Beyenal, H

ABSTRACT Biofilms hinder wound infection healing, making wound infections a healthcare challenge for which innovative treatments are needed. A 1.77 cm 2 electrochemical bandage (e-bandage), which generates the biocide hypochlorous acid (HOCl), has shown promising anti-biofilm activity in vitro , ex vivo , and in vivo . The e-bandage is a three-electrode system operationalized by a hydrogel. In addition to providing a conductive electrolyte for the electrodes, the hydrogel provides a protective barrier for the wound. How different hydrogels impact e-bandage functionality is not completely defined. Here, the in vitro activity of seven clinically available hydrogels (xanthan gum, 3M, Duoderm, Prontosan, Purilon, Skintegrity, and Solosite) was evaluated with 1.77 cm 2 HOCl-generating e-bandages using a potentiostat with a multiplexer in a membrane biofilm model against methicillin-resistant Staphylococcus aureus IDRL-6169 and Acinetobacter baumannii ATCC-17978. When applied with polarized HOCl-producing e-bandages, all evaluated hydrogels except Solosite yielded ≥5.5 log 10 CFU/cm 2 reductions in bacterial quantities after 6 h of treatment, with most being below the detection limit at that time point. Prontosan exhibited antibacterial activity independent of e-bandage. Results of this study inform the selection of clinically available hydrogels that may be suitable for use with a 1.77 cm 2 HOCl-producing e-bandage. IMPORTANCE In this study, it was shown that some clinically available hydrogels, combined with a 1.77 cm 2 HOCl-generating electrochemical bandage, deliver time-dependent antimicrobial activity against methicillin-resistant Staphylococcus aureus and Acinetobacter baumannii biofilms, offering a potential approach to treat wound infections, including those caused by antimicrobial-resistant bacteria.

ABSTRACT An electrochemical bandage (e‐bandage) that generates hydrogen peroxide (H 2 O 2 ) through a combination of working, counter and reference electrodes used with an electrolyte‐providing hydrogel is being developed for wound infection management. e‐Bandage biocidal activity was previously demonstrated using Xanthan gum hydrogel. For clinical use, clinically used hydrogels would be ideal, but their use with the described e‐bandage has not been shown. The goal of this work was to evaluate the biocidal activity of off‐the‐shelf, clinically used hydrogels when used with a 1.77 cm 2 H 2 O 2 ‐producing e‐bandage. e‐Bandage electrochemical parameters and activity against methicillin‐resistant Staphylococcus aureus IDRL‐6169 and Acinetobacter baumannii ATCC‐17978 biofilms were assessed with six off‐the‐shelf hydrogels. Variations in hydrogel composition affected electrochemical parameters, which was associated with differences in biocidal activity. Results of this study inform the selection of off‐the‐shelf hydrogels for use with H 2 O 2 ‐producing e‐bandages.

ABSTRACTIntraluminal infection of central venous catheters, used for long‐term treatment, can result in central line‐associated bloodstream infection (CLABSI). These infections can be challenging to prevent and treat due to formation of biofilms within catheter lumens, which shield bacteria from the human immune response and conventional antimicrobial therapies. Preventing bacterial colonization of catheter hubs is a strategy to prevent CLABSI. To address this, we developed a nonantibiotic, animal‐ready electrochemical catheter hub (e‐catheter hub), operated by a wearable, battery‐powered micropotentiostat (MP), that internally generates tunable hypochlorous acid (HOCl) for preventing intraluminal infection. The design evaluated three different electrode materials—titanium, platinum, and gold—for HOCl generation and biocidal activity, using working and counter electrodes of the same materials and a silver/silver chloride‐plated wire as a quasi‐reference electrode. e‐catheter hubs operated by MPs at 1.5 VAg/AgCl for 3 h generated HOCl, reducing Acinetobacter baumannii ATCC‐17978 below the detection limit (average reduction of 4.40 ± 0.05 log10 CFU/mL). The efficacy of e‐catheter hubs operated by MPs in generating HOCl and achieving biocidal activity is comparable to that of a commercial potentiostat. This study represents the first step in developing a localized, nonantibiotic strategy to mitigate CLABSI risk.

ABSTRACT Wound infections, exacerbated by the prevalence of antibiotic-resistant bacterial pathogens, necessitate innovative antimicrobial approaches. Polymicrobial infections, often involving Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus (MRSA), present challenges due to biofilm formation and antibiotic resistance. Hypochlorous acid (HOCl), a potent antimicrobial agent, holds promise as an alternative therapy. An electrochemical bandage (e-bandage) that generates HOCl in situ via precise polarization controlled by a miniaturized potentiostat was evaluated for the treatment of murine wound biofilm infections containing both P. aeruginosa with “difficult-to-treat” resistance and MRSA. Previously, HOCl-producing e-bandage was shown to reduce murine wound biofilms containing P. aeruginosa alone. Here, in 5-mm excisional skin wounds containing 48-h biofilms comprising MRSA and P. aeruginosa combined, polarized e-bandage treatment reduced MRSA by 1.1 log 10 CFU/g ( P = 0.026) vs non-polarized e-bandage treatment (no HOCl production), and 1.4 log 10 CFU/g (0.0015) vs Tegaderm only controls; P. aeruginosa was similarly reduced by 1.6 log 10 CFU/g ( P = 0.0032) and 1.6 log 10 CFU/g ( P = 0.0015), respectively. For wounds infected with MRSA alone, polarized e-bandage treatment reduced bacterial load by 1.1 log 10 CFU/g ( P = 0.0048) and 1.3 log 10 CFU/g ( P = 0.0048) compared with non-polarized e-bandage and Tegaderm only, respectively. The e-bandage treatment did not negatively impact wound healing or cause tissue toxicity. The addition of systemic antibiotics did not enhance the antimicrobial efficacy of e-bandages. This study provides additional evidence for the HOCl-producing e-bandage as a novel antimicrobial strategy for managing wound infections, including in the context of antibiotic resistance and polymicrobial infections. IMPORTANCE New approaches are needed to combat the rise of antimicrobial-resistant infections. The HOCl-producing electrochemical bandage (e-bandage) leverages in situ generation of HOCl, a natural biocide, for broad-spectrum killing of wound pathogens. Unlike traditional therapies that may exhibit limited activity against biofilms and antimicrobial-resistant organisms, the e-bandage offers a potent, standalone solution that does not contribute to further resistance or require adjunctive antibiotic therapy. Here, we show the ability of the e-bandage to address polymicrobial infection by antimicrobial resistant clinical isolates of Staphylococcus aureus and Pseudomonas aeruginosa , two commonly isolated, co-infecting wound pathogens. Effectiveness of the HOCl-producing e-bandage in reducing pathogen load while minimizing tissue toxicity and avoiding the need for systemic antibiotics underscores its potential as a tool in managing complex wound infections.

ABSTRACT Chronic wound infections can be difficult to treat and may lead to impaired healing and worsened patient outcomes. Novel treatment strategies are needed. This study evaluated the effects of intermittently produced hydrogen peroxide (H 2 O 2 ) and hypochlorous acid (HOCl), generated via an electrochemical bandage (e-bandage), against methicillin-resistant Staphylococcus aureus biofilms in an agar membrane biofilm model. By changing the working electrode potential, the e-bandage generated either HOCl (1.5 V Ag/AgCl ) or H 2 O 2 (−0.6 V Ag/AgCl ). The degree of biocidal activity of intermittent treatment with HOCl and H 2 O 2 correlated with HOCl treatment time; HOCl treatment durations of 0, 1.5, 3, 4.5, and 6 hours (with the rest of the 6-hour total treatment time devoted to H 2 O 2 generation) resulted in mean biofilm reductions of 1.36 ± 0.2, 2.22 ± 0.16, 3.46 ± 0.38, 4.63 ± 0.74, and 7.66 ± 0.5 log CFU/cm 2 , respectively, vs. non-polarized controls, respectively. However, application of H 2 O 2 immediately after HOCl treatment was detrimental to biofilm removal. For example, 3 hours HOCl treatment followed by 3 hours H 2 O 2 resulted in a 1.90 ± 0.84 log CFU/cm 2 lower mean biofilm reduction than 3 hours HOCl treatment followed by 3 hours non-polarization. HOCl generated over 3 hours exhibited biocidal activity for at least 7.5 hours after e-bandage operation ceased; 3 hours of HOCl generation followed by 7.5 hours of non-polarization resulted in a biofilm cell reduction of 7.92 ± 0.12 log CFU/cm 2 vs. non-polarized controls. Finally, intermittent treatment with HOCl (i.e., interspersed with periods of e-bandage non-polarization) for various intervals showed similar effects (approximately 6 log CFU/cm 2 reduction vs. non-polarized control) to continuous treatment with HOCl for 3 hours, followed by 3 hours of non-polarization. These findings suggest that timing and sequencing of HOCl and H 2 O 2 treatments are crucial for maximizing biofilm control when using an e-bandage strategy.

AbstractBiofilms formed by antibiotic‐resistant bacteria in wound beds challenge the treatment of wound infections. In this work, the activity of a novel electrochemical bandage (e‐bandage) composed of carbon fabric and controlled by a wearable potentiostat, designed to continuously deliver low amounts of hydrogen peroxide (H2O2), is evaluated against methicillin‐resistant Staphylococcus aureus (MRSA), multidrug‐resistant Pseudomonas aeruginosa (MDR‐PA) and mixed‐species (MRSA and MDR‐PA) wound infections. Wounds created on Swiss Webster mice are infected with the above‐named bacteria and biofilms allow to establish in wound beds for 3 days. e‐Bandages, which electrochemically reduce dissolved oxygen to H2O2 when polarized at −0.6 VAg/AgCl, are placed atop infected wound beds and polarized continuously for 48 h. Polarized e‐bandage treatment results in significant reductions (p <0.001) of both monospecies and mixed‐species wound infections. After e‐bandage treatment, electron microscopy shows degradation of bacterial cells, and histopathology shows no obvious alterations to the inflammatory host response. Blood biochemistries show no abnormalities. Taken all together, results of this work suggest that the described H2O2‐producing e‐bandage can reduce in vivo MRSA, MDR‐PA, and mixed‐species wound biofilms, and should be further developed as a potential antibiotic‐free strategy for treatment of wound infections.

Chronic wound biofilm infections represent a major clinical challenge which results in a substantial burden to patients and healthcare systems. Treatment with topical antibiotics is oftentimes ineffective as a result of antibiotic‐resistant microorganisms and biofilm‐specific antibiotic tolerance. Use of biocides such as hypochlorous acid (HOCl) has gained increasing attention due to the lack of known resistance mechanisms. An HOCl‐generating electrochemical bandage (e‐bandage) is designed that delivers HOCl continuously at low concentrations targeting infected wound beds in a similar manner to adhesive antimicrobial wound dressings. A battery‐operated wearable potentiostat is developed that controls the e‐bandage electrodes at potentials suitable for HOCl generation. It is demonstrated that e‐bandage treatment is tunable by changing the applied potential. HOCl generation on electrode surfaces is verified using microelectrodes. The developed e‐bandage shows time‐dependent responses against in vitro Acinetobacter baumannii and Staphylococcus aureus biofilms, reducing viable cells to nondetectable levels within 6 and 12 h of treatment, respectively. The developed e‐bandage should be further evaluated as an alternative to topical antibiotics to treat wound biofilm infections.

AbstractThe activity of a hypochlorous acid‐producing electrochemical bandage (e‐bandage) in preventing methicillin‐resistant Staphylococcus aureus infection (MRSA) infection and removing biofilms formed by MRSA was assessed using a porcine explant biofilm model. e‐Bandages inhibited S. aureus infection (p = 0.029) after 12 h (h) of exposure and reduced 3‐day biofilm viable cell counts after 6, 12, and 24 h exposures (p = 0.029). Needle‐type microelectrodes were used to assess HOCl concentrations in explant tissue as a result of e‐bandage treatment; toxicity associated with e‐bandage treatment was evaluated. HOCl concentrations in infected and uninfected explant tissue varied between 30 and 80 µM, decreasing with increasing distance from the e‐bandage. Eukaryotic cell viability was reduced by an average of 71% and 65% in fresh and day 3‐old explants, respectively, when compared to explants exposed to nonpolarized e‐bandages. HOCl e‐bandages are a promising technology that can be further developed as an antibiotic‐free treatment for wound biofilm infections.

Abstract Aims Effects of H2O2 producing electrochemical-bandages (e-bandages) on methicillin-resistant Staphylococcus aureus colonization and biofilm removal were assessed using a porcine explant biofilm model. Transport of H2O2 produced from the e-bandage into explant tissue and associated potential toxicity were evaluated. Methods and Results Viable prokaryotic cells from infected explants were quantified after 48 h treatment with e-bandages in three ex vivo S. aureus infection models: (1) reducing colonization, (2) removing young biofilms and (3) removing mature biofilms. H2O2 concentration-depth profiles in explants/biofilms were measured using microelectrodes. Reductions in eukaryotic cell viability of polarized and nonpolarized noninfected explants were compared. e-Bandages effectively reduced S. aureus colonization (p = 0.029) and reduced the viable prokaryotic cell concentrations of young biofilms (p = 0.029) with limited effects on mature biofilms (p &gt; 0.1). H2O2 penetrated biofilms and explants and reduced eukaryotic cell viability by 32–44% compared to nonpolarized explants. Conclusions H2O2 producing e-bandages were most active when used to reduce colonization and remove young biofilms rather than to remove mature biofilms. Significance and Impact of Study The described e-bandages reduced S. aureus colonization and young S. aureus biofilms in a porcine explant wound model, supporting their further development as an antibiotic-free alternative for managing biofilm infections.

Abstract Aims Wound infections involving Candida albicans can be challenging to treat because of the fungus’ ability to penetrate wound tissue and form biofilms. The goal of this study was to assess the activity of a hypochlorous acid (HOCl)-generating electrochemical scaffold (e-scaffold) against C. albicans biofilms in vitro and on porcine dermal explants (ex vivo). Methods and Results C. albicans biofilms were grown either on acrylic-bottom six-well plates (in vitro) or on skin tissue excised from porcine ears (ex vivo), and the polarized e-scaffold was used to generate a continuous supply of low concentration HOCl near biofilm surfaces. C. albicans biofilms grown in vitro were reduced to undetectable amounts within 24 h of e-scaffold exposure, unlike control biofilms (5·28 ± 0·034 log10 (CFU cm-2); P &lt; 0·0001). C. albicans biofilms grown on porcine dermal explants were also reduced to undetectable amounts in 24 h, unlike control explant biofilms (4·29 ± 0·057 log10 (CFU cm-2); P &lt; 0·0001). There was a decrease in the number of viable mammalian cells (35·6 ± 6·4%) in uninfected porcine dermal explants exposed to continuous HOCl-generating e-scaffolds for 24 h compared to explants exposed to nonpolarized e-scaffolds (not generating HOCl) (P &lt; 0·05). Conclusions Our HOCl-generating e-scaffold is a potential antifungal-free strategy to treat C. albicans biofilms in chronic wounds. Significance and Impact of the Study Wound infections caused by C. albicans are difficult to treat due to presence of biofilms in wound beds. Our HOCl producing e-scaffold provides a promising novel approach to treat wound infections caused by C. albicans.