Validation Of Biofilm Formation On Human Wound Models And Confirmation Of Their Usability In Skin-relevant Biofilm Studies
Mohammed Ashrafi1, Lilyann Novak-Frazer1, Mohamed Baguneid2, Teresa Alonso-Rasgado1, Guoqing Xia1, Riina Rautemaa-Richardson1, Ardeshir Bayat1.
1The University of Manchester, Manchester, United Kingdom, 2Manchester University NHS Foundation Trust, Manchester, United Kingdom.
BACKGROUND - We previously presented novel formation of common wound bacterial biofilms on human ex-vivo cutaneous wound models and their bacterial-specific volatile organic compound (VOC) profiles. The aims here were to further validate bacterial biofilm formation in human ex-vivo incisional and excisional cutaneous wound models using a further bacterial strain along with its biofilm-forming deficient derivative and biomass assay for qualitative and quantitative comparisons, respectively.
METHODS - Explant viability was assessed using XTT assay. Staphylococcus aureus (MSSA), Pseudomonas aeruginosa (PA) and Streptococcus pyogenes (SP) biofilms, along with biofilm-deficient Staphylococcus aureus mutant (SA113ΔtagO) derived from the wild type Staphylococcus aureus strain (SA113) were formed on plastic, incisional and excisional cutaneous wound substrates. Biofilm development was further determined at day 0, 1, 3 and 5 using histological assessment, fluorescence microscopy and biomass (PicoGreen) assay. Metabolic activity (XTT) and VOC data used in the analyses have been previously presented.
RESULTS - There were no significant differences in explant viability in ex-vivo models confirming their suitability over the time period (P>0.05). MSSA, PA and SP biofilm comparisons to SA113 and SA113ΔtagO further confirmed biofilm development and maturity. There were significant inter-strain and inter-model variations in biomass at all time points (P<0.05). MSSA biofilm metabolic activity correlated with biomass across all biofilm models (R=-0.5). PA (R≥0.5) and SP (R=-1) biofilm metabolic activity correlated with biomass across the two cutaneous wound models. MSSA biomass correlated with 3-methyl-1-butanol (R=-1) across the two cutaneous wound models. PA biomass correlated with 3-methyl-1-butanol (R=-0.5) across the two cutaneous wound models and with hydrogen cyanide (R≥0.5), 2-methyl-1-propanol (R≤-0.5) and 5-methyl-2-heptanamine (R≥0.5) and 2-nonanone (R=0.5) across all three models.
CONCLUSIONS - Further validation of biofilm formation on human cutaneous wound models confirms that these models provide a vehicle for human skin-relevant biofilm studies and VOC detection has potential clinical translatability in efficient non-invasive wound infection diagnosis.
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