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Many bacterial species readily develop biofilms that act as a protectivematrix against external challenge, e.g. from antimicrobial treatment.Therefore, biofilms are often responsible for persistent and recurringinfections. Established methods for studying biofilms are either destructive orthey focus on the biofilm surface. A non-destructive method that is sensitiveto the underside of the biofilm is highly desirable, as it allows studying thepenetration of antibiotics through the film. Here, we demonstrate that the highsurface sensitivity of resonant hyperspectral imaging provides this capability.The method allows us to monitor the early stages of Escherichia coli biofilmformation, cell attachment and microcolony formation, in-situ and in real time.We study the response of the biofilm to a number of different antibiotics andverify our observations using confocal microscopy. Based on this ability toclosely monitor the surface-bound cells, resonant hyperspectral imaging givesnew insights into the antimicrobial resistance of biofilms.

Attachment and antibiotic response of early-stage biofilms studied using resonant hyperspectral imaging