Here, we demonstrate the use of a recently developed electrochemical sensing platform using transparent carbon ultramicroelectrode arrays (T-CUAs) for the in vitro detection of pyocyanin and other phenazine metabolites. Pyocyanin is a bacterial warfare toxin secreted by the opportunistic human pathogen Pseudomonas aeruginosa, which is commonly found in chronic wounds and in the lungs of cystic fibrosis patients. Pyocyanin and other phenazine species are highly diffusible signaling molecules that are toxic to surrounding host cells and other competing microorganisms. Therefore, the detection of these species is of high importance as it could provide insights regarding P. aeruginosa virulence. T-CUAs are low-cost sensing platform and provide rapid, sensitive, reproducible and direct detection of pyocyanin compared to conventional, time-consuming, and expensive spectrophotometric methods. We quantified pyocyanin concentrations on T-CUAs and determined limits of detection (LODs) and linear dynamic ranges (LDRs), which fall within reported micromolar ranges for a variety of in vitro and in vivo cellular environments. Pyocyanin and other redox-active phenazine metabolites play fundamental roles in cellular functions and P. aeruginosa virulence mechanisms, however, very little is known about how their concentrations vary and direct cellular behavior over time. Thus, we present real-time electrochemical monitoring of pyocyanin and another highly reactive, redox-active phenazine metabolite from a highly virulent laboratory P. aeruginosa strain, PA14, using T-CUAs. Our quantitative results show increasing phenazine concentrations over a 48-hour period as well as changes in metabolism dynamics over time in two different growth media. Finally, desorption electrospray ionization mass spectrometry imaging (DESI-MSI) and nano-electrospray ionization (nano-ESI) are used to confirm the identity of phenazine species observed and quantified with our electrochemical sensors.

Real-time electrochemical detection of Pseudomonas aeruginosa phenazine metabolites using transparent carbon ultramicroelectrode arrays

2019 Spring National Meeting

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