Adv Biochem Engin/Biotechnol (2005) 97: 1–28 DOI 10.1007/b135821 © Springer-Verlag Berlin Heidelberg 2005 Published online: 8 August 2005 Aquaculture of “Non-Food Organisms” for Natural Substance Production Gerd Liebezeit Research Centre Terramare, Schleusenstrasse 1, 26382 Wilhelmshaven, Germany [email protected] 1Introduction................................... 2 2CultureAspects................................. 5 2.1 Medium..................................... 5 2.2 Food....................................... 6 2.3 Currents..................................... 7 2.4 LarvalProductionandSettlement....................... 7 2.5 Example Flustra foliacea ............................ 7 3Organisms.................................... 9 3.1 Porifera...................................... 9 3.2 Bryozoa...................................... 11 3.3 Molluscs..................................... 11 3.3.1Ophistobranchs................................. 12 3.4 Others...................................... 12 4 Applications Other than Pharmaceutical ................... 13 4.1 MarineCements................................. 13 4.2 Biominerals................................... 13 4.3 AntifoulingCompounds............................ 14 4.4 OtherApplications............................... 14 5 Interactions with Microautotrophs and -heterotrophs ............ 16 6 Further Considerations ............................. 17 7Conclusions................................... 18 References ....................................... 18 Abstract Marine invertebrates are already sources of commercially important secondary metabolites and may become even more so as knowledge on marine natural products and chemical ecology develops. Among the producers of these compounds predominantly sponges, bryozoa and molluscs have received the attention of academic and industrial research and development. For all these invertebrate groups culture techniques have been developed encompassing in situ, laboratory and cell culture approaches for the production of natural products. Potential applications of these are not restricted to phar- maceuticals but include marine cements, biominerals and antifouling compounds. In addition, markets exist for ornamental species. All culture approaches require sound eco- logical knowledge about the organisms to be cultured and possible symbiotic interactions between host invertebrates and microheterotrophs. 2 G. Liebezeit Keywords Aquaculture · Porifera · Bryozoa · Molluscs · Natural product 1 Introduction Invertebrates are defined as any animal lacking a backbone. The invertebrates include the tunicates and lancelets of the phylum Chordata, as well as all animal phyla other than Chordata including members of the phyla Porifera (sponges), Cnidaria (coelenterates), Ctenophora, Platyhelmintes (flatworms), Nematoda (roundworms), annelida (segmented worms), arthropoda, mol- lusca, echinodermata, endo- and ectoprocta and protochordates. Approxi- mately 95% of all the earth’s animal species are invertebrates; of these the vast majority are insects and other arthropods. Invertebrates are important as par- asites and are key players in all ecological communities, e.g. [1–6]. According to Brusca and Brusca [7] more than 151 000 species of invertebrates have been reported in the aquatic environment. Services and natural products from marine organisms have elicited con- siderable interest (Fig. 1), e.g. in cancer research and treatment [8–13]. These and other aspects of marine natural products have been reviewed by a.o. Baslow [14], Scheuer [15–19], Bohlin [20], Faulkner [21–51], Guyot [52], Cart [53], Olson [54], Abad and Bermejo [55], Blunden [56, 57], Proksch et al. [58–60] and Jha and Zi-rong [61]. Invertebrates provide the vast major- ity of active marine metabolites [25]. To further illustrate this point from 1969 to 1995 approximately 200 new patents were issued worldwide for marine- derived biochemicals with potential therapeutic activities. Between 1996 and 1999 the rate of discovery and patenting increased considerably with close Fig. 1 Diagram illustrating the services rendered by various classes of marine organisms to man Aquaculture of “Non-Food Organisms”for Natural Substance Production 3 to 100 new compounds patented in just these 3 years [62]. The rate of new discoveries will certainly be increasing in the future as marine biomedical research matures and more and more researchers and companies turn their attention to the seas [63–65]. Nevertheless, as mankind negatively influences the oceanic ecosystems through for example pollution, species introduction, overfishing and destruc- tive fishing methods, concerns have been expressed that opportunities to learn more about marine organisms and their commercial potential may be- come limited in the future [63, 66]. Thus, increased research and development efforts are necessary. Reviews of the natural product chemistry of bryozoa have been given by Cristophersen [67] and Blackman and Walls [68] while information on nat- ural products from Porifera can be found in Faulkner [23, 38, 41, 47], Sarma et al. [69], Proksch [59, 70], Engel and Pawlik [71], Guyot [72] as well as Kobayashi and Ishibashi [73]. While the first publications on chemical aspects of sponges date from 1882 onwards describing pigments, steroids, guani- dines, amines, and related compounds [74–81] an important break-through in the discovery of medicinal properties of natural products of sponge ori- gin was made with the isolation of sponge arabinose nucleosides such as spongouridine from Tethya crypta [82, 83]. A synthetic modification of this compound is now clinically used against Hodgkins lymphoma, acute myelo- cytic leukaemia, and the herpes virus [84] providing an early example of the applicability and commercial success of marine-derived pharmaceuticals. Molluscs and here especially ophistobranchs have also provoked consider- able interest. More than 400 compounds which might be of pharmacological value have been described in the literature [85, 86]. These include hypoten- sive agents, cardioactive substances, muscle relaxants, antibiotics, antiviral and antitumour agents. Toxins of marine snails are also of interest, espe- cially conotoxins in signal transmission research and due to their analgetic properties [87, 88]. Tuncates and other invertebrate phyla have received less attention [89–91]. Despite these considerable academic efforts, only a few metabolites found in marine invertebrates have so far entered into any commercial activities (Table 1). In addition, a few more metabolites are presently under clinical investiga- tions (Table 2, see also the compilation by Haefner [92]). Due to the wide-ranging potential applications of marine bioactive com- pounds (Tables 1,2) the need for a reliable and continuous supply of these and other compounds from marine invertebrates arises. Providing an adequate supply of raw material for marine pharmaceutical compounds has so far been solved by in situ collection of large quantities of invertebrates [113, 114] although limitations in available biomass [77–81] and its usually patchy dis- tribution may impede the permanent success of this approach. Furthermore, bioactive compounds are normally present in minor quantities, e.g. produc- 4 G. Liebezeit Table 1 Examples of commercially available compounds from marine invertebrates Product Source Application Ara-A Cryptotethya crypta,sponge antiviral Ara-C Cryptotethya crypta,sponge anticancer Manoalide Luffariella variabilis, sponge phospholipase-A inhibition Aequorin Aequora victoria, jellyfish bioluminescent calcium indicator Green Fluorescent Protein Aequora victoria, jellyfish reporter gene Resilience® (Estée Lauder) Pseudopterogorgia additive elisabethae,toskincreams gorgonian Table 2 Selected examples of compounds from marine invertebrates presently in clinical trials (see also [10, 12, 58, 93]) Compound Source References ecteinascidin 743 Ecteinascidia turbinata, tunicate [94–96] bryostatin-1 Bugula neritina, bryozoan [97–100] aplidine (dehydrodidemnin B) Aplidium albicans, tunicate [101, 102] kahalalide F Elysia rufescens, mollusc [103, 104]∗ didemnin B Trididemnum solidum, tunicate [105–107] discodermolide Discodermia dissoluta, sponge [108–112] ∗ This appears to be one example of dietary uptake of bioactive metabolites, see 5 tion of 1 g of ecteinascidin-743 or E-743, an antitumour agent from a tunicate, would require collection of about 1 ton of organism [115, 116]. Given the fact that annual demands for secondary metabolites from marine sources which have passed all clinical trials and are ready to enter the market will fall in the range of 1 to 5 kg [59] an amount impossible to obtain from natural sources, alternative techniques for obtaining these amounts are required. Thus, aquaculture, either under controlled or natural conditions, and chemical synthesis [117, 118] may develop into (commercially) attractive al- ternatives [113]. Furthermore, application of the latter techniques would also protect natural resources and biodiversity. Chemosynthesis is, however, both a challenging and difficult task as the complex molecular structure of marine metabolites usually gives rise to complex synthesis pathways and low yields. Marine peptides such as the conotoxins are a noticeable ex- ception and can be produced in virtually unlimited amounts [119]. Thus “marine organisms should probably be used as inspiration [for] and not as the source of the chemicals, [...]” (Faulkner in [120]). Hence, present ap- proaches attempt to characterise