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Mechanically Resolved Imaging Of Bacteria Using Expansion Microscopy

Youngbin Lim, Margarita Khariton, Keara M. Lane, Anthony L. Shiver, Katharine M. Ng, Samuel R. Bray, Jian Qin, Kerwyn Casey Huang, Bo Wang

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AbstractImaging dense and diverse microbial communities has broad applications in basic microbiology and medicine, but remains a grand challenge due to the fact that many species adopt similar morphologies. While prior studies have relied on techniques involving spectral labeling, we have developed an expansion microscopy method (μExM) in which cells are physically expanded prior imaging and their expansion patterns depend on the structural and mechanical properties of their cell walls, which vary across species and conditions. We use this phenomenon as a quantitative and sensitive phenotypic imaging contrast orthogonal to spectral separation in order to resolve bacterial cells of different species or in distinct physiological states. Focusing on host-microbe interactions that are difficult to quantify through fluorescence alone, we demonstrate the ability of μExM to distinguish species within a dense community throughin vivoimaging of a model gut microbiota, and to sensitively detect cell-envelope damage caused by antibiotics or previously unrecognized cell-to-cell phenotypic heterogeneity among pathogenic bacteria as they infect macrophages.