Strongly enhanced bacterial bioluminescence with the <i>ilux</i> operon for single-cell imaging.

Abstract

Bioluminescence imaging of single cells is often complicated by the requirement of exogenous luciferins that can be poorly cell-permeable or produce high background signal. Bacterial bioluminescence is unique in that it uses reduced flavin mononucleotide as a luciferin, which is abundant in all cells, making this system purely genetically encodable by the <i>lux</i> operon. Unfortunately, the use of bacterial bioluminescence has been limited by its low brightness compared with other luciferases. Here, we report the generation of an improved <i>lux</i> operon named <i>ilux</i> with an approximately sevenfold increased brightness when expressed in <i>Escherichia coli</i>; <i>ilux</i> can be used to image single <i>E. coli</i> cells with enhanced spatiotemporal resolution over several days. In addition, since only metabolically active cells produce bioluminescent signal, we show that <i>ilux</i> can be used to observe the effect of different antibiotics on cell viability on the single-cell level.