Rapid Determination of Antimicrobial Susceptibility Using Gel Microdroplets and Flow Cytometry.

摘要

This new method, currently under development in our laboratory, is unique in that no initial pre-culture step is required, thus eliminating a significant rate-limiting step required by other methods currently in use. This makes it possible, in principal, to obtain results quickly enough to guide a clinician's initial choise of antimicrobial agent. Our approach is based on methods of culturing individual cells in gel microdroplets (GMDs). Applying these methods to antimicrobial susceptibility testing, individual cells or colony-forming units are immobilized in GMDs, and then cultured in the presence of candidate antimicrobial agents at several concentrations. Fluorescence-based assays are used to monitor subsequent formation and growth of microcolonies from single cells under control conditions, as well as the inhibition of microcolony formation from cells which are susceptible to the agent under test, at the concentration used. Qualitative effects can be observed with fluorescence microscopy. However, in order to quantify the effects and draw statistically meaningful conclusions, we measure microcolony growth in large numbers of GMDs using flow cytometric analysis.