Marine Biotechnology

, Volume 16, Issue 5, pp 594–603 | Cite as

Application of Diffusion Growth Chambers for the Cultivation of Marine Sponge-Associated Bacteria

  • Georg Steinert
  • Susanna Whitfield
  • Michael W. Taylor
  • Carsten Thoms
  • Peter J. Schupp
Original Article


Marine sponges contain dense and diverse microbial communities, which are renowned as a source of bioactive metabolites. The biological activities of sponge-microbe natural products span a broad spectrum, from antibacterial and antifungal to antitumor and antiviral applications. However, the potential of sponge-derived compounds has not been fully realized, due largely to the acknowledged “supply issue.” Most bacteria from environmental samples have resisted cultivation on artificial growth media, and cultivation of sponge-associated bacteria has been a major focus in the search for novel marine natural products. One approach to isolate so-called “uncultivable” microorganisms from different environments is the diffusion growth chamber method. Here, we describe the first application of diffusion growth chambers for the isolation of cultivable and previously uncultivated bacteria from sponges. The study was conducted by implanting diffusion growth chambers in the tissue of Rhabdastrella globostellata reef sponges. In total, 255 16S rRNA gene sequences were obtained, with phylogenetic analyses revealing their affiliations with the Alpha- and Gammaproteobacteria, Bacteroidetes, Actinobacteria, and Firmicutes. Fifteen sequences represented previously uncultivated bacteria belonging to the Bacteroidetes and Proteobacteria (Alpha and Gamma classes). Our results indicate that the diffusion growth chamber approach can be successfully applied in a natural, living marine environment such as sponges.


Cultivation-dependent Marine sponge Bacterial symbionts Diffusion growth chamber In vivo cultivation Rhabdastrella globostellata Sponge-specific cluster 16S rRNA 



PJS acknowledges funding by NIH MBRS SCORE grant S06-GM-44796. CT acknowledges the support by a Fedor-Lynen-Fellowship from the Alexander-von-Humboldt Foundation. We thank the University of Guam Marine Laboratory staff for assisting with field work. GS acknowledges the funding for phylogenetic analyses at the University of Auckland in the authors' laboratory by the German Academic Exchange Service (DAAD) short-term fellowship 'Microbial Symbiosis and Diversity in Marine Sponges' from 02/2013 to 06/2013.

Supplementary material

10126_2014_9575_MOESM1_ESM.jpg (486 kb)
Supplementary Fig. 1 16S rRNA-based phylogeny of R. globostellata-associated Gammaproteobacteria organisms. Details are the same as those provided in Fig. 3 (JPEG 485 kb)
10126_2014_9575_MOESM2_ESM.jpg (458 kb)
Supplementary Fig. 2 16S rRNA-based phylogeny of R. globostellata-associated Gammaproteobacteria organisms. Details are the same as those provided in Fig. 3 (JPEG 457 kb)
10126_2014_9575_MOESM3_ESM.jpg (599 kb)
Supplementary Fig. 3 16S rRNA-based phylogeny of R. globostellata-associated Alphaproteobacteria organisms. Details are the same as those provided in Fig. 3 (JPEG 599 kb)
10126_2014_9575_MOESM4_ESM.jpg (526 kb)
Supplementary Fig. 4 16S rRNA-based phylogeny of R. globostellata-associated Alphaproteobacteria organisms. Details are the same as those provided in Fig. 3 (JPEG 526 kb)
10126_2014_9575_MOESM5_ESM.jpg (784 kb)
Supplementary Fig. 5 16S rRNA-based phylogeny of R. globostellata-associated Actinobacteria organisms. Details are the same as those provided in Fig. 3 (JPEG 784 kb)
10126_2014_9575_MOESM6_ESM.jpg (467 kb)
Supplementary Fig. 6 16S rRNA-based phylogeny of R. globostellata-associated Firmicutes organisms. Details are the same as those provided in Fig. 3 (JPEG 466 kb)
10126_2014_9575_MOESM7_ESM.xlsx (91 kb)
Supplementary Table 1 Sequence information for all isolates. Novel cultured strains are highlighted in bold. It includes the GenBank accession number (ACC), the length of the sequence (long = >1199, short ≤1199), the isolate DGC source of the sequence (direct or DGC1 to DGC4), the different media types, and the GenBank blast results with the taxonomic classification from phylum to genus, the maximum identity and the related GenBank accession number. (XLSX 90 kb)


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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Georg Steinert
    • 1
  • Susanna Whitfield
    • 1
  • Michael W. Taylor
    • 2
  • Carsten Thoms
    • 3
  • Peter J. Schupp
    • 1
  1. 1.Institute for Chemistry and Biology of the Marine EnvironmentUniversity of OldenburgOldenburgGermany
  2. 2.Centre for Microbial Innovation, School of Biological SciencesUniversity of AucklandAucklandNew Zealand
  3. 3.Department of Marine Science & Technology, Faculty of Fisheries & Marine Sciences, Bogor Agricultural UniversityBogorIndonesia

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