Identification and accession of novel sources of microbial natural product diversity

This thesis reports attempts to improve the identification and accession of novel and understudied sources of microbial natural product chemical diversity. The cryptophycin-1 pathway from an understudied source, the genus Nostoc, constitutes an exceptional case to view natural chemical diversity, as more than thirty cryptophycins are made from the cryptophycin biosynthetic pathway. In this thesis the Nostoc polyketide (PK) and nonribosomal peptide (NRP) gene cluster for cryptophycin-1 from the lichen symbiont Nostoc sp. ATCC 53789 was identified through a subtractive cloning approach using driver sequences for PK and NRP gene clusters from the cycad symbiont, Nostoc punctiforme ATCC 29133. The cloned and sequenced portion (33 kb) of the cryptophycin-1 gene cluster provides insights on how cryptophycin-1 and the other derivatives are made from this one pathway. The cryptophycin-1 gene cluster was also localized to a 175 kb megaplasmid. This localization and identification of alien genes within the gene cluster provides evidence concerning the evolution of cryptophycin-1 evolved from its likely ancestor, the marine natural product arenastatin-A.; A broader view of Nostoc PK and NRP chemical diversity and variation of the PK and NRP biosynthetic pathways from different ecotypes (free-living and symbiotic) was made possible by analysis (comparative and in silico PK and NRP pathway reconstructions) of previously sequenced Nostoc genomes (Nostoc sp. PCC 7120 and Nostoc punctiforme ATCC 29133). Comparative analyses indicated that symbionts share many PK and NRP pathways and overall can be viewed as a larger source of chemical diversity.; The second aim of this thesis work was to culture previously uncultured marine microorganisms which are rich in bioactive compounds. In these studies a novel selective enrichment technique was developed for marine sediments. Using this technique resulted in access to a totally novel group of actinobacteria (PNG 1 clade). A complete polyphasic taxonomic characterization of members of the PNG 1 clade is presented here and show the PNG 1 clade is a new genus (Solwaraspora). Not only are these strains novel they also constitute a new rich source of bioactive compounds.