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1. Hypochlorous acid-generating electrochemical scaffold eliminates Candida albicans biofilms

   https://doi.org/10.1111/jam.14656  

   Zmuda, H. M. ; Mohamed, A. ; Raval, Y. S. ; Call, D. R. ; Schuetz, A. N. ; Patel, R. ; Beyenal, H. ( October 2020 , Journal of Applied Microbiology)

   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).

   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 < 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 < 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 < 0·05).

   Our HOCl-generating e-scaffold is a potential antifungal-free strategy to treat C. albicans biofilms in chronic wounds.

   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.

    

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2. Non‐invasive imaging of oxygen concentration in a complex in vitro biofilm infection model using 19 F MRI: Persistence of an oxygen sink despite prolonged antibiotic therapy

   https://doi.org/10.1002/mrm.27888  

   Simkins, Jeffrey W. ; Stewart, Philip S. ; Codd, Sarah L. ; Seymour, Joseph D. ( August 2019 , Magnetic Resonance in Medicine)

   Oxygen availability is a critical determinant of microbial biofilm activity and antibiotic susceptibility. However, measuring oxygen gradients in these systems remains difficult, with the standard microelectrode approach being both invasive and limited to single‐point measurement. The goal of the study was to develop a¹⁹F MRI approach for 2D oxygen mapping in biofilm systems and to visualize oxygen consumption behavior in real time during antibiotic therapy.

   Oxygen‐sensing beads were created by encapsulating an emulsion of oxygen‐sensing fluorocarbon into alginate gel.Escherichia colibiofilms were grown in and on the alginate matrix, which was contained inside a packed bed column subjected to nutrient flow, mimicking the complex porous structure of human wound tissue, and subjected to antibiotic challenge.

   The linear relationship between¹⁹F spin‐lattice relaxation rateR₁and local oxygen concentration permitted noninvasive spatial mapping of oxygen distribution in real time over the course of biofilm growth and subsequent antibiotic challenge. This technique was used to visualize persistence of microbial oxygen respiration during continuous gentamicin administration, providing a time series of complete spatial maps detailing the continued bacterial utilization of oxygen during prolonged chemotherapy in an in vitro biofilm model with complex spatial structure.

   Antibiotic exposure temporarily causes oxygen consumption to enter a pseudosteady state wherein oxygen distribution becomes fixed; oxygen sink expansion resumes quickly after antibiotic clearance. This technique may provide valuable information for future investigations of biofilms by permitting the study of complex geometries (typical of in vivo biofilms) and facilitating noninvasive oxygen measurement.

    

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3. Contribution of Pseudomonas aeruginosa Exopolysaccharides Pel and Psl to Wound Infections

   https://doi.org/10.3389/fcimb.2022.835754  

   Fleming, Derek ; Niese, Brandon ; Redman, Whitni ; Vanderpool, Emily ; Gordon, Vernita ; Rumbaugh, Kendra P. ( April 2022 , Frontiers in Cellular and Infection Microbiology)

   Biofilms are the cause of most chronic bacterial infections. Living within the biofilm matrix, which is made of extracellular substances, including polysaccharides, proteins, eDNA, lipids and other molecules, provides microorganisms protection from antimicrobials and the host immune response. Exopolysaccharides are major structural components of bacterial biofilms and are thought to be vital to numerous aspects of biofilm formation and persistence, including adherence to surfaces, coherence with other biofilm-associated cells, mechanical stability, protection against desiccation, binding of enzymes, and nutrient acquisition and storage, as well as protection against antimicrobials, host immune cells and molecules, and environmental stressors. However, the contribution of specific exopolysaccharide types to the pathogenesis of biofilm infection is not well understood. In this study we examined whether the absence of the two main exopolysaccharides produced by the biofilm former Pseudomonas aeruginosa would affect wound infection in a mouse model. Using P. aeruginosa mutants that do not produce the exopolysaccharides Pel and/or Psl we observed that the severity of wound infections was not grossly affected; both the bacterial load in the wounds and the wound closure rates were unchanged. However, the size and spatial distribution of biofilm aggregates in the wound tissue were significantly different when Pel and Psl were not produced, and the ability of the mutants to survive antibiotic treatment was also impaired. Taken together, our data suggest that while the production of Pel and Psl do not appear to affect P. aeruginosa pathogenesis in mouse wound infections, they may have an important implication for bacterial persistence in vivo. 

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4. Hypochlorous-Acid-Generating Electrochemical Scaffold for Treatment of Wound Biofilms

   https://doi.org/10.1038/s41598-019-38968-y  

   Kiamco, Mia Mae ; Zmuda, Hannah M. ; Mohamed, Abdelrhman ; Call, Douglas R. ; Raval, Yash S. ; Patel, Robin ; Beyenal, Haluk ( February 2019 , Scientific Reports)

   Biofilm formation causes prolonged wound infections due to the dense biofilm structure, differential gene regulation to combat stress, and production of extracellular polymeric substances.Acinetobacter baumannii,Staphylococcus aureus, andPseudomonas aeruginosaare three difficult-to-treat biofilm-forming bacteria frequently found in wound infections. This work describes a novel wound dressing in the form of an electrochemical scaffold (e-scaffold) that generates controlled, low concentrations of hypochlorous acid (HOCl) suitable for killing biofilm communities without substantially damaging host tissue. Production of HOCl near the e-scaffold surface was verified by measuring its concentration using needle-type microelectrodes. E-scaffolds producing 17, 10 and 7 mM HOCl completely eradicatedS. aureus,A. baumannii, andP. aeruginosabiofilms after 3 hours, 2 hours, and 1 hour, respectively. Cytotoxicity and histopathological assessment showed no discernible harm to host tissues when e-scaffolds were applied to explant biofilms. The described strategy may provide a novel antibiotic-free strategy for treating persistent biofilm-associated infections, such as wound infections.

    

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5. Reverse diauxie phenotype in Pseudomonas aeruginosa biofilm revealed by exometabolomics and label-free proteomics

   https://doi.org/10.1038/s41522-019-0104-7  

   Yung, Yeni P. ; McGill, S. Lee ; Chen, Hui ; Park, Heejoon ; Carlson, Ross P. ; Hanley, Luke ( October 2019 , npj Biofilms and Microbiomes)

   Microorganisms enhance fitness by prioritizing catabolism of available carbon sources using a process known as carbon catabolite repression (CCR). Planktonically grownPseudomonas aeruginosais known to prioritize the consumption of organic acids including lactic acid over catabolism of glucose using a CCR strategy termed “reverse diauxie.”P. aeruginosais an opportunistic pathogen with well-documented biofilm phenotypes that are distinct from its planktonic phenotypes. Reverse diauxie has been described in planktonic cultures, but it has not been documented explicitly inP. aeruginosabiofilms. Here a combination of exometabolomics and label-free proteomics was used to analyze planktonic and biofilm phenotypes for reverse diauxie.P. aeruginosabiofilm cultures preferentially consumed lactic acid over glucose, and in addition, the cultures catabolized the substrates completely and did not exhibit the acetate secreting “overflow” metabolism that is typical of many model microorganisms. The biofilm phenotype was enabled by changes in protein abundances, including lactate dehydrogenase, fumarate hydratase, GTP cyclohydrolase, L-ornithine N(5)-monooxygenase, and superoxide dismutase. These results are noteworthy because reverse diauxie-mediated catabolism of organic acids necessitates a terminal electron acceptor like O₂, which is typically in low supply in biofilms due to diffusion limitation. Label-free proteomics identified dozens of proteins associated with biofilm formation including 16 that have not been previously reported, highlighting both the advantages of the methodology utilized here and the complexity of the proteomic adaptation forP. aeruginosabiofilms. Documenting the reverse diauxic phenotype inP. aeruginosabiofilms is foundational for understanding cellular nutrient and energy fluxes, which ultimately control growth and virulence.

    

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