The biotechnological potential of deep sea sponges and their associated microbiome

Shallow water sponges are known to be a prolific source of bioactive compounds and interesting enzymes. In particular shallow water sponges from temperate and warm environments have been investigated in the last couple of decades due to their easy accessibility. Sponges have been shown to harbour dense microbial communities, which were subsequently identified to be the source of most of the isolated bioactive compounds and enzymes. Marine sponges are widespread in our oceans, the biggest interconnected habitat on our whole planet. Sponges can be found not only in shallow water regions but also in the deep sea. The deep sea, comprising approximately anything deeper than 200 m with respect to sea level, makes up an immense area of the oceans, keeping in mind that the mean depth of the oceans is 3800 m. The biodiversity of the deep sea is hard to assess as 95% of the oceans are hypothesized to be unexplored, in this respect it is interesting to note that we have send more people to the moon than to the Marianas Trench, the deepest part of the oceans. Nonetheless already a few deep sea studies have changed our perception from a supposedly very hostile living environment, due to the huge pressure, low temperature and absence of sunlight to a treasure trove of to date largely unexplored marine life, especially with hot spots for living beings and biological diversity like hydrothermal vents, sponge and coral gardens. The marine life in the deep sea has in millions of years adapted to the aforementioned conditions and is therefore believed to be considerably different from other environments, therefore novel or considerably different chemistry particularly with respect to small molecules and novel modes of action for enzymes of industrial interest and antimicrobial compounds can be expected. The study presented here aims to provide a better understanding of the microbiota of deep sea sponges via applying different next generation sequencing approaches (MiSeq, PacBio, 454 pyrosequencing) as well as standard marine cultivation methods and various enzyme activity assays. In chapter two the metagenomes of three different deep sea sponge species (Inflatella pellicula, Stelletta normani and Poecillastra compressa) have been investigated for their potential to encode conserved domains of polyketide synthases and non-ribosomal peptide synthetase clusters. These clusters are involved in the production of secondary metabolites that are 6 beneficial to the sponges as defence mechanism, but could also be used in the pharmacological industry as novel drug leads for example as anticancer or antimicrobial medicines. A huge number of potentially novel adenylation and especially ketosynthase domains were oberverd in the metagenome of the investigated sponge species. Sequence similarities to domains from gene clusters known to be involved in the production of different classes of antibiotics and other bioactive compounds including lipopeptides, glycopeptides, macrolides and hepatotoxins have been identified. The next chapter studies a common marine microbial isolate that can be retrieved from various marine sources. The Pseudoalteromonas spp. isolates described herein have all been isolated from deep sea sponges (Inflatella pellicula, Sericolophus hawaiicus and Poecillastra compressa). The isolates were studied with respect to their biotechnological potential, with a particular focus on their enzymatic activity profiles and their potential for cold adaptation. Furthermore the whole genomes of these isolates and two reference strains were compared with a particular focus on genes potentially involved in symbiosis and secondary metabolism. The isolated Pseudoalteromonas spp. were shown to be cold adapted and to express various enzymatic activities, with only one activity being truly cold active. The genome comparison revealed an open pan-genome for all investigated isolates, but no enrichment in symbiosis related genes in the sponge isolates was observed. Nonetheless all the isolates harboured a highly conserved bacteriocin gene cluster with a tetratricopeptide repeat domain, which can be involved in host-association. Chapter four describes the screening and characterization of a novel cold-active esterase found via a function-based screening of a metagenomic fosmid library of the deep sea sponge Stelletta normani. Besides the enzyme defining activity parameters, the esterase was compared to other lipolytic enzymes and in situ docking studies were performed. The newly described esterase is part of the type IV hormone sensitive lipase family and is to the best of our knowledge the first truly cold active esterase of this family. The esterase is most active at alkaline pH, mimicking seawater conditions and displays a wide range of halotolerance; coupled with its cold activity this enzyme is potentially desirable for industrial applications in bioremediation and production of biodiesel