Programmable digital microfluidic assay for simultaneous detection of multiple anti-microbial resistance genes

The rapid emergence of antimicrobial resistant bacteria requires the development of new diagnostic tests. Nucleic acid based assay determine antimicrobial susceptibility by detecting genes that encode for the resistance. In this study, we demonstrate rapid and simultaneous detection of three genes that confer resistance in bacteria to extended spectrum β-lactam and carbapenem antibiotics; CTX-M-15, KPC and NDM-1. The assay uses isothermal DNA amplification (Recombinase Polymerase Amplification, RPA) implemented on a programmable digital microfluidics (DMF) platform. Automated dispensing protocols are used to simultaneously manipulate 45 droplets of nL volume containing sample DNA, reagents and controls. The droplets are processed and mixed under electronic control on the DMF devices with positive amplification measured by fluorescence. The assay is fast, has two orders of magnitude improved sensitivity compared with a benchtop assay.