Fast and Sensitive Antibiotic Susceptibility Testing Using Interferometry
A Krueger1*, P Yunker1, D Weiss2, (1) Georgia Institute of Technology, Atlanta, GA, (2) Emory University, Atlanta, GA
Presentations
PO-GePV-M-282 (Sunday, 7/10/2022) [Eastern Time (GMT-4)]
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Purpose: Our goal is to improve the speed and sensitivity of antibiotic susceptibility testing, and thus detect even small numbers of resistant cells. Doing so will enable faster, more informed treatment plans, better able to combat growing threats of antibiotic resistance.
Methods: Interferometry was used to provide a significant improvement in out-of-plane resolution of biofilm topographies compared to typical imaging techniques with three-dimensional resolution like confocal microscopy. We mix two genetically modified Vibrio cholerae strains, one that produces LacZ, making it resistant to Kanamycin and one that does not produce LacZ and is thus susceptible to Kanamycin to create an ‘artificial’ strain heteroresistant to Kanamycin. We mix the two strains at varying proportions of resistant to susceptible bacteria – 1:10, 1:100, and 1:1000. These heterogenous populations were inoculated onto Kanamycin agar plates and placed under the objective of an interferometer while incubated at 37C. Topographic images with single nanometer height resolution were taken at regular intervals, creating time-lapses lasting two to three hours after inoculation.
Results: Analysis of these images illuminates the biophysically important features of heteroresistant biofilm topographies. Mean height and biomass weighted histograms are among the most descriptive features of these images to detect bacterial growth. With p-value less than 0.0001, resistance was differentiated from susceptible strains in as little as 45 minutes, and with p-value less than .0025, high levels of heteroresistance were differentiated from susceptible in 90 minutes.
Conclusion: This work using interferometrically imaged topographies shows the potential for a major shift in antibiotic susceptibility testing methods to drastically decrease the time to classification. Even when surrounded by susceptible cells, resistant cells can be identified, demonstrating the potential to detect heteroresistance. More work is being done to improve the sensitivity of these measurements by optimizing the biophysical features associated with growth in a biofilm.
Funding Support, Disclosures, and Conflict of Interest: 1R35GM138354 from NIH/NIGMS
Keywords
Fast Fourier Transformation, Fluorescence, Health Physics
Taxonomy
IM- Optical : Microscope-based imaging
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