Determination of antibiotic EC₅₀using a zero‐flow microfluidic chip based growth phenotype assay

Abstract

Current existing assay systems for evaluating antimicrobial activity suffer from several limitations including excess reagent consumption and inaccurate concentration gradient preparation. Recently, microfluidic systems have been developed to provide miniaturized platforms for antimicrobial susceptibility assays. However, some of current microfluidic based assays require continuous flows of reagents or elaborate preparation steps during concentration preparation. In this study, we introduce a novel microfluidic chip based growth phenotype assay that automatically generates a logarithmic concentration gradient and allows observing the growth of pathogenic bacteria under different concentrations of antibiotics in nanoliter batch culture reactors. We chose pathogen bacterium Pseudomonas aeruginosa as a model strain and evaluated the inhibitory effects of gentamicin and ciprofloxacin. We determined the EC50 values and confirmed the validity of the present system by comparing the EC50 values obtained through conventional test tube method. We demonstrated that the EC50 values acquired from present assay are comparable to those obtained from conventional test tube cultures. The potential application of present assay system for investigating combinatorial effects of antibiotics on multidrug resistant pathogenic bacteria is discussed and it can be further used for systematic evaluation of antifungal or antiviral agents.

Microfluidic systems for rapid antibiotic susceptibility tests (ASTs) at the single-cell level

“…73 To rapidly test the performance of these newly identied drug candidates, the high throughput microuidic systems show great potential. 74,75 The single-cell AST also holds huge potential for high throughput single bacterial heterogeneity analysis. 76 Since the antimicrobial susceptibility may differ among individual cells, the adequate detection of single bacterial heterogeneity is of great clinical importance.…”

Microfluidic advances in phenotypic antibiotic susceptibility testing

“…have reported on a microfluidic AST device that measured the EC 50 values of antibiotics against GFP-expressing P. aeruginosa in a zero-flow microfluidic chip (Dai et al 2015). Each of the 14 chambers has an antibiotic metering channel, for concentration gradient formation, and a cell channel (3.5×10 8 CFU/mL).…”

Micro- and Nanoscale Approaches in Antifungal Drug Discovery

“…Since microfluidics promises several advantages over existing macro-scale methods, several microfluidic platforms that can perform rapid antimicrobial susceptibility tests have been developed during the last years (Table 3) [132,135]. The recent development of microfluidic platforms was mostly focused on devices for antibiotics susceptibility testing and much less on antifungals susceptibility testing, although the same designs could, usually, be used for both (e.g., see [136,137]).…”