Red-Shifted Environmental Fluorophores and Their Use for the Detection of Gram-Negative Bacteria

Abstract

Two novel, water-soluble, merocyanine fluorophores were readily prepared by microwave-assisted synthesis. Full optical characterization was performed in a series of protic and aprotic solvents, and the dyes displayed fluorescence in the red region with up to a 20-fold decrease in brightness in water, demonstrating a strong environmental sensitivity hereby termed as solvato-fluorogenicity (to distinguish from solvatochromism). Shorter fluorescent lifetimes were also measured in water, which confirmed this character. These dyes were conjugated to a modified polymyxin scaffold that allowed fluorescence “switch-on” upon binding to Gram-negative bacterial membranes, and selective fluorescence detection of bacteria in a wash-free protocol.

Rapid detection of major Gram-positive pathogens in ocular specimens using a novel fluorescent vancomycin-based probe

“…By determining  and  f values for each PBS/DMSO mixture (Table S1), we found a progressive, 22fold, increase in brightness ( f ) in pure DMSO compared to aqueous buffers with negligible variations in excitation and emission wavelengths. This solvato-fluorogenic character is the key to labelling, selectively, pathogen membranes/cell walls without the need for washing steps, 16 as the low fluorescence in aqueous media reduces background signal and "lights-up" the bacteria. The probe showed good labelling of bacteria down to 0.1 µM for the target microbe (Figure 2), which is lower than the reported minimum inhibitory concentration (MIC) of parent vancomycin compound.…”

“…Several strategies have been used to develop Gram-specific fluorescent probes, [4][5][6][7] including the development of positively charged dyes 8 or aggregation-induced emission systems, [9][10][11][12] or the direct covalent conjugation of known fluorophores to pathogen-binding ligands. [13][14][15][16] In this exploratory study, we designed and evaluated a fluorescent probe for the rapid detection of Gram-positive bacteria in ocular specimens in comparison with the conventional Gram stain. 17,18 The fluorescent probe developed (a vancomycin-NBD conjugate here called Van-Green) discriminated Gram-positive bacteria from other bacteria with excellent selectivity and vitally did not require any modification to the readily available microscope systems widely used in the laboratory.…”

“…17,18 The fluorescent probe developed (a vancomycin-NBD conjugate here called Van-Green) discriminated Gram-positive bacteria from other bacteria with excellent selectivity and vitally did not require any modification to the readily available microscope systems widely used in the laboratory. The optical molecular imaging agent used here, Van-Green, was designed with a solvato-fluorogenic dye 16 conjugated to the clinically approved glycopeptide antibiotic vancomycin. The design of the probe imitates the structure of oritavancin, a semi-synthetic vancomycin derivative in which the vancosamine moiety (i.e.…”

Applications of Machine Learning in time-domain Fluorescence Lifetime Imaging: a Review

“…Such capabilities underpin critical applications like cancer analysis and drug-target engagement [119][120][121]. Examples include, segmenting, detecting, and tracking microscopic images of viruses (e.g., SARS-CoV and its variants [122]), bacteria like E. coli and Staphylococcus aureus [123], as well as various types of cells [124]. Essentially, the aim is to identify and quantify objects of interest to provide an enriched image semantic analysis.…”

“…In clinical microscopy imaging, precise object recognition can be intricate due to problem-specific factors. For instance, detecting specific bacteria [ 123 ] is often subject to expert opinion, making it more challenging than problems with clear visual cues. In the field of fluorescence imaging, many researchers have utilized computer vision techniques specifically for steady-state systems, as indicated by [ 135 ].…”