Antimicrobial resistance (AMR) is a rapidly evolving threat to human health, predicted to cause 10 million deaths annually by 2050 [1]. A sparse clinical pipeline mainly populated by new versions of existing antibiotics emphasises the need for novel therapeutic strategies. However, the development of new antibiotics is hindered by the lack of understanding of how existing antibiotics actually work, despite decades of use.
Over the past decade we have been developing fluorescent probes derived from all major classes of antibiotics. We attach the small fluorophores nitrobenzoxadiazole (NBD: green) and dimethylaminocoumarin-2-acetic acid (DMACA: blue) via Cu-catalyzed azide-alkyne cycloaddition reaction to azide-functionalised antibiotics, with the site of derivatisation selected so the adducts retain activity. We have previously reported derivatives of vancomycin, trimethoprim, roxithromycin, linezolid and ciprofloxacin, and now describe probes of the peptide antibiotics polymyxin B, octapeptin C4, tachyplesin, arenicin-3 and daptomycin, as well as several classes of lactams. Notably, most derivatised antibiotics retained similar activity and resistance profiles as the parent antibiotic, confirming their utility for investigations of antibiotic-bacteria interactions.
High resolution microscopy of bacteria labelled with these probes demonstrated strikingly different localisation patterns between the peptide antibiotics, despite all nominally working via membrane disruption. The polymyxin and daptomycin probes are able to quickly distinguish between bacteria with varying levels of resistance, providing a useful rapid assay to assess bacterial sensitivity. Applications of the probes in single cell microfluidics, biofilm uptake, and detection of bacteria within macrophages will be presented. The data to date suggests that the probes are valuable tools to aid in understanding antibiotic-bacteria interactions, supporting the fight against antimicrobial resistance.