Electrical Stimulation Signficantly Impacts Biofilm Viability, Metabolism, Biomass And Volatile Organic Compound Profiles
Mohammed Ashrafi1, Lilyann Novak-Frazer1, Mohamed Baguneid2, Teresa Alonso-Rasgado1, Riina Rautemaa-Richardson1, Ardeshir Bayat1.
1The University of Manchester, Manchester, United Kingdom, 2Manchester University NHS Foundation Trust, Manchester, United Kingdom.
BACKGROUND - Antibiotic resistance and inefficiencies in monitoring response to treatment of wound infections are of significant concern. Electrical stimulation (ES) has anti-bacterial effects and could be used as an alternative to antibiotics. Microorganisms produce volatile organic compounds (VOCs) and we have previously identified specific VOC profiles of biofilms formed on human ex-vivo cutaneous wound models. The aim here was to assess the antimicrobial effects of ES and monitor variations in VOCs in response to ES and antibiotic therapy.
METHODS - Staphylococcus aureus and Pseudomonas aeruginosa biofilms were formed in vitro and subjected to either direct current ES, high dose ciprofloxacin or no treatment at early (24h) and late (72h) phases. Response to treatment was quantified using bacterial counts, metabolic (XTT) and biomass (PicoGreen) assays. Biofilm headspace was sampled at 24 hours post treatment and VOCs separated by gas chromatography and detected by mass spectrometry. VOCs were chemically identified using the National Institute of Standards and Technology library and relative abundances compared.
RESULTS - ES treated S. aureus biofilms showed significant reduction in bacterial counts (n=6, P<0.001; 24 and 72h) and biomass (P<0.001; 72h) compared to controls. ES treated P. aeruginosa biofilms showed a significant reduction in bacterial counts (P≤0.008; 24 and 72h), metabolic activity (P=0.014; 24h) and biomass (P=0.026; 72h) compared to controls. There were significant variations in relative abundances of VOCs between treatment groups (P<0.05). Butanedione and acetic acid ethenyl ester were specific to ciprofloxacin treated S. aureus biofilms (P<0.001). S. aureus production of 2-methyl-1-propanol and 3-methyl-1-butanol; and P. aeruginosa production of hydrogen cyanide, 5-methyl-2-hexanamine, 5-methyl-2-heptanamine, 1-undecene, 3-methyl-1-butanol and 2-nonanone correlated with biofilm metabolic activity (r≥0.92; P≤0.01) and biomass (r≥0.98; P≤0.0004).
CONCLUSIONS - We conclude that ES has potential as an antimicrobial and VOC detection has prospective clinical translatability in the non-invasive monitoring of response to treatment; and human ex-vivo studies are currently underway.
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