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SATURDAY - AAR-683 - Ajoene a First in Class Quorum Sensing Inhibitor that Prevents, Biofilm Formation, the Production of Virulence Factors, Spreading of Infections and Modulates Established Biofilms in an Ex-Vivo Chronic Wound Infection Model

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Author Block: D. M. J. Houston, D. Neef, P. Rice, D. Williams, H. Taleb, L. Jones, J. Preece, L. Sykes, W. W. Nichols, G. Dixon; Neem Biotech, Abertillery, United Kingdom
Disclosure Block: D.M.J. Houston: None. D. Neef: None. P. Rice: None. D. Williams: None. H. Taleb: None. L. Jones: None. J. Preece: None. L. Sykes: None. W.W. Nichols: None. G. Dixon: None.
Background: In chronically-infected wounds, the prevention of biofilm formation, the disruption of mature biofilms, the reduction of virulence factors and thus the spreading of infection remains clinically elusive. Quorum sensing (QS) pathways regulate microbial motility, virulence factor production and the formation and maturation of biofilms and thus present a potential mode of therapeutic intervention. Here we present the activity of ajoene, a QS Inhibitor, and its effects on motility, biofilm formation, virulence factor production and spreading infection, including activity on ex-vivo wound specific mature biofilm models. Methods: Ajoene effects on planktonic bacteria and biofilms of Pseudomonas aeruginosa (Pa) and Staphylococcus aureus (Sa) were tested using QS inhibition (Pa lasB-gfp % inhibition in liquid medium, and Sa spa-lacZ zone of inhibition in an agar-diffusion assay), biofilm formation (minimum biofilm inhibitory concentration, MBIC) and eradication assays (minimum biofilm eradication concentration, MBEC). The modulation of secretion of extracellular toxins in Sa and Pa was tested. The effect of ajoene on the motility of Pa was analysed via swimming and swarming assays. Ajoene was tested against novel biofilm skin and chronic wound spreading infection models using a mature Pa + Sa biofilm consortium. Results: At concentrations that did not inhibit growth, ajoene inhibited QS in Pa (80% at 75 µM) and Sa (33 mm zone of inhibition at 3 mM) and inhibited the formation of biofilms of Pa (IC50 31 ± 10 µM) and Sa (1 ± 0.5 µM). Pre-treatment with ajoene enhanced the susceptibility of Pa and Sa to both tobramycin and iodine in vitro. Ajoene reduced the secretion hemolysin in Sa of lasB, RL, pyocyanin and proteases in Pa. Ajoene reduced microbial viability in a biofilm infection model of ex-vivo porcine skin (>5 log reduction of colony forming units of both Pa and Sa). A clinically relevant ajoene-loaded hydrogel prevented the spread of bacteria from the initial inoculation site in the chronic wound ex vivo model. Conclusion:Ajoene, a first in class QS inhibitor, reduced secretion of virulence factors, inhibited the spread of infection and destabilised mature biofilms of Pa and Sa, re-sensitizing them to antibacterial agents. Ajoene disrupted the contamination-infection continuum of an ex-vivo skin wounds and could potentially aid in the maintenance or return of chronic wounds to a healing state.