Single-Cell Real-Time Visualization and Quantification of Perylene Bioaccumulation in Microorganisms

Bioaccumulation of perylene in Escherichia coli and Staphylococcus aureus was visualized and quantified
in real time with high sensitivity at high temporal resolution. For
the first time, single-molecule fluorescence microscopy (SMFM) with
a microfluidic flow chamber and temperature control has enabled us
to record the dynamic process of perylene bioaccumulation in single
bacterial cells and examine the cell-to-cell heterogeneity. Although
with identical genomes, individual E. coli cells
exhibited a high degree of heterogeneity in perylene accumulation
dynamics, as shown by the high coefficient of variation (C.V = 1.40).
This remarkable heterogeneity was exhibited only in live E.
coli cells. However, the bioaccumulation of perylene in live
and dead S. aureus cells showed similar patterns
with a low degree of heterogeneity (C.V = 0.36). We found that the
efflux systems associated with Tol C played an essential role in perylene
bioaccumulation in E. coli, which caused a significantly
lower accumulation and a high cell-to-cell heterogeneity. In comparison
with E. coli, the Gram-positive bacteria S. aureus lacked an efficient efflux system against perylene.
Therefore, perylene bioaccumulation in S. aureus was
simply a passive diffusion process across the cell membrane.