Saudi Cultural Missions Theses & Dissertations
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Item Restricted Polymer based inhibition of Quroum sensing in Gram negative bacteria(2023-07-02) Alshalan, Rawan; Shepherd, JoeyBackground: Bacterial quorum sensing (QS) is involved in several pathogenic bacterial processes, such as production of tissue-destroying enzymes, pyocyanin, and rhamnolipids, and biofilm formation. QS -mediated infectious diseases caused by bacteria make QS an important target for the development of materials with a novel antibacterial mode of action. Aim: To evaluate the anti- QS ability of the polymer HB-PINPAM-HL, a hyperbranched poly(NIPAM) polymer with chain ends uniquely functionalised with homoserine lactone (HL), on two Gram-negative bacteria Chromobacterium violaceum CV026 and Pseudomonas aeruginosa PAO1. Method: The polymer HB-PNIPAM-HL was first assessed for inhibition of QS in the model organism Chromobacterium violaceum CV026, and then in the pathogen Pseudomonas aeruginosa PAO1 indirectly by measuring different virulence factors (VFs). Then, the differential gene expression of bacterial genes involved in QS and VFs pathways regulated by QS systems was directly assessed following treatment with HB-PNIPAM-HL and HBPNIPAM-succ, an intermediate polymer formed during the process of HB-PNIPAM-HL production . Further analyses of the effects of the polymers on human cells were performed, which included a cytotoxicity assay and a wound healing assay (migration assay). Subsequently, the anti-QS actions of the two polymers were tested in early (2h) and late (24h) P.aeruginosa infections in a tissue-based wound infection model. Results: HB-PNIPAM-HL at 128 μg/ml can significantly reduce production of the violacein pigment in C.violaceum CV026 and down-regulate the gene cviR, encoding the QS receptor, in addition to three genes directly encoding the violacein genes. HB-PNIPAM-HL incubated with P.aeruginosa at 256 μg/ml can significantly reduce the production of several VFs such as protease, elastase, pyocyanin, rhamnolipids and biofilm, anti-QS. There was no cytotoxic effect on either of the cell lines tested at up to 512 μg/ml. In the wound healing assay, there was no significant delay in cell migration and wound healing at up to 512 μg/ml of HBPNIPAM-HL, and 256 μg/ml of HB-PNIPAM -succ. In a three-dimensional tissue engineered human skin wound infection model infected with P.aeruginosa, application of HB-PNIPAM-HL at up to 512 μg/ml resulted in less tissue destruction when applied 2 hours after infection, but not when applied late, 24 hours after infection. Conclusion: HB- PNIPAM-HL and HB-PNIPAM -succ show promising anti- QS properties by reducing the virulence and pathogenicity of P.aeruginosa. These polymers could be an alternative to conventional antibiotics and a promising non-antibiotic therapy for wound infections.20 0