Phylogenetic Diversity of Bacteria Associated with the Marine Sponge Gelliodes carnosa Collected from the Hainan Island Coastal Waters of the South China Sea


Several molecular techniques were employed to document the bacterial diversity associated with the marine sponge Gelliodes carnosa. Cultivation-dependent and cultivation-independent methods were used to obtain the 16S rRNA gene sequences of the bacteria. Phylogenetic analysis based on the 16S rRNA gene sequences showed that the bacterial community structure was highly diverse with representatives of the high G + C Gram-positive bacteria, cyanobacteria, low G + C Gram-positive bacteria, and proteobacteria (α-, β-, and γ-), most of which were also found in other marine environments, including in association with other sponges. Overall, 300 bacterial isolates were cultivated, and a total of 62 operational taxonomic units (OTUs) were identified from these isolates by restriction fragment length polymorphism (RFLP) analysis and DNA sequencing of the 16S rRNA genes. Approximately 1,000 16S rRNA gene clones were obtained by the cultivation-independent method. A total of 310 clones were randomly selected for RFLP analysis, from which 33 OTUs were acquired by further DNA sequencing and chimera checking. A total of 12 cultured OTUs (19.4% of the total cultured OTUs) and 13 uncultured OTUs (39.4% of the total uncultured OTUs) had low sequence identity (≤97%) with their closest matches in GenBank and were probably new species. Our data provide strong evidence for the presence of a diverse variety of unidentified bacteria in the marine sponge G. carnosa. A relatively high proportion of the isolates exhibited antimicrobial activity, and the deferred antagonism assay showed that over half of the active isolates exhibited a much stronger bioactivity when grown on medium containing seawater. In addition to demonstrating that the sponge-associated bacteria could be a rich source of new biologically active natural products, the results may have ecological implications. This study expands our knowledge of the diversity of sponge-associated bacteria and contributes to the growing database of the bacterial communities within sponges.

This is a preview of subscription content, log in to check access.

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5


  1. 1.

    Agogué H, Joux F, Obernosterer I, Lebaron P (2005) Resistance of marine bacterioneuston to solar radiation. Appl Environ Microbiol 71:5282–5289

    Article  PubMed  PubMed Central  Google Scholar 

  2. 2.

    Amann RI, Ludwig W, Schleifer KH (1995) Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Rev 59:143–169

    CAS  PubMed  PubMed Central  Google Scholar 

  3. 3.

    Baker GC, Smith JJ, Cowan DA (2003) Review and re-analysis of domain-specific 16S primers. J Microbiol Methods 55:541–555

    CAS  Article  PubMed  Google Scholar 

  4. 4.

    Bhadury P, Mohammad BT, Wright PC (2006) The current status of natural products from marine fungi and their potential as anti-infective agents. J Ind Microbiol Biotechnol 33:325–337

    CAS  Article  PubMed  Google Scholar 

  5. 5.

    Carballo JL, Naranjo SA, Garciagomez JC (1996) Use of marine sponges as stress indicators in marine ecosystems at Algeciras Bay (southern Iberian Peninsula). Mar Ecol Prog Ser 135:109–122

    Article  Google Scholar 

  6. 6.

    Carballo JL, Naranjo S (2002) Environmental assessment of a large industrial marine complex based on a community of benthic filter-feeders. Mar Pollut Bull 44:605–610

    CAS  Article  PubMed  Google Scholar 

  7. 7.

    Cole JR, Chai B, Farris RJ, Wang Q, Kulam-Syed-Mohideen AS, McGarrell DM, Bandela AM, Cardenas E, Garrity GM, Tiedje JM (2007) The ribosomal database project (RDP-II): introducing myRDP space and quality controlled public data. Nucleic Acids Res 35:169–172

    Article  Google Scholar 

  8. 8.

    Lee OO, Lau SC, Qian PY (2006) Consistent bacterial community structure associated with the surface of the sponge Mycale adhaerens Bowerbank. Microb Ecol 52:693–707

    Article  PubMed  Google Scholar 

  9. 9.

    Du H, Jiao N, Hu Y, Zeng Y (2006) Diversity and distribution of pigmented heterotrophic bacteria in marine environments. FEMS Microbiol Ecol 57:92–105

    CAS  Article  PubMed  Google Scholar 

  10. 10.

    Elyakov GB, Kuznetsova T, Mikhailov VV, Maltsev II, Voinov VG, Fedoreyev SA (1991) Brominated diphenyl ethers from a marine bacterium associated with the sponge Dysidea sp. Experientia 47:632–633

    CAS  Article  Google Scholar 

  11. 11.

    Erin AG, William F, Paul RJ (2007) Phylogenetic diversity of Gram-positive bacteria cultured from marine sediments. Appl Environ Microbiol 73:3272–3282

    Article  Google Scholar 

  12. 12.

    Erwin PM, Thacker RW (2008) Cryptic diversity of the symbiotic cyanobacterium Synechococcus spongiarum among sponge hosts. Mol Ecol 17:2937–2947

    CAS  Article  PubMed  Google Scholar 

  13. 13.

    Faulkner DJ (2000) Marine natural products. Nat Prod Rep 17:7–55

    CAS  Article  PubMed  Google Scholar 

  14. 14.

    Friedrich AB, Fischer I, Proksch P, Hacker J, Hentschel U (2001) Temporal variation of the microbial community associated with the Mediterranean sponge Aplysina aerophoba. FEMS Microbiol Ecol 38:105–113

    CAS  Article  Google Scholar 

  15. 15.

    Gontang EA, Fenical W, Jensen PR (2007) Phylogenetic diversity of Gram-positive bacteria cultured from marine sediments. Appl Environ Microbiol 73:3272–3282

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Haefner B (2003) Drugs from the deep: marine natural products as drug candidates. Drug Discov Today 8:536–544

    CAS  Article  PubMed  Google Scholar 

  17. 17.

    Hansen MC, Tolker-Neilson T, Givskov M (1998) Biased 16S rDNA PCR amplication caused by interference from DNA flanking template region. FEMS Microbiol Ecol 26:141–149

    CAS  Article  Google Scholar 

  18. 18.

    Hentschel U, Fieseler L, Wehrl M, Gernert C, Steinert M, Hacker J, Horn M (2003) Microbial diversity of marine sponges. In: Müller WEG (ed) Sponges (Porifera). Springer, Berlin, pp 59–88

    Google Scholar 

  19. 19.

    Hirayama H, Sunamura M, Takai K, Nunoura T, Noguchi T, Oida H, Furushima Y, Yamamoto H, Oomori T, Horikoshi K (2007) Culture-dependent and -independent characterization of microbial communities associated with a shallow submarine hydrothermal system occurring within a coral reef off Taketomi Island, Japan. Appl Environ Microbiol 73:7642–7656

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  20. 20.

    Imamura N, Nishijima M, Adachi K, Sano H (1993) Novel antimycin antibiotics, urauchimycins A and B, produced by marine actinomycete. J Antibiot (Tokyo) 46:241–246

    CAS  Article  Google Scholar 

  21. 21.

    Ivanova EP, Onyshchenko OM, Christen R, Lysenko AM, Zhukova NV, Shevchenko LS, Kiprianova EA (2005) Marinomonas pontica sp. nov., isolated from the Black Sea. Int J Syst Evol Microbiol 55:275–279

    CAS  Article  PubMed  Google Scholar 

  22. 22.

    Izumi H, Gauthier ME, Degnan BM, Ng YK, Hewavitharana AK, Shaw PN, Fuerst JA (2010) Diversity of Mycobacterium species from marine sponges and their sensitivity to antagonism by sponge-derived rifamycin-synthesizing actinobacterium in the genus Salinispora. FEMS Microbiol Lett 313:33–40

    CAS  Article  PubMed  Google Scholar 

  23. 23.

    Jayatilake GS, Thornton MP, Leonard AC, Grimwade JE, Baker BJ (1996) Metabolites from an Antarctic sponge-associated bacterium, Pseudomonas aeruginosa. J Nat Prod 59:293–296

    CAS  Article  PubMed  Google Scholar 

  24. 24.

    Jensen PR, Fenical WF (1996) Marine bacterial diversity as a resource for novel microbial products. J Ind Microbiol Biotechnol 17:346–351

    CAS  Article  Google Scholar 

  25. 25.

    Kan J, Evans SE, Chen F, Suzuki MT (2008) Novel estuarine bacterioplankton in rRNA operon libraries from the Chesapeake Bay. Aquat Microb Ecol 51:55–56

    Article  Google Scholar 

  26. 26.

    Kennedy J, Marchest J (2007) Metagenomic approaches to exploit the biotechnological potential of the microbial consortia of marine sponges. Appl Microbiol Biotechnol 75:11–20

    CAS  Article  PubMed  Google Scholar 

  27. 27.

    Kennedy J, Codling CE, Jones BV, Dobson AD, Marchesi JR (2008) Diversity of microbes associated with the marine sponge, Haliclona simulans, isolated from Irish waters and identification of polyketide synthase genes from the sponge metagenome. Environ Microbiol 10:1888–1902

    CAS  Article  PubMed  Google Scholar 

  28. 28.

    Kennedy J, Baker P, Piper C, Cotter PD, Walsh M, Mooij MJ, Bourke MB, Rea MC, O'Connor PM, Ross RP, Hill C, O'Gara F, Marchesi JR, Dobson AD (2009) Isolation and analysis of bacteria with antimicrobial activities from the marine sponge Haliclona simulans collected from Irish waters. Mar Biotechnol 11:384–396

    CAS  Article  PubMed  Google Scholar 

  29. 29.

    Könneke M, Bernhard AE, de la Torre JR, Walker CB, Waterbury JB, Stahl DA (2005) Isolation of an autotrophic ammonia-oxidizing marine archaeon. Nature 437:543–546

    Article  PubMed  Google Scholar 

  30. 30.

    Levi C, Levi P (1965) Populations bacteriennes dans les eponges. J Microsc (Paris) 4:60

    Google Scholar 

  31. 31.

    Li Q, Wang G (2009) Diversity of fungal isolated from three Hawaiian marine sponges. Microbiol Res 164:233–241

    CAS  Article  PubMed  Google Scholar 

  32. 32.

    Li Z, Liu Y (2006) Marine sponge Craniella austrialiensis associated bacterial diversity revelation based on 16S rDNA library and biologically active Actinomycetes screening, phylogenetic analysis. Lett Appl Microbiol 43:410–416

    CAS  Article  PubMed  Google Scholar 

  33. 33.

    Liu WC, Li CQ, Zhu P, Yang JL, Cheng KD (2010) Phylogenetic diversity of culturable fungi associated with two sponges: Haliclona simulans and Gelliodes carnosa, collected from the Hainan Island coastal waters of the South China Sea. Fungal Divers 42:1–15

    Article  Google Scholar 

  34. 34.

    Maldonado LA, Stach JEM, Pathom-aree W, Ward AC, Bull AT, Goodfellow M (2005) Diversity of culturable actinobacteria in geographically widespread marine sediments. Antonie van Leeuwenhoek 87:11–18

    Article  PubMed  Google Scholar 

  35. 35.

    Michaud L, Di Cello F, Brilli M, Fani R, Lo Giudice A, Bruni V (2004) Biodiversity of cultivable psychrotrophic marine bacteria isolated from Terra Nova Bay (Ross Sea, Antarctica). FEMS Microbiol Lett 230:63–71

    CAS  Article  PubMed  Google Scholar 

  36. 36.

    Mincer TJ, Jensen PR, Kauffman CA, Fenical W (2002) Widespread and persistent populations of a major new marine actinomycete taxon in ocean sediments. Appl Environ Microbiol 68:5005–5011

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Muscholl-Silberborn A, Thiel V, Imhoff JF (2008) Abundance and bioactivity of cultured sponge-associated bacteria from the Mediterranean Sea. Microb Ecol 55:94–106

    Article  Google Scholar 

  38. 38.

    Nicolas GG, Frenette M, Lavoie MC (2010) Streptococcus salivarius mutants defective in mannose phosphotransferase systems show reduced sensitivity to mutacins I-T9 and R-3B. Can J Microbiol 56:692–696

    CAS  Article  PubMed  Google Scholar 

  39. 39.

    Nubel U, Garcia-Pichel F, Muyzer G (1997) PCR primers to amplify 16S rRNA genes from cyanobacteria. Appl Environ Microbiol 63:3327–3332

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Oren M, Steindler L, Ilan M (2005) Transmission, plasticity and molecular identification of cyanobacterial symbionts in the Red Sea sponge Diacarnus erythraenus. Mar Biol 148:35–41

    CAS  Article  Google Scholar 

  41. 41.

    Proksch P, Putz A, Ortlepp S, Kjer J, Bayer M (2010) Bioactive natural products from marine sponges and fungal endophytes. Phytochem Rev 9:475–489

    CAS  Article  Google Scholar 

  42. 42.

    Reiswig HM (1973) Water transport, respiration and energetics of 3 tropical marine sponges. J Exp Mar Biol Ecol 14:231–249

    Article  Google Scholar 

  43. 43.

    Santavy DL, Willenz P, Colwell RR (1990) Phenotypic study of bacteria associated with the Caribbean sclerosponge Ceratoporella nicholsoni. Appl Environ Microbiol 56:1750–1762

    CAS  PubMed  PubMed Central  Google Scholar 

  44. 44.

    Schmidt EW, Obraztsova AY, Davidson SK, Faulkner DJ, Haygood MG (2000) Identification of the antifungal peptide-containing symbiont of the marine sponge Theonella swinhoei as a novel δ-proteobacterium, “Candidatus Entotheonella palauensis”. Mar Biol 136:969–977

    CAS  Article  Google Scholar 

  45. 45.

    Sekar R, Mills DK, Remily ER, Voss JD, Richardson LL (2006) Microbial communities in the surface mucopolysaccharide layer and the black band microbial mat of black band-diseased Siderastrea siderea. Appl Environ Microbiol 72:5963–5973

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  46. 46.

    Sharp KH, Eam B, Faulkner DJ, Haygood MG (2007) Vertical transimission of diverse microbes in the tropical sponge Corticium sp. Appl Environ Microbiol 73:622–629

    CAS  Article  PubMed  Google Scholar 

  47. 47.

    Taylor MW, Radax R, Steger D, Wagner M (2007) Sponge-associated microorganisms: evolution, ecology, and biotechnological potential. Microbiol Mol Biol Rev 71:295–347

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  48. 48.

    Thiel V, Leininger S, Schmaljohann R, Brümmer F, Imhoff JF (2007) Sponge-specific bacterial associations of the Mediterranean sponge Chondrilla nucula (Demospongiae, Tetractinomorpha). Microb Ecol 54:101–111

    Article  PubMed  Google Scholar 

  49. 49.

    Usher KM, Kuo J, Fromont J, Sutton DC (2001) Vertical transmission of cyanobacterial symbionts in the marine sponge Chondrilla australiensis (Demospongiae). Hydrobiologia 461:15–23

    Article  Google Scholar 

  50. 50.

    Usher KM, Sutton DC, Toze S, Kuo J, Fromont J (2005) Intergenerational transmission of microbial symbionts in the marine sponge Chondrilla australiensis (Demospongiae). Mar Freshw Res 56:125–131

    Article  Google Scholar 

  51. 51.

    Vacelet J (1975) Electron microscope study of the association between bacteria and sponges of the genus Verongia (Dictyoceratida). J Microscope Biol Cell 23:271–288

    Google Scholar 

  52. 52.

    Vacelet J, Donadey C (1977) Electron microscope study of the association between some sponges and bacteria. J Exp Mar Ecol 30:301–314

    Article  Google Scholar 

  53. 53.

    Watanabe K, Teramoto M, Harayama S (1999) An outbreak of nonflocculating catabolic populations caused the breakdown of a phenol-digesting activated-sludge process. Appl Environ Microbiol 65:2813–2819

    CAS  PubMed  PubMed Central  Google Scholar 

  54. 54.

    Webster NS, Webb RI, Ridd MJ, Hill RT, Negri AP (2001) The effects of copper on the microbial community of a coral reef sponge. Environ Microbiol 3:19–31

    CAS  Article  PubMed  Google Scholar 

  55. 55.

    Webster NS, Taylor MW, Behnam F, Lucker S, Rattei T, Whalan S, Horn M, Wagner M (2010) Deep sequencing reveals exceptional diversity and modes of transmission for bacterial sponge symbionts. Environ Microbiol 12:2070–2082

    CAS  PubMed  PubMed Central  Google Scholar 

  56. 56.

    Zhang HT, Lee YK, ZhangW LHK (2006) Culturable actinobacteria from the marine sponge Hymeniacidon perleve: isolation and phylogenetic diversity by 16S rRNA gene-RFLP analysis. Antonie van Leeuwenhoek 90:159–169

    CAS  Article  PubMed  Google Scholar 

  57. 57.

    Zhang H, Zhang W, Jin Y, Jin M, Yu X (2008) A comparative study on the phylogenetic diversity of culturable actinobacteria isolated from five marine sponge species. Antonie van Leeuwenhoek 93:241–248

    CAS  Article  PubMed  Google Scholar 

  58. 58.

    Zhang R, Wang Y, Gu JD (2006) Identification of environmental plasmid-bearing Vibrio species isolated from polluted and pristine marine reserves of Hong Kong, and resistance to antibiotics and mercury. Antonie van Leeuwenhoek 89:307–315

    CAS  Article  PubMed  Google Scholar 

  59. 59.

    Zhu P, Li Q, Wang G (2008) Unique microbial signatures of the alien Hawaiian marine sponge Suberites zeteki. Microb Ecol 55:406–414

    Article  PubMed  Google Scholar 

Download references


This work was supported in part by the National Science and Technology Project of China (no. 2009ZX09301-003-4-1, 2009ZX09301-003-11-1). We thank Professor Jin-He Li at the Institute of Oceanology, Chinese Academy of Science, for the sponge identification and gratefully acknowledge the help of the personnel at Sanya Nanfan Sci-Tech Institute during the sampling of the sponge. We also thank Mr. Rui-shan Wang and Ms. Ruo-meng Song in our lab for assistance in the antimicrobial assay. Finally, we thank Ms. Amy Blass at Brigham and Women’s Hospital, Harvard Medical School, USA, for proofreading this manuscript.

Author information



Corresponding author

Correspondence to Ping Zhu.

Electronic Supplementary Materials

Below is the link to the electronic supplementary material.

Suppl. 1

Phylogenetic affiliation of the cultured isolates from the sponge G. carnosa (DOC 121 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Li, C., Liu, W., Zhu, P. et al. Phylogenetic Diversity of Bacteria Associated with the Marine Sponge Gelliodes carnosa Collected from the Hainan Island Coastal Waters of the South China Sea. Microb Ecol 62, 800 (2011).

Download citation


  • Sponge
  • Operational Taxonomic Unit
  • Marine Sponge
  • Restriction Fragment Length Polymorphism Analysis
  • Restriction Fragment Length Polymorphism Pattern