Abstract
[Truncated] The increasing occurrence of antibiotic resistance and the decline in the discovery of new antibiotics is fuelling the search for antibiotic alternatives, including compounds that inhibit bacterial quorum sensing. Quorum sensing is a chemical communication system used by bacteria to control the expression of genes in relation to cell density. Genes under quorum sensing control include those associated with virulence and biofilm formation, so compounds that inhibit quorum sensing, termed quorum quenching compounds, could potentially be used to inhibit bacterial disease.
This study sought to detect and investigate quorum quenching compounds produced by culturable marine bacteria from the Perth metropolitan area (Western Australia). This involved the development of a novel rapid and direct screening assay that facilitated selective isolation and purification of marine bacteria associated with the production of bioactive compounds. The assay showed that up to 46% of culturable bacteria present in Perth marine waters produced quorum quenching bioactivity. A total of 107 bacterial isolates inhibiting growth (18) or quorum sensing-regulated bioluminescence (89) of Vibrio harveyi Vh1 (Vh1) were obtained. The isolates were identified biochemically, and by 16S rDNA sequencing and cellular fatty acid content, as members of the Gram negative genera Enterovibrio, Flammeovirga, Halomonas, Marinomonas, Phaeobacter, Photobacterium, Proteus, Pseudoalteromonas, Reinekea and Vibrio, and the Gram positive genus Bacillus, and appear to include several novel species.
Two Pseudoalteromonas isolates (K1 and B2) showed strong bioluminescence inhibitory activity and were chosen for further study. Extracts of the growth media of these isolates inhibited bioluminescence by Vh1 and pigment production by Chromobacterium violaceum at concentrations that did not inhibit growth. Extracts of the isolates also inhibited growth and pigment production of Serratia marcescens. Confirmation that these bioactive compounds directly inhibited quorum sensing required purification of the bioactive components and retesting in pure form. The bioactive compounds produced by the isolates were extracted, purified by HPLC and identified using nuclear magnetic resonance spectroscopy and mass spectrometry. Two types of bioactive compounds were identified; diketopiperazines and desferrioxamine (DFO) siderophores. DFOs have not previously been reported as quorum quenching compounds and so were selected for further investigation. Commercially available DFO G, DFO E and DFO B inhibited Vh1 bioluminescence in a qualitative bioassay; DFO B was used in all further bioassays.
This study sought to detect and investigate quorum quenching compounds produced by culturable marine bacteria from the Perth metropolitan area (Western Australia). This involved the development of a novel rapid and direct screening assay that facilitated selective isolation and purification of marine bacteria associated with the production of bioactive compounds. The assay showed that up to 46% of culturable bacteria present in Perth marine waters produced quorum quenching bioactivity. A total of 107 bacterial isolates inhibiting growth (18) or quorum sensing-regulated bioluminescence (89) of Vibrio harveyi Vh1 (Vh1) were obtained. The isolates were identified biochemically, and by 16S rDNA sequencing and cellular fatty acid content, as members of the Gram negative genera Enterovibrio, Flammeovirga, Halomonas, Marinomonas, Phaeobacter, Photobacterium, Proteus, Pseudoalteromonas, Reinekea and Vibrio, and the Gram positive genus Bacillus, and appear to include several novel species.
Two Pseudoalteromonas isolates (K1 and B2) showed strong bioluminescence inhibitory activity and were chosen for further study. Extracts of the growth media of these isolates inhibited bioluminescence by Vh1 and pigment production by Chromobacterium violaceum at concentrations that did not inhibit growth. Extracts of the isolates also inhibited growth and pigment production of Serratia marcescens. Confirmation that these bioactive compounds directly inhibited quorum sensing required purification of the bioactive components and retesting in pure form. The bioactive compounds produced by the isolates were extracted, purified by HPLC and identified using nuclear magnetic resonance spectroscopy and mass spectrometry. Two types of bioactive compounds were identified; diketopiperazines and desferrioxamine (DFO) siderophores. DFOs have not previously been reported as quorum quenching compounds and so were selected for further investigation. Commercially available DFO G, DFO E and DFO B inhibited Vh1 bioluminescence in a qualitative bioassay; DFO B was used in all further bioassays.
| Original language | English |
|---|---|
| Qualification | Doctor of Philosophy |
| Publication status | Unpublished - 2012 |
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