Marine Microbial Resources
The ocean and its living resources are a treasure box for biotechnology and new medicine. It has so far not been systematically explored for its microbial biodiversity and only in recent years serious attempts have been to systematically explore its resources. Marine organisms are a particularly rich source of new bioactive compounds. This has initiated the intensive exploration of marine organisms, including relatively recently also from marine microorganisms (Imhoff et al., 2011). There is a continued and urgent need for new drugs for the treatment of severe human diseases such as cancer, inflammatory disorders, and neurodegenerative, metabolic and infectious diseases. In addition, the pipelines for new drugs have faded during the past few decades, primarily due to the decreasing research efforts of the pharmaceutical industry. Among other issues these consideration gave strong support for the initiation of the KiWiZ project in which basic research on marine natural products and microbial interactions is combined with the search for new bioactive compounds and the strong desire to promote suitable candidates thereof into application (see chapters “Overarching Research Activities” and “Applied Research”).
We have studied communities of microorganisms associated with various marine macroorganisms, in particular those associated with sponges, macroalgae, bryozoa and corals were intensively studied by us both by genetic analysis with the 16S rRNA gene as marker molecule and by pure culture investigations. Our studies revealed that the bacteria and fungi associated with these macroorganisms provide a rich source of bioactive strains (e.g. Schneemann et al., 2010; Heindl et al., 2010; Goecke et al., 2013). Fungal strains derived from marine sponges produced an immense variety of different bioactive substances (see research highlight III). A sponge-associated bacterium was shown to be the producer of the mayamycin exhibiting antibiotic activity against methicillin-resistant Staphylococcus aureus (MRSA) (Schneemann et al., 2010). A number of new bioactive compounds were identified in isolates from marine sediment organisms. Among them are eutypoids B-E, produced by marine fungi, which are inhibitors of the glycogen synthase kinase 3β (Schulz et al., 2011), the geranylphenazinediol showing promising inhibitory activity against the enzyme acetylcholinesterase (Ohlendorf et al., 2012) and the novel cytotoxic macrolide levantilide A and B produced by a deep-sea Micromonospora strain (Gärtner et al., 2011).
Genetic approaches to detect pathways for natural product biosynthesis
Genetic approaches for the detection of secondary metabolite pathways are promising tools for the selection of biosynthetically talented microorganisms. Main targets in this respect were genes encoding for polyketide synthases (PKSs) or non-ribosomal peptide synthetases (NRPSs), which are involved in two prominent biosynthetic pathways for natural products. In addition to these groups phenazines turned out to be good drug candidates and therefore are promising secondary metabolites. They are heterocyclic, nitrogenous compounds that are substituted at different sites of the core ring system and therefore display a wide range of structural derivatives and biological activities. A fast and efficient approach was established to identify bacteria possessing the potential to biosynthesize phenazines. Sequences of the phzE gene, which codes for one of the enzymes of the phenazine biosynthetic pathway, were used to design a universal primer system and to analyse the ability of bacteria to produce phenazines (Schneemann et al., 2011).
Genomic approaches offer great chances in the discovery of new natural products
During the past years, we have initiated several genome projects of marine bacteria and fungi to unravel capacities for natural product biosynthesis and to study their genetic capabilities in detail. Genomes of five bacteria have been established, two of these within a project on antibacterial peptides, and genomes of three marine fungi are currently under investigation within the EU-project “Marine Fungi” (coordinated by MI) in order to unravel their potential for secondary metabolite biosynthesis.
One of the bacteria selected for genome analysis was identified as a Bacillus subtilis strain. Bacillus subtilis is a model organism of aerobic spore-forming Gram-positive bacteria and is of great industrial significance as the source of natural antibiotic peptides as well as diverse functional molecules. The genome sequence of the marine isolate of Bacillus subtilis gtP20b contains a subset of candidate genes and gene clusters, which are potentially involved in production of ribosomal and non-ribosomal antimicrobial peptides, demonstrating the great potential of this strain as a source for novel antimicrobial peptides. There is significant variation at both DNA- and amino acid level of the genes when compared with those of other B. subtilis strains, suggesting that the investigated B. subtilis strain is a unique source for antimicrobial peptides. At least 59 genes were found to be involved potentially in bacterial secondary metabolism. Noticeably, a set of open reading frames (ORFs) was retrieved from antimicrobial peptides (AMPs) databases including subtilisin A (sboA), surfactin (sfp), beta-lactamase precursor (penP) and replicative DNA helicase (dnaC) with high sequence similarity (Fan et al., 2011).
Natural products application
In order to unravel the potential for application of natural products, their biological activity is of crucial importance. Therefore, a broad range of bioassays has been established to determine antibacterial, antifungal, antitumor and other activities that are specifically relevant for human diseases, protection against plant diseases or biofilm inhibition. For reproducible and sustainable production of natural substances, the microorganisms are preserved in the large culture collections, methods for cultivation and fermentation up to 250 Liter scale, chemical analytical methods for purification and substance identification, purification at the preparative scale for substance production, establishment and maintenance of a substance library are key methods available. These cover the whole range of methods required to reach from the source at the marine habitat to the pure substance product, available for external use and testing in preclinical studies. Approx. 1000 substances have been isolated and purified, most of which were included into the KiWiZ library of pure substances and quite a number of new substances have been published and/or are patented together with their activity profiles for the possible application. Among the new compounds identified at the KiWiZ since 2010 are the diacidine, a polyene dicarboxylic (Ohlendorf et al., 2012).
Three new bioactive compounds and their derivatives were selected according to the bioactivity profiles as potential relevant for application. These chemical structures, the production of the substances and their bioactivity spectrum relevant for application as cancer drugs or antibiotics have been patented at international levels.
The Kiel Center for marine natural products covers the work of RU MI in the field of marine natural compounds.
Head of the Research Unit:
Prof. Dr. Johannes F. Imhoff
GEOMAR | Helmholtz Centre for Ocean Research Kiel
Düsternbrooker Weg 20
Phone: ++49-431 600-4450
Fax: ++49-431 600-4482
Personal Assistant / Office Management:
Phone: ++49-431 600-4481
Fax: ++49-431 600-4482