Cryo-Scanning Electron Microscopy (SEM) and Scanning Transmission Electron Microscopy (STEM)-in-SEM for Bio- and Organo-Mineral Interface Characterization in the Environment
Résumé
Understanding biofilm interactions with surrounding substratum and pollutants/particles can benefit
from the application of existing microscopy tools. Using the example of biofilm interactions with zero-valent iron
nanoparticles (nZVI), this study aims to apply various approaches in biofilm preparation and labeling for
fluorescent or electron microscopy and energy dispersive X-ray spectrometry (EDS) microanalysis for accurate
observations. According to the targeted microscopy method, biofilms were sampled as flocs or attached biofilm,
submitted to labeling using 4’,6-diamidino-2-phenylindol, lectins PNA and ConA coupled to fluorescent dye or
gold nanoparticles, and prepared for observation (fixation, cross-section, freezing, ultramicrotomy). Fluorescent
microscopy revealed that nZVI were embedded in the biofilm structure as aggregates but the resolution was
insufficient to observe individual nZVI. Cryo-scanning electron microscopy (SEM) observations showed nZVI
aggregates close to bacteria, but it was not possible to confirm direct interactions between nZVI and cell
membranes. Scanning transmission electron microscopy in the SEM (STEM-in-SEM) showed that nZVI
aggregates could enter the biofilm to a depth of 7–11 μm. Bacteria were surrounded by a ring of extracellular
polymeric substances (EPS) preventing direct nZVI/membrane interactions. STEM/EDS mapping revealed a
co-localization of nZVI aggregates with lectins suggesting a potential role of EPS in nZVI embedding. Thus, the
combination of divergent microscopy approaches is a good approach to better understand and characterize
biofilm/metal interactions.
Domaines
Matériaux
Origine : Fichiers produits par l'(les) auteur(s)
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