Microbial biodiscovery: Exploring venomous animal associated microbes as sources of new chemical diversity

Abstract

Microorganisms produce a large number of medically relevant small molecules through unique biosynthetic mechanisms. These small molecules have been widely used to develop drugs to treat a range of infectious diseases, cancers and other pathologies. In their natural environment microorganisms use these small molecules for signalling or defence. As microorganisms have adapted to different environments, they have greatly expanded their chemical profile and thus the investigation of microorganisms from new environments has often led to the discovery of novel chemistry. Microbes often exist in association with higher eukaryotic hosts and are therefore exposed to unique environments and thus produce unique chemistry. The host organism can benefit by utilising microbial metabolites for their own defence against infections, predators, and for chemical signalling. Interestingly, some venomous animals such as puffer fish and the blue ringed octopus utilise toxins produced by associated microbes to defend against predators. However this chemistry remains largely unexplored for the vast majority of venomous animals. In this study, we investigated metabolites from microbes associated with venomous animals including spiders, centipedes, cone snails, scorpions and wasps. We cultivated microbes from 15 venomous animals to produce a library of ~360 strains. The microbe library was cultured under a range of conditions and the resultant solvent extracts were subjected to chemical, biological and genomic studies. This PhD thesis focuses on metabolites produced by three wasp-associated fungi, Aspergillus sp. (CMB-W031), Talaromyces sp. (CMB- W045) and Quambalaria sp. (CMB-W001) and two cone snail-derived bacteria, Streptomyces sp. (CMB-CS038 and CMB-CS150) and Bacillus sp. (CMB-CS100). Chapter 1 Reviews the current status of drug discovery and symbiotic microbial strains involved in producing bioactive metabolites Chapter 2 Describes our approach to venomous animal microbial cultivation and microbe library biological and chemical profiling Chapter 3 Describes the identification and structure elucidation of a new nitrated diketopiperazine, waspergillamide A from a wasp-derived fungi Aspergillus sp. (CMB-W031) Chapter 4 Describes the identification and structure elucidation of new siderophores and azaphilones from a wasp-derived fungi Talaromyces sp. (CMB-W045) ii Chapter 5 Describes the chemical investigation of several cone snail-derived Streptomyces sp. (CMB-CS038, CMB-CS150, CMB-CS138, CMB-CS143 and CMB-CS145) and their common biosynthetic capacity. Chapter 6 Describes the identification and characterization of a pheromone from the wasp-derived fungi Quambalaria sp. (CMB-W001) Chapter 7 Describes the identification of a novel biosynthetic gene cluster in the cone snail-derived Bacillus sp. (CMB-CS100), through in silico approaches Chapter 8 Conclusions and outlook Our studies showed that venomous animals are a rich source of fungi and bacteria with the capacity to produce diverse metabolites. Chemical and biological screening on microbes demonstrated their ability to synthesise bioactive components. The isolation of new metabolites from wasp-derived fungi reveals the unexplored potential of venomous animal associated microbes as a source of new chemistry. The discovery of an insect pheromone from a wasp-associated fungi and antifungal compounds from cone snail bacteria hints at their use as signalling molecules and as anti-infective agents respectively by their host organisms. To explore otherwise silent biosynthetic pathways, we sequenced and analysed the genome of a cone snail bacterium and identified a novel lantipeptide gene cluster whose expression in traditional culture conditions was validated by proteomics. Our results indicate this approach can also be applied to other isolates in combination with chemical approaches to discover and characterise new metabolites. In summary, this thesis highlights the potential of venomous animal-associated microbes as sources of new bioactive metabolites.