Nitroreductases (NTRs)
mediate the reduction of nitroaromatic compounds
to the corresponding nitrite, hydroxylamine, or amino derivatives.
The activity of NTRs in bacteria facilitates the metabolic activation
and antibacterial activity of 5-nitroimidazoles. Therefore, NTR activity
correlates with the drug susceptibility and resistance of pathogenic
bacteria. As such, it is important to develop a rapid and visual assay
for the real-time sensing of bacterial NTRs for the evaluation and
development of antibiotics. Herein, an activatable near-infrared fluorescent
probe (
HC–NO
2
) derived
from a hemicyanine fluorophore was designed and developed based on
two evaluation factors, including the calculated partition coefficient
(Clog
P
) and fluorescence wavelength. Using
HC–NO
2
as the special substrate
of NTRs, NTR activity can be assayed efficiently, and then, bacteria
can be imaged based on the detection of NTRs. More importantly, a
sensitive in-gel assay using
HC–NO
2
has been developed to selectively identify NTRs and sensitively
determine NTR activity. Using the in-gel assay, NTRs from various
bacterial species have been profiled visually from the “fluorescence
fingerprints”, which facilitates the rapid identification of
NTRs from bacterial lysates. Thus, various homologous NTRs were identified
from three metronidazole-susceptible bacterial species as well as
seven unsusceptible species, which were confirmed by the whole-genome
sequence. As such, the evaluation of NTRs from different bacterial
species should help improve the rational usage of 5-nitroimidazole
drugs as antibiotics.