WAMSI Node 5.4 - (PhD) Quorum quenching compounds from marine bacteria

Full description

Antibiotic resistance is of enormous concern in disease control and is one of the factors driving the search for new strategies for controlling bacterial growth. Quorum sensing (QS) is a bacterial communication process that allows control of gene expression in relation to cell density. Bacterial activities under QS control include the production of pigments and toxins, bioluminescence, swarming motility and biofilm formation. Quorum quenching (QQ) compounds inhibit the QS process, with the potential to control detrimental bacterial activities. QQ compounds have been isolated from several plants, algae, fungi and several bacteria. The aim of the project was to isolate and characterise QQ compounds from marine bacteria and investigating the mechanism by which the compound interferes with quorum sensing.

Objectives of project were:

(i)    to purify and identify quorum quenching compounds (QQCs) from marine invertebrates and bacteria;
(ii)    to assess the diversity of quorum quenching effects and mechanisms of action amongst QQCs.
(iii)    to investigate ecological aspects of the formation and activity of QQCs
(iv)    to assess potential applications of QQCs.

Lineage

Statement: Bioassay for detection of QQ bacteria: Natural marine samples were screened for the presence of bacteria with QQ activity using a bioluminescence inhibition assay developed specifically for the purpose. Bacterial isolation, culture and purification: Bacteria testing positive in the screening assay were isolated, purified and preserved using standard microbiological culturing techniques. Bacterial identification: Bacterial isolates were identified using exhaustive biochemical testing to establish phenotypic characteristics, volatile fatty acid analysis, and molecular techniques based on 16S RNA analysis of genomic DNA. Test results were compared with published data on bacterial phenotypes (phenotypic characters) and online BLAST sequences (molecular analysis). Known and unknown isolates were grouped using phylogenetic analysis. Confirmation of QQ activity and qualitative QQ assay: The purified bacterial isolates were assessed for QQ activity in a quantitative assay (based on spectrophotometric methods to detect both growth and bioluminescence, enabling assessment of relative bioluminescence inhibition. Extraction, purification and identification of QQCs: Using a bioassay-directed procedure the QQCs from selected bacterial isolates were purified and identified using a combination of chemical procedures including solvent extraction, HPLC, GC, NMR and MS. Assay of QQCs against additional quorum sensing processes: Purified QQCs from the selected bacterial isolates were screened for their effects in a range of QQ assays including inhibition of bioluminescence by Vibrio harveyi, pigment production by Serratia marcescens and Chromobacterium violaceum, and biofilm formation by Pseudomonas aeruginosa.