Zhang, Y; Kepiro, I; Ryadnov, M G; Pagliara, S (2023) Single cell killing kinetics differentiate phenotypic bacterial responses to different antibacterial classes. Microbiology Spectrum, 11 (1). e03667-22

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Abstract

Antibiotics save millions of lives every year by preventing bacterial infections. With the spread of multi-drug resistant bacteria, there is an increasing focus on molecular classes that have not had an antibiotic before. A key capability in assessing and prescribing new antibacterial treatments is to compare the effects antibacterial agents have on bacterial growth at a phenotypic, single-cell level. Here we combine time-lapse microscopy with microfluidics to investigate concentration-dependent killing kinetics of Escherichia coli cells using antibacterial agents from three different molecular classes – b-lactams and fluoroquinolones, with known antibiotics ampicillin and ciprofloxacin, and a new experimental class of protein Ψ-capsids. We found that bacteria elongated when treated with ampicillin and ciprofloxacin used at their minimum growth inhibitory concentrations. This was in contrast to Ψ-capsids, which arrested bacterial elongation within the first two hours of treatment. At concentrations exceeding minimum inhibitory concentrations all the antibacterial agents tested arrested bacterial growth within the first two hours of treatment. Further, the tests revealed differences in modes of action for the agents. At minimum inhibitory concentrations ampicillin and ciprofloxacin caused the lysis of bacterial cells, whereas at higher concentrations the mode of action shifted towards membrane disruption. Ψ-capsids killed bacterial by disrupting their membranes at all concentrations tested. Finally, with increasing concentrations ampicillin and Ψ-capsids reduced the number of bacteria that survived treatment in a viable but non culturable state, whereas ciprofloxacin increased this number. The study introduces an effective capability to differentiate killing kinetics of antibacterial agents from different molecular classes and offers a high content analysis of antibacterial mechanisms at the single-cell level.

Item Type:	Article
Keywords:	antibiotics, Escherichia coli, microfluidics, antibiotic susceptibility testing, bacterial membranes, minimum inhibitory concentration, single-cell analysis, ampicillin, antimicrobial resistance, ciprofloxacin, persisters, viable but nonculturable
Subjects:	Biotechnology > Biopharmaceutical Manufacturing and Characterisation
Divisions:	Chemical & Biological Sciences
Identification number/DOI:	10.1128/spectrum.03667-22
Last Modified:	02 May 2023 13:13
URI:	https://eprintspublications.npl.co.uk/id/eprint/9695