Advanced imaging techniques for assessment of structure, composition and function in biofilm systems

Neu, Thomas R.; Manz, B.; Volke, F.; Dynes, James J.; Hitchcock, Adam P.; Lawrence, John R.

doi:10.1111/j.1574-6941.2010.00837.x

Cited by 195 publications

(137 citation statements)

References 180 publications

(190 reference statements)

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“…For instance, although confocal laser scanning microscopy (CLSM)44 achieves excellent sub‐micrometer resolution and can be applied to a wide field of view,45 it lacks sufficient penetration depth (max. 200 μm in opaque biological samples, but considerably less in chromophore‐containing tissues), thus often not allowing for accurate volumetric analysis 41, 45, 46. Scanning transmission X‐ray microscopy (STXM) and scanning/transmission electron microscopy (SEM/TEM) have been applied to reveal high‐resolution information about sub‐micrometer structure and composition of biofilms, but require fixation or cryogenic preparation of samples 45, 47.…”

Section: Introductionmentioning

confidence: 99%

In situ Biofilm Quantification in Bioelectrochemical Systems by using Optical Coherence Tomography

Molenaar

Sleutels²,

Pereirã³

et al. 2018

ChemSusChem

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Detailed studies of microbial growth in bioelectrochemical systems (BESs) are required for their suitable design and operation. Here, we report the use of optical coherence tomography (OCT) as a tool for in situ and noninvasive quantification of biofilm growth on electrodes (bioanodes). An experimental platform is designed and described in which transparent electrodes are used to allow real‐time, 3D biofilm imaging. The accuracy and precision of the developed method is assessed by relating the OCT results to well‐established standards for biofilm quantification (chemical oxygen demand (COD) and total N content) and show high correspondence to these standards. Biofilm thickness observed by OCT ranged between 3 and 90 μm for experimental durations ranging from 1 to 24 days. This translated to growth yields between 38 and 42 mgCODbiomass gCODacetate −1 at an anode potential of −0.35 V versus Ag/AgCl. Time‐lapse observations of an experimental run performed in duplicate show high reproducibility in obtained microbial growth yield by the developed method. As such, we identify OCT as a powerful tool for conducting in‐depth characterizations of microbial growth dynamics in BESs. Additionally, the presented platform allows concomitant application of this method with various optical and electrochemical techniques.

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Section: Introductionmentioning

confidence: 99%

In situ Biofilm Quantification in Bioelectrochemical Systems by using Optical Coherence Tomography

Molenaar

Sleutels²,

Pereirã³

et al. 2018

ChemSusChem

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“…20 For more comprehensive reviews of the individual techniques, readers are referred elsewhere. [3][4][5][6][7] …”

Section: Introductionmentioning

confidence: 99%

Correlative electron and X-ray microscopy: probing chemistry and bonding with high spatial resolution

et al. 2015

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Two powerful and complementary techniques for chemical characterisation of nanoscale systems are electron energy-loss spectroscopy in the scanning transmission electron microscope, and X-ray absorption spectroscopy in the scanning transmission X-ray microscope. A correlative approach to spectromicroscopy may not only bridge the gaps in spatial and spectral resolution which exist between the two instruments, but also offer unique opportunities for nanoscale characterisation. This review will discuss 10 the similarities of the two spectroscopy techniques and the state of the art for each microscope. Case studies have been selected to illustrate the benefits and limitations of correlative electron and X-ray microscopy techniques. In situ techniques and radiation damage are also discussed. IntroductionGrowth in the development and applications of nanotechnology 15 brings with it an increasing need for characterisation methods capable of providing both structural and chemical analysis on the nanometre scale. Most laboratory based characterisation techniques such as ultra-violet and infra-red spectroscopies, mass spectrometry and thermogravimetric analysis provide spatially 20 averaged information from comparatively large volumes of sample. While these techniques provide a representative overview of the sample, their averaged signals often probe not only the nanomaterial of interest, but also any support material, debris and impurities in heterogeneous structures. In addition, averaged 25 properties miss statistical outliers, which may be critical for the function of the nanomaterial. For characterisation of interfaces or individual nanostructures, techniques with much higher spatial resolutions are essential.There are few available techniques capable of providing 30 chemical information on the nanometre scale. This review will discuss two such methods: electron energy-loss spectroscopy in the scanning transmission electron microscope (STEM-EELS) and X-ray absorption spectroscopy in the scanning transmission X-ray microscope (STXM-XAS). These spectroscopies study the 35 primary processes of inelastic electron scattering or X-ray absorption to obtain chemical information, from chemical speciation analysis to local bonding environments and electronic structure. While the secondary process of X-ray emission also provides chemical information in the electron and X-ray 40 microscopes (by energy dispersive X-ray spectroscopy and X-ray probe X-ray micro-and nano-analysis respectively), these techniques, whilst being very sensitive to low concentrations, are limited to elemental analysis, and they will not be discussed in this review. Although the probe-specimen interactions of EELS 45 and XAS differ (electron scattering and photon absorption, respectively), they provide remarkably similar chemical information. Due to recent advances in diffractive optics for soft (100 eV to 2 keV 1 ) X-rays, as well as monochromators and aberration correctors for electron microscopes, the spatial and 50 spectral resolutions of STXM-XAS...

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“…The inability to answer these and other basic questions stems from a lack of understanding of the cellular-scale architecture of biofilms, which, in turn, highlights a lack of technology capable of tracking the time evolution of biofilms at the single-cell level, in stark contrast to the established tools available for analogous studies of development in eukaryotic organisms (11). Despite tremendous progress in imaging biofilms (12), bottlenecks remain due to resolution and phototoxicity issues. Thus, our understanding of biofilms primarily comes from high-resolution optical images of immature biofilms that are only a few cell layers thick (13), low-resolution images of the 3D contour (12), or electron microscopy images of processed samples (14), which lack the key spatiotemporal information required to understand the biofilm developmental process at the level of the basic unit: the individual cell.…”

mentioning

confidence: 99%

“…Despite tremendous progress in imaging biofilms (12), bottlenecks remain due to resolution and phototoxicity issues. Thus, our understanding of biofilms primarily comes from high-resolution optical images of immature biofilms that are only a few cell layers thick (13), low-resolution images of the 3D contour (12), or electron microscopy images of processed samples (14), which lack the key spatiotemporal information required to understand the biofilm developmental process at the level of the basic unit: the individual cell.…”

mentioning

confidence: 99%

Vibrio cholerae biofilm growth program and architecture revealed by single-cell live imaging

Yan

Sharo

Stone

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

158

209

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Biofilms are surface-associated bacterial communities that are crucial in nature and during infection. Despite extensive work to identify biofilm components and to discover how they are regulated, little is known about biofilm structure at the level of individual cells. Here, we use state-of-the-art microscopy techniques to enable live singlecell resolution imaging of a Vibrio cholerae biofilm as it develops from one single founder cell to a mature biofilm of 10,000 cells, and to discover the forces underpinning the architectural evolution. Mutagenesis, matrix labeling, and simulations demonstrate that surface adhesion-mediated compression causes V. cholerae biofilms to transition from a 2D branched morphology to a dense, ordered 3D cluster. We discover that directional proliferation of rod-shaped bacteria plays a dominant role in shaping the biofilm architecture in V. cholerae biofilms, and this growth pattern is controlled by a single gene, rbmA. Competition analyses reveal that the dense growth mode has the advantage of providing the biofilm with superior mechanical properties. Our single-cell technology can broadly link genes to biofilm fine structure and provides a route to assessing cell-to-cell heterogeneity in response to external stimuli. biofilm | single cell | self-organization | community | biomechanics

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Advanced imaging techniques for assessment of structure, composition and function in biofilm systems

Cited by 195 publications

References 180 publications

In situ Biofilm Quantification in Bioelectrochemical Systems by using Optical Coherence Tomography

In situ Biofilm Quantification in Bioelectrochemical Systems by using Optical Coherence Tomography

Correlative electron and X-ray microscopy: probing chemistry and bonding with high spatial resolution

Vibrio cholerae biofilm growth program and architecture revealed by single-cell live imaging

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