Lei Shi

Among pathogenic bacteria, Escherichia coli, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa were the six leading causes for the deaths associated with antibiotic resistance in 2019. Although new treatment options are urgently needed, the precise identification of the bacterial species remains pivotal for an accurate diagnosis and effective treatment. Clinically, mass spectrometry is used to distinguish these bacteria based on their protein mass pattern at the genus and species level. Herein, we report an alternative approach to identify these bacteria using the nitroreductase-based "turn-on" fluorescent probes (ETH1-NO₂ and ETH2-NO₂), with potential visual indicators for the six individual bacteria species. The limits of detection (LODs) of the probes for NTRs are 0.562 (ETH1-NO₂) and 0.153 μg/mL (ETH2-NO₂), respectively. They respond effectively to both Gram-positive and Gram-negative bacteria, with the lowest LOD at 1.2 × 10⁶ CFU/mL for E. coli. In particular, different bacteria show noticeable difference in the apparent color of ETH1-NO₂ samples, allowing possible identification of these bacteria visually. In addition, ETH1-NO₂ also has potential applications in bacterial fluorescence imaging. Thus, our study provides an alternative approach for bacteria identification and new reagents for bacteria imaging.

Viscosity, at the subcellular level, plays a crucial role as a physicochemical factor affecting microenvironment homeostasis. Abnormal changes in mitochondrial viscosity often lead to various diseases in the organism. Based on the twisted intramolecular charge transfer mechanism, four hemicyanine dye fluorescent probes (HT-SA, HT-SA-S, HT-Bzh, and HT-NA) were designed and synthesized for viscosity response. The single bond between the nitrogen-containing heterocycle and the carbon–carbon double in the structure of the probe bond served as the viscosity response site. Finally, the probe HT-Bzh was screened as the optimal mitochondrial viscosity probe according to its responsiveness, targeting, and interference resistance. The fluorescence intensity of the probe HT-Bzh increased 22-fold when the viscosity was increased from 13.75 to 811.2 cP. In summary, all four viscosity probes we have developed can be used in different applications depending on the external environment, providing a valuable reference for the design of potential tools to address viscosity monitoring in biological systems.

The fluorescence of F-0 is susceptible to quenching by Cu²⁺, but can be restored upon GLY complexation with Cu²⁺.

SO₂, a gas signaling molecule, can be produced endogenously in mitochondria. Its hydrolysate, HSO₃^-, plays a key role in food preservation, cardiovascular relaxation, and other fields, suggesting that it is important to achieve its detection. Here, based on the Michael addition mechanism, four hemicyanine dye fluorescent probes (ETN, ETB, STB, and EIB) were designed and synthesized for responding to HSO₃^-. We evaluated the reaction ability of different probes with HSO₃^- and tried to explain the reasons for the significantly different response effects between probes and HSO₃^- according to the structure-activity relationship. The influence of different substituents of probes on the properties of mitochondria-targeting was also discussed. Finally, we screened out ETN as the optimal HSO₃^- probe due to its high sensitivity, rapid reactivity, and good mitochondria-targeting, and it could sensitively respond to HSO₃^- in living cells. The LODs of ETN for HSO₃^- were calculated by both absorption and fluorescence methods, respectively, which were 2.727 and 0.823 μM. Our work provided valuable references for designing strategies and potential tools for response to SO₂ derivatives in biosystems.

A mitochondria-targeted fluorescent probe based on hemicyanine showed a dual-response for viscosity and HSO₃⁻in living cells.