The rapid emergence of antimicrobial resistant bacteria requires the development of new diagnostic tests. Nucleic acid-based assays 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 on these devices is significantly improved with a Time to Positivity (TTP) half that of the benchtop assay.
Bibliographical noteFunding Information:
The authors would like to thank Ben Hadwen, Chris J. Brown, and Jonathan Buse of Sharp Laboratories Europe for many useful discussions and development of a measurement jig. This work was supported by National Institute for Health Research (NIHR) Invention for Innovation (i4i) Programme grant II-ES-0511-21002 "Rapid detection of infectious agents at point of triage (PoT)". The views expressed in this publication are those of the authors and not necessarily those of the NHS, the National Institute for Health Research, Public Health England or the Department of Health. All data supporting this study are openly available from the University of Southampton repository at http://doi.org/10.5258/SOTON/404856.
© 2017 by the authors.
- Antimicrobial resistance (AMR)
- Digital microfluidics (DMF)
- Isothermal amplification