Fluorescence microscopy has illuminated countless discoveries in biology, but its utility has been restricted in uncovering live-cell biological phenomena in oxygen-free or oxygen-limited environments. Fluorescent proteins like GFP depend on oxygen to produce fluorescence and cannot be used to study anaerobic systems like the gut microbiome. Here, we demonstrate the use of the fatty-acid-binding fluorescent protein UnaG to implement live-cell imaging of the obligate anaerobe gut bacterium Bacteroides thetatiotaomicron. UnaG binds a fluorogenic ligand, which enables fluorescent labeling in live-cell environments with diminished background noise. We have evaluated cytosolic and membrane UnaG labeling in mono-cultures of B. thetatiotaomicron and in mixed-species cultures of B. thetatiotaomicron and Ruminococcus bromii. We have also assessed the versatility of UnaG fluorescence by screening other fluorogenic ligands to look for novel pairings with UnaG that produce different color labeling.

Using a fatty-acid-binding fluorescent-protein for oxygen-independent live-cell imaging of microbiomes

2019 Fall National Meeting

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