A multiplexed microfluidic platform for rapid antibiotic susceptibility testing

Effective treatment of clinical infections is critically dependent on the ability to rapidly screen patient samples to identify antibiograms of infecting pathogens. Existing methods for antibiotic susceptibility testing suffer from several disadvantages, including long turnaround times, excess sample and reagent consumption, poor detection sensitivity, and limited combinatorial capabilities. Unfortunately, these factors preclude the timely administration of appropriate antibiotics, complicating management of infections and exacerbating the development of antibiotic resistance. Here, we seek to address these issues by developing a microfluidic platform that relies on fluorescence detection of bacteria that express green fluorescent protein for highly sensitive and rapid antibiotic susceptibility testing. This platform possesses several advantages compared to conventional methods: (1) analysis of antibiotic action in two to four hours, (2) enhanced detection sensitivity (≈1 cell), (3) minimal consumption of cell samples and antibiotic reagents (<6 µL), and (4) improved portability through the implementation of normally closed valves. We employed this platform to quantify the effects of four antibiotics (ampicillin, cefalexin, chloramphenicol, tetracycline) and their combinations on Escherichia coli. Within four hours, the susceptibility of bacteria to antibiotics can be determined by detecting variations in maxima of local fluorescence intensity over time. As expected, cell density is a major determinant of antibiotic efficacy. Our results also revealed that combinations of three or more antibiotics are not necessarily better for eradicating pathogens compared to pairs of antibiotics. Overall, this microfluidic based biosensor technology has the potential to provide rapid and precise guidance in clinical therapies by identifying the antibiograms of pathogens.