FTIR micro-spectroscopy using synchrotron-based and thermal source-based radiation for probing live bacteria
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RESEARCH PAPER
FTIR micro-spectroscopy using synchrotron-based and thermal source-based radiation for probing live bacteria Julie Meneghel 1 & Stéphanie Passot 1 & Frédéric Jamme 2 & Stéphane Lefrançois 2 & Pascale Lieben 1 & Paul Dumas 2 & Fernanda Fonseca 1 Received: 3 April 2020 / Revised: 17 July 2020 / Accepted: 21 July 2020 # Springer-Verlag GmbH Germany, part of Springer Nature 2020
Abstract Fourier transform infrared (FTIR) spectroscopy has proven to be a non-invasive tool to analyse cells without the hurdle of employing exogenous dyes or probes. Nevertheless, the study of single live bacteria in their aqueous environment has long remained a big challenge, due to the strong infrared absorption of water and the small size of bacteria compared to the micronrange infrared wavelengths of the probing photons. To record infrared spectra of bacteria in an aqueous environment, at different spatial resolutions, two setups were developed. A custom-built attenuated total reflection inverted microscope was coupled to a synchrotron-based FTIR spectrometer, using a germanium hemisphere. With such a setup, a projected spot size of 1 × 1 μm2 was achieved, which allowed spectral acquisition at the single-cell level in the 1800–1300 cm−1 region. The second setup used a demountable liquid micro-chamber with a thermal source-powered FTIR microscope, in transmission geometry, for probing clusters of a few thousands of live cells in the mid-IR region (4000–975 cm−1). Both setups were applied for studying two strains of a model lactic acid bacterium exhibiting different cryo-resistances. The two approaches allowed the discrimination of both strains and revealed population heterogeneity among bacteria at different spatial resolutions. The multivariate analysis of spectra indicated that the cryo-sensitive cells presented the highest cell heterogeneity and the highest content of proteins with the α-helix structure. Furthermore, the results from clusters of bacterial cells evidenced phosphate and peptidoglycan vibrational bands associated with the cell envelope, as potential markers of resistance to environmental conditions. Keywords Lactic acid bacteria . Freezing . FTIR . Aqueous environment . Population heterogeneity
Introduction Fourier transform infrared (FTIR) micro-spectroscopy is a powerful analytical technique used for characterizing cells and tissues. It is a non-invasive tool since it does not require exogenous probes or contrast agents and it does not induce radiation damage to cells, even if high brightness infrared beam is used [1, 2]. The mid-IR region (4000–400 cm−1) Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00216-020-02835-x) contains supplementary material, which is available to authorized users. * Fernanda Fonseca [email protected] 1
INRAE, AgroParisTech, UMR SayFood, Université Paris-Saclay, 78850 Thiverval-Grignon, France
2
Synchrotron SOLEIL, L’Orme des Merisiers, Saint Aubin, BP 489, 91192 Gif-sur-Yvette, France
provides a non-restrictive chemi
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