Impedimetric detection of Pseudomonas aeruginosa attachment on flexible ITO-coated polyethylene terephthalate substrates

Biofilm monitoring in environmental and biomedical applications remains a challenge. Currently, conventional biochemical methods do not provide a quick quantitative measure of attached biomass. Thus, there is a need for rapid in situ detection tools for routine biofilm characterization. Electrochemical impedance spectroscopy (EIS) characterizes the electroactivity of bacteria within a biofilm and has been extensively used to monitor strong electroactive biofilms. Yet, studies on weak electricigens such as Pseudomonas aeruginosa remain underrepresented. Here, conductive indium tin oxide coated polyethylene terephthalate (ITO:PET) sheets were used as flexible growth substrates instead of more conventional carbonaceous or gold materials. EIS was compared with standard optical methods for the detection of P. aeruginosa biofilms formed on ITO:PET under static growth conditions. Relaxation time analysis showed a dominant time constant at approximately 1 s and confirmed the validity of a two time constant equivalent circuit model for the biofilm impedance. The interfacial resistance calculated from the equivalent circuit analysis showed a rapid drop after bacterial attachment whereas capacitance of the biofilm was masked by the capacitance of ITO:PET. The trends for interfacial resistance and capacitance were independent to the geometry of the ITO:PET working electrode. Moreover, most robust behaviour was observed for rectangular electrodes. EIS across a broad range of potentials with and without inhibitors showed a marked difference between the interfacial resistance of viable and energy inhibited biofilms. Moreover, EIS of exopolysaccharide Δpsl mutant showed a substantial drop in current. Overall, our results indicated that EIS enabled the detection of biofilm formation across large surface areas as early as 24 h after inoculation for the weak electroactive species P. aeruginosa using a flexible polymeric substrate