Issue 24, 2024

Label-free single-cell antimicrobial susceptibility testing in droplets with concentration gradient generation

Abstract

Bacterial communities exhibit significant heterogeneity, resulting in the emergence of specialized phenotypes that can withstand antibiotic exposure. Unfortunately, the existence of subpopulations resistant to antibiotics often goes unnoticed during treatment initiation. Thus, it is crucial to consider the concept of single-cell antibiotic susceptibility testing (AST) to tackle bacterial infections. Nevertheless, its practical application in clinical settings is hindered by its inability to conduct AST efficiently across a wide range of antibiotics and concentrations. This study introduces a droplet-based microfluidic platform designed for rapid single-cell AST by creating an antibiotic concentration gradient. The advantage of a microfluidic platform is achieved by executing bacteria and antibiotic mixing, cell encapsulation, incubation, and enumeration of bacteria in a seamless workflow, facilitating susceptibility testing of each antibiotic. Firstly, we demonstrate the rapid determination of minimum inhibitory concentration (MIC) of several antibiotics with Gram-negative E. coli and Gram-positive S. aureus, which enables us to bypass the time-consuming bacteria cultivation, speeding up the AST in 3 h from 1 to 2 days of conventional methods. Additionally, we assess 10 clinical isolates including methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant Staphylococcus aureus (MDRSA) against clinically important antibiotics for analyzing the MIC, compared to the gold standard AST method from the United States Clinical and Laboratory Standards Institute (CLSI), which becomes available only after 48 h. Furthermore, by monitoring single cells within individual droplets, we have found a spectrum of resistance levels among genetically identical cells, revealing phenotypic heterogeneity within isogenic populations. This discovery not only advances clinical diagnostics and treatment strategies but also significantly contributes to the field of antibiotic stewardship, underlining the importance of our approach in addressing bacterial resistance.

Graphical abstract: Label-free single-cell antimicrobial susceptibility testing in droplets with concentration gradient generation

Supplementary files

Article information

Article type
Paper
Submitted
28 Jul 2024
Accepted
19 Sep 2024
First published
20 Sep 2024

Lab Chip, 2024,24, 5274-5289

Label-free single-cell antimicrobial susceptibility testing in droplets with concentration gradient generation

J. S. Kim, J. Kim, J. Kim, W. Kim and C. Lee, Lab Chip, 2024, 24, 5274 DOI: 10.1039/D4LC00629A

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