Electrotaxis or galvanotaxis refers to the migration pattern of cells induced in response to electrical potential. Although it has been extensively studied in mammalian cells, electrotaxis has not been explored in detail in bacterial cells; information regarding the impact of current on pathogenic bacteria is severely lacking. Therefore, we designed a series of single and multi-cue experiments to assess the impact of varying currents on bacterial motility dynamics in pathogenic multi-drug resistant (MDR) strains of Pseudomonas aeruginosa and Escherichia coli using a microfluidic platform. Motility plays key roles in bacterial migration and the colonization of surfaces during the formation of biofilms, which are inherently recalcitrant to removal and resistant to traditional disinfection strategies (e.g. antibiotics). Use of the microfluidic platform allows for exposure to current, which can be supplied at a range that is biocidal to bacteria, yet physiologically safe in humans (single cue). This system also allows for multi-cue experiments where acetic acid, a relatively safe compound with anti-fouling/antimicrobial properties, can be combined with current to enhance disinfection. These strategies may offer substantial therapeutic benefits, specifically for the treatment of biofilm infections, such as those found in the wound environment. Furthermore, microfluidic systems have been successfully used to model the unique microfluidic dynamics present in the wound environment, suggesting that these investigations could be extended to more complex biological systems. Our results showed that the application of current in combination with acetic acid has profound inhibitory effects on MDR strains of P. aeruginosa and E. coli, even with brief applications. Specifically, E. coli motility dynamics and cell survival were significantly impaired starting at a concentration of 125 μA DC and 0.31% acetic acid, while P. aeruginosa was impaired at 70 μA and 0.31% acetic acid. As these strains are relevant wound pathogens, it is likely that this strategy would be effective against similar strains in vivo and could represent a new approach to hasten wound healing.

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Electrotaxis, or galvanotaxis, refers to the migration pattern of cells induced in response to electrical potential. Electrotaxis has not been explored in detail in bacterial cells; information regarding the impact of current on pathogenic bacteria is severely lacking. Using microfluidic platforms and optical microscopy, we designed a series of single- and multi-cue experiments to assess the impact of varying electrical currents and acetic acid concentrations on bacterial motility dynamics in pathogenic multi-drug resistant (MDR) strains of Pseudomonas aeruginosa and Escherichia coli. The use of the microfluidic platform allows for single-cue experiments where electrical current is supplied at a range that is biocidal to bacteria and multi-cue experiments where acetic acid is combined with current to enhance disinfection. These strategies may offer substantial therapeutic benefits, specifically for the treatment of biofilm infections, such as those found in the wound environment. Our results showed that an application of current in combination with acetic acid has profound inhibitory effects on MDR strains of P. aeruginosa and E. coli, even with brief applications. Specifically, E. coli motility dynamics and cell survival were significantly impaired starting at a concentration of 0.125 mA of direct current (DC) and 0.31% acetic acid, while P. aeruginosa was impaired at 0.70 mA and 0.31% acetic acid. As these strains are relevant wound pathogens, it is likely that this strategy would be effective against similar strains in vivo and could represent a new approach to hasten wound healing.

7Biofilms are natural, resilient films formed when microorganisms adhere to a surface and form a 8 complex three-dimensional structure that allows them to persist in a wide variety of environments. 9 Readily forming in hospitals and on medical equipment, biofilms are frequent causes of infections 10 and their subsequent complications. Due to the complexity of these structures, systematically 11 studying individual bacterial cells and their interactions with their surrounding environment will 12 provide a deeper understanding of the processes occurring within the biofilm as whole versus bulk 13 population based methods that do not differentiate individual cells or species. Methods based on 14 atomic force microscopy (AFM) are particularly suited to the study of individual cells, but are 15 underutilized for the study of bacterial electrical properties. The ability of electrical currents to 16 impair bacterial attachment is well documented, but to utilize electrical current as an effective 17 antibacterial treatment, it is important to understand the electrical properties of bacteria. Therefore, 18 we used AFM, Kelvin probe force microscopy, and ResiScope to measure the surface potential 19 and conductance of Psuedomonas aeruginosa and methicillin resistance Staphylococcus aureus 20(MRSA) on gold and stainless steel. This is the first study to directly measure the electrical 21 resistance of single bacterial cells using ResiScope. Our goal was to develop a framework for 22 measuring biological molecules using conductive atomic force microscopy. We found that the 23 2 average resistance for P. aeruginosa was 135.4±25.04 GΩ, while MRSA had an average of 24 173.4±16.28 GΩ. Using KPFM, the surface potential of MRSA shifted from -0.304 V to 0.153 V 25 and from -0.280 V to 0.172 V for P. aeruginosa on gold versus stainless steel substrates, 26 respectively. In an attempt to identify a potential charge carrier, peptidoglycan was also measured 27 with the ResiScope module and shown to have a resistance of 105 GΩ. 28

Biofilms are natural, resilient films formed when microorganisms adhere to a surface and form a complex three-dimensional structure that allows them to persist in a wide variety of environments. Readily forming in hospitals and on medical equipment, biofilms are frequent causes of infections and their subsequent complications. Due to the complexity of these structures, systematically studying individual bacterial cells and their interactions with their surrounding environment will provide a deeper understanding of the processes occurring within the biofilm as whole versus bulk population based methods that do not differentiate individual cells or species. Methods based on atomic force microscopy (AFM) are particularly suited to the study of individual cells, but are underutilized for the study of bacterial electrical properties. The ability of electrical currents to impair bacterial attachment is well documented, but to utilize electrical current as an effective antibacterial treatment, it is important to understand the electrical properties of bacteria. Therefore, we used AFM, Kelvin probe force microscopy, and ResiScope to measure the surface potential and conductance of Psuedomonas aeruginosa and methicillin resistance Staphylococcus aureus (MRSA) on gold and stainless steel. This is the first study to directly measure the electrical resistance of single bacterial cells using ResiScope. Our goal was to develop a framework for measuring biological molecules using conductive atomic force microscopy. We found that the. CC-BY-NC-ND 4.