acquired at each pixel of the image, to study the DUV excited fluorescence emitted from quinolones within single bacteria. Measuring spectra allowed us to separate the antibiotic fluorescence from the autofluorescence contribution. By performing spectroscopic analysis, the quantification of the antibiotic signal was possible. To our knowledge, this is the first time that the intracellular accumulation of a clinical antibiotic could be determined and discussed in relation with the level of drug susceptibility for a multiresistant strain. This method is especially important to follow the behavior of quinolone molecules at individual cell level, to quantify the intracellular concentration of the antibiotic and develop new strategies to combat the dissemination of MDR-bacteria. In addition, this original approach also indicates the heterogeneity of bacterial population when the same strain is under environmental stress like antibiotic attack.https://www.ncbi.nlm.nih.gov/pmc/articles/pmid/22719907/?tool=EBI |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Slávka Kaščáková Laure Maigre Jacqueline Chevalier Matthieu Réfrégiers Jean-Marie Pagès |
spellingShingle |
Slávka Kaščáková Laure Maigre Jacqueline Chevalier Matthieu Réfrégiers Jean-Marie Pagès Antibiotic transport in resistant bacteria: synchrotron UV fluorescence microscopy to determine antibiotic accumulation with single cell resolution. PLoS ONE |
author_facet |
Slávka Kaščáková Laure Maigre Jacqueline Chevalier Matthieu Réfrégiers Jean-Marie Pagès |
author_sort |
Slávka Kaščáková |
title |
Antibiotic transport in resistant bacteria: synchrotron UV fluorescence microscopy to determine antibiotic accumulation with single cell resolution. |
title_short |
Antibiotic transport in resistant bacteria: synchrotron UV fluorescence microscopy to determine antibiotic accumulation with single cell resolution. |
title_full |
Antibiotic transport in resistant bacteria: synchrotron UV fluorescence microscopy to determine antibiotic accumulation with single cell resolution. |
title_fullStr |
Antibiotic transport in resistant bacteria: synchrotron UV fluorescence microscopy to determine antibiotic accumulation with single cell resolution. |
title_full_unstemmed |
Antibiotic transport in resistant bacteria: synchrotron UV fluorescence microscopy to determine antibiotic accumulation with single cell resolution. |
title_sort |
antibiotic transport in resistant bacteria: synchrotron uv fluorescence microscopy to determine antibiotic accumulation with single cell resolution. |
publisher |
Public Library of Science (PLoS) |
series |
PLoS ONE |
issn |
1932-6203 |
publishDate |
2012-01-01 |
description |
A molecular definition of the mechanism conferring bacterial multidrug resistance is clinically crucial and today methods for quantitative determination of the uptake of antimicrobial agents with single cell resolution are missing. Using the naturally occurring fluorescence of antibacterial agents after deep ultraviolet (DUV) excitation, we developed a method to non-invasively monitor the quinolones uptake in single bacteria. Our approach is based on a DUV fluorescence microscope coupled to a synchrotron beamline providing tuneable excitation from 200 to 600 nm. A full spectrum was acquired at each pixel of the image, to study the DUV excited fluorescence emitted from quinolones within single bacteria. Measuring spectra allowed us to separate the antibiotic fluorescence from the autofluorescence contribution. By performing spectroscopic analysis, the quantification of the antibiotic signal was possible. To our knowledge, this is the first time that the intracellular accumulation of a clinical antibiotic could be determined and discussed in relation with the level of drug susceptibility for a multiresistant strain. This method is especially important to follow the behavior of quinolone molecules at individual cell level, to quantify the intracellular concentration of the antibiotic and develop new strategies to combat the dissemination of MDR-bacteria. In addition, this original approach also indicates the heterogeneity of bacterial population when the same strain is under environmental stress like antibiotic attack. |
url |
https://www.ncbi.nlm.nih.gov/pmc/articles/pmid/22719907/?tool=EBI |
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