Matrix comparison of isolation conditions for secondary metabolite producing marine sponge associated bacteria

Abstract

The discovery of novel secondary metabolites has declined significantly in recent years whereas there is a rise in the number of multi-drug resistant pathogens and other types of diseases. The decline in natural product discovery was due to high rediscovery of already known compounds and the costs in developing natural products. As a result pharmaceutical companies lost interest in investing in natural product discovery. However, there is a renewed interest in marine sponge associated microorganisms as a rich and untapped source of secondary metabolites. The objective of this study was to design a matrix to investigate the extent to which the One Strain-Many Compounds (OSMAC) approach applies to a collection of marine sponge isolates harvested from two South African marine sponge samples. Terminal restriction fragment length polymorphisms (T-RFLP) analysis was used to investigate and ascertain the two marine sponges which hosted the highest microbial diversities to be used for further culture-dependent studies. The culture-dependent studies, using 33 media which included liquid enrichment, heat treatments and antibiotic treatments, resulted in 400 sponge isolates from the two marine sponges Isodictya compressa and Higginsia bidentifera. Using antibacterial overlay assays, 31 dereplicated isolates showed antibacterial activity. Bioactivities were also exhibited against E. coli 1699 which is genetically engineered for resistance against 52 antibiotics which implies that some of the bioactive compounds could be novel. The 16S rRNA gene sequences revealed that the microbial phyla isolated from the marine sponges belonged to Actinobacteria, Firmicutes and Proteobacteria (Alphaproteobacteria and Gammaproteobacteria).Thirty isolates were selected for an OSMAC-based matrix study, 17 of which showed noantibacterial activities in preliminary screening. The application of the OSMAC approach using co-culture and 36 culture conditions resulted in 6 isolates showing antibacterial activities, three of which did not show activities in preliminary screening. One of these, a Bacillus pumilus isolated from I. compressa displayed antibacterial activity against 5 indicator strains whereas in preliminary screening it had not shown activity. The results show that marine sponges can host novel microbial species which may produce novel bioactive compounds. The results also confirm that traditional methods employing a single culture condition restricts the expression of some biosynthetic pathways of microorganisms and as a result many metabolites have yet to be identified.