Technologies for High-Throughput Identification of Antibiotic Mechanism of Action

Abstract

There are two main strategies for antibiotic discovery: target-based and phenotypic screening. The latter has been much more successful in delivering first-in-class antibiotics, despite the major bottleneck of delayed Mechanism-of-Action (MOA) identification. Although finding new antimicrobial compounds is a very challenging task, identifying their MOA has proven equally challenging. MOA identification is important because it is a great facilitator of lead optimization and improves the chances of commercialization. Moreover, the ability to rapidly detect MOA could enable a shift from an activity-based discovery paradigm towards a mechanism-based approach. This would allow to probe the grey chemical matter, an underexplored source of structural novelty. In this study we review techniques with throughput suitable to screen large libraries and sufficient sensitivity to distinguish MOA. In particular, the techniques used in chemical genetics (e.g., based on overexpression and knockout/knockdown collections), promoter-reporter libraries, transcriptomics (e.g., using microarrays and RNA sequencing), proteomics (e.g., either gel-based or gel-free techniques), metabolomics (e.g., resourcing to nuclear magnetic resonance or mass spectrometry techniques), bacterial cytological profiling, and vibrational spectroscopy (e.g., Fourier-transform infrared or Raman scattering spectroscopy) were discussed. Ultimately, new and reinvigorated phenotypic assays bring renewed hope in the discovery of a new generation of antibiotics.

High-Throughput Screening of Natural Product and Synthetic Molecule Libraries for Antibacterial Drug Discovery

“…Other emerging technologies such as biolayer interferometry (BLI), microscale thermophoresis (MST), surface acoustic waves, second-harmonic generation, waveguide-based grating-coupled interferometry, and electrically switchable nanolevers that are based on photometry, fluorometry, wave, and temperature gradient may be explored in the future for antibacterial drug discovery research. Irrespective of the various innovative strategies for HTS assays, finding new antibacterial agents, as well as determining their mechanisms of action, have been very challenging, and da Cunha et al reviewed various techniques such as overexpression and knockout-based genetics, promoter–receptor libraries, transcriptomics, proteomics, metabolomics, bacterial cytological profiling, and vibrational spectroscopy that are currently being used in mechanism-based antibacterial drug discovery [ 415 ] that the readers are encouraged to consult.…”

The Phytochemical Tactics for Battling Antibiotic Resistance in Microbes: Secondary Metabolites and Nano Antibiotics Methods

“…Much of the attention has been given in the recent past to developing molecules which, when administered at low concentrations, can inhibit or kill a large number of pathogenic organisms by keeping the adverse effects on patients as low as possible. [24] This search yielded only a few types of acceptable antibiotics among a larger group of antibacterial agents, and more specifically anti-filarial, malaria, and antifungal compounds. Overall, very few suitable antibacterial agents have been found, and the same antibacterial agents have been repeatedly discovered, suggesting a low overall variety of such relatively deadly compounds.…”

Antibacterial mechanism of vanillin against Escherichia coli O157: H7

“…As a new type of omics [ 27 ], transcriptomics is different from most traditional techniques (dye penetration, electron microscopy imaging and in vitro DNA binding assay) that cannot elucidate the mechanism of antibacterial molecules of natural antibacterial substances. Transcriptomics can generate large amounts of data through high-throughput sequencing technology, which can analyze extremely complex features [ 28 ]. The molecular mechanism was also elucidated.…”