Quorum sensing in Australian soft corals

Quorum Sensing (QS) has emerged as a potential pathway of interaction between eukaryotic host organisms and their associated microbial communities. QS is the term that has been applied to describe the indirect regulation of gene expression in many micro-organisms. This is achieved by the use of small diffusible molecules that enable coordination of gene expression across an entire population. The extracellular nature of these compounds also allows their interception, interference and mimicry. Consequently, the molecules that regulate QS, and those produced to interfere with QS, are of increasing interest for biotechnology products. This thesis targeted QS interference in soft coral holobionts, as many soft coral species commonly contain compounds with structural similarities to well-studied bacterial QS molecules (acyl homoserine lactones). The presence of QS activity within the soft coral holobiont was investigated in order to determine if there were chemical or taxonomic patterns to the QS interference capability. Extracts (across three polarity ranges) of fifteen species of soft coral from four different families were screened against two QS bacterial biosensors, Agrobacterium tumefaciens A136 and Chromobacterium violaceum CV026, for both induction and inhibition of QS. The QS interference observed in the soft coral species differed between the two biosensor strains and was not restricted to one family, but rather extended across taxonomic bounds. Bioassay-guided fractionation of these extracts revealed chemical patterns that related to the observed activity, particularly in the induction of QS. To investigate the source of the chemical patterns, cembranoid diterpenes from active fractions were purified and tested for QS interference activity to confirm their presence as one source of QS activity in soft coral holobionts. Interestingly, the form of this activity (induction or inhibition) was found to correlate with structural variability of the secondary oxygen ring of these compounds. The potential for non-cembranoid diterpene QS activity in soft corals was investigated with an extract of Nephthea chabroli that had previously demonstrated strong QS inductive activity in A. tumefaciens A136 but was not known to contain cembranoid diterpenes. Bioassay guided fractionation identified one QS compound as 17,22-dihydroxy-24-methylene cholesterol. Finally, bacteria within the mucosal layer of the soft coral species Sinularia flexibilis and Lobophytum compactum were cultured to investigate the potential for the previously observed QS interference of cembranoid diterpenes to have an in situ ecological role. Isolobophytolide, which is the dominant cembranoid diterpene of L. compactum and has demonstrated QS activity, was used as an optional selection agent in the isolation media for strains of this species. The isolated strains were identified by their 16s rRNA gene and screened for induction and inhibition of QS using their culture medium. The identity and QS activity of the isolates, varied between the host corals and with the presence of isolobophytolide. These results suggest that QS interference could be a mechanism of bacterial regulation in soft corals and that soft coral holobionts could be a good model system to explore QS interference. The identification of QS interference in cembranoid diterpenes demonstrates their potential for inter-kingdom interference with bacterial QS gene expression. In addition, the structure activity information revealed in these compounds may be transferable to other model systems and for the design of biotechnology products. The selective effect of isolobophytolide on the growth of QS bacterial strains in the mucosal layers of L. compactum is further evidence of the possible ecological role these compounds may play in influencing the composition of the associated bacterial community and their QS regulated gene expression.