Article

Force volume and stiffness tomography investigation on the dynamics of stiff material under bacterial membranes

Laboratory of Physics of Living Matter, EPFL, Lausanne, Switzerland.
Journal of Molecular Recognition (Impact Factor: 2.34). 05/2012; 25(5):278-84. DOI: 10.1002/jmr.2171
Source: PubMed

ABSTRACT The determination of the characteristics of micro-organisms in clinical specimens is essential for the rapid diagnosis and treatment of infections. A thorough investigation of the nanoscale properties of bacteria can prove to be a fundamental tool. Indeed, in the latest years, the importance of high resolution analysis of the properties of microbial cell surfaces has been increasingly recognized. Among the techniques available to observe at high resolution specific properties of microscopic samples, the Atomic Force Microscope (AFM) is the most widely used instrument capable to perform morphological and mechanical characterizations of living biological systems. Indeed, AFM can routinely study single cells in physiological conditions and can determine their mechanical properties with a nanometric resolution. Such analyses, coupled with high resolution investigation of their morphological properties, are increasingly used to characterize the state of single cells. In this work, we exploit the capabilities and peculiarities of AFM to analyze the mechanical properties of Escherichia coli in order to evidence with a high spatial resolution the mechanical properties of its structure. In particular, we will show that the bacterial membrane is not mechanically uniform, but contains stiffer areas. The force volume investigations presented in this work evidence for the first time the presence and dynamics of such structures. Such information is also coupled with a novel stiffness tomography technique, suggesting the presence of stiffer structures present underneath the membrane layer that could be associated with bacterial nucleoids.

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Available from: Giovanni Longo, Aug 05, 2014
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    • "Any images of this microorganism are therefore impossible to record. Recent works by Longo et al. (2012) present nanoindentation images of E. coli cells. This is also a way to image this bacterial species, however, the lateral resolution shown by the authors is of 32 px 2 , which is very low and does not allow a detailed observation of the bacterial cell wall. "
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    ABSTRACT: Since the last 10 years, AFM has become a powerful tool to study biological samples. However, the classical modes offered (imaging or tapping mode) often damage sample that are too soft or loosely immobilized. If imaging and mechanical properties are required, it requests long recording time as two different experiments must be conducted independently. In this study we compare the new QI™ mode against contact imaging mode and force volume mode, and we point out its benefit in the new challenges in biology on six different models: Escherichia coli, Candida albicans, Aspergillus fumigatus, Chinese hamster ovary cells and their isolated nuclei, and human colorectal tumor cells.
    Micron 02/2013; DOI:10.1016/j.micron.2013.02.003 · 2.06 Impact Factor
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    • "Any images of this microorganism are therefore impossible to record. Recent works by Longo et al. (2012) present nanoindentation images of E. coli cells. This is also a way to image this bacterial species, however, the lateral resolution shown by the authors is of 32 px 2 , which is very low and does not allow a detailed observation of the bacterial cell wall. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Since the last 10 years, AFM has become a powerful tool to study biological samples. However, the classical modes offered (imaging or tapping mode) often damage sample that are too soft or loosely immobilized. If imaging and mechanical properties are required, it requests long recording time as two different experiments must be conducted independently. In this study we compare the new QI™ mode against contact imaging mode and force volume mode, and we point out its benefit in the new challenges in biology on six different models: E. coli, C. albicans, A. fumigatus, Chinese Hamster Ovary cells and their isolated nuclei, and Human Colorectal Tumor cells.
    Micron 02/2013; · 2.06 Impact Factor
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    • "These features were present on all the analyzed bacteria, and their complex structure is evidenced in the higher resolution, 1 × 1 μm, images (Fig. 4B and D, zooms of the higher and lower structures respectively). A complete characterization of such structures is still underway, but the most probable origin of such features is the presence of agglomerates of stiffer internal cell components (mainly DNA), inside the cytoplasm (nucleoids) (Frenkiel-Krispin et al., 2004; Longo et al., 2012). To better characterize such areas, force tomography analyses were performed. "
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    ABSTRACT: In the latest years the importance of high resolution analysis of the microbial cell surface has been increasingly recognized. Indeed, in order to better understand bacterial physiology and achieve rapid diagnostic and treatment techniques, a thorough investigation of the surface modifications induced on bacteria by different environmental conditions or drugs is essential. Several instruments are nowadays available to observe at high resolution specific properties of microscopic samples. Among these, AFM can routinely study single cells in physiological conditions, measuring the mechanical properties of their membrane at a nanometric scale (force volume). Such analyses, coupled with high resolution investigation of their morphological properties, are increasingly used to characterize the state of single cells. In this work we exploit such technique to characterize bacterial systems. We have performed an analysis of the mechanical properties of bacteria (Escherichia coli) exposed to different conditions. Such measurements were performed on living bacteria, by changing in real-time the liquid environment: standard phosphate buffered saline, antibiotic (ampicillin) in PBS and growth medium. In particular we have focused on the determination of the membrane stiffness modifications induced by these solutions, in particular between stationary and replicating phases and what is the effect of the antibiotic on the bacterial structure.
    Journal of microbiological methods 02/2013; 93(2). DOI:10.1016/j.mimet.2013.01.022 · 2.10 Impact Factor