Marine Biology

, Volume 152, Issue 3, pp 657–666 | Cite as

Larval metamorphosis of the mussel Mytilus galloprovincialis in response to Alteromonas sp. 1: evidence for two chemical cues?

  • Wei-Yang BaoEmail author
  • Jin-Long Yang
  • Cyril Glenn Satuito
  • Hitoshi Kitamura
Research Article


Bacterial isolates from multi-species biofilms were identified by 16S rDNA gene sequences and investigated for their inductive effects as monospecific biofilms on larval metamorphosis of Mytilus galloprovincialis. Alteromonas sp. 1 biofilm was found to have inductive activity, which increased with increasing cell density. The cue(s) of Alteromonas sp. 1 biofilm responsible for inducing larval metamorphosis was further investigated. Treatment of the biofilm with formalin, ethanol, heat or ultraviolet irradiation resulted in a significant reduction in the inductive activity of Alteromonas sp. 1, and the crude extract of surface-bound products of the biofilm showed no activity. These results indicated that if the cue was a surface-bound chemical cue, it was unstable, or susceptible to the treatments or the extraction process. On the other hand, the inductive activity of treated biofilms had a linear regression to the cell survival of bacteria, indicating a metabolically active biofilm was a requirement for larval metamorphosis. Conditioned water of the biofilm did not induce larvae to metamorphose. However, larval crawling behavior in the conditioned water was the same as that in the biofilm prior to larval metamorphosis, and significantly different to larval behavior in seawater. This indicated that a potential or partial waterborne cue existed, but remained inactive when alone. A synergistic effect of the conditioned water with formalin-fixed Alteromonas sp. 1 biofilm resulted in a significant increase in larval metamorphosis. Heat treatment and fractionation of the conditioned water demonstrated that the waterborne cue was heat-stable and <3,000 Da in molecular weight. Platinum-coating, Lentil Lectin and Wheat Germ Agglutinin treatments of the formalin-fixed biofilm significantly reduced its synergistic effect with the conditioned water, suggesting that a surface-bound cue was present on the biofilm and that the cue might be associated with the bacterial exopolysaccharide or glycoprotein. Evidence presented here suggests that two chemical cues derived from bacteria act synergistically on larval metamorphosis of Mytilus galloprovincialis.


Inductive Activity Alteromonas Larval Behavior Conditioned Water Larval Metamorphosis 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



The authors are grateful to Dr. K. Kanai of Nagasaki University for guidance in the identification of the bacteria and the staff of the Nagasaki Prefectural Institute of Fisheries for cooperation in the collection of adult mussels. The authors thank 2 anonymous reviewers for their helpful comments. The first author acknowledges Japanese Ministry of Education, Culture, Sports, Science and Technology for providing a scholarship during the study period. All experiments complied with the current Japanese laws.


  1. Bao WY, Satuito CG, Yang JL, Kitamura H (2007) Larval settlement and metamorphosis of the mussel Mytilus galloprovincialis in response to biofilms. Mar Biol 150:565–574CrossRefGoogle Scholar
  2. Cáceres-Martínez J, Robledo JA, Figueras A (1994) Settlement and post-larvae behavior of Mytilus galloprovincialis: field and laboratory experiments. Mar Ecol Prog Ser 112:107–117CrossRefGoogle Scholar
  3. Dahms HU, Dobretsov S, Qian PY (2004) The effect of bacterial and diatom biofilms on the settlement of the bryozoan Bugula neritina. J Exp Mar Biol Ecol 313:191–209CrossRefGoogle Scholar
  4. Fitt WK, Coon SL, Walch M, Weiner RM, Colwell RR, Bonar DB (1990) Settlement behavior and metamorphosis of oyster larvae (Crassostrea gigas) in response to bacterial supernatants. Mar Biol 106:389–394CrossRefGoogle Scholar
  5. García-Lavandeira M, Silva A, Abad M, Pazos AJ, Sánchez JL, Pérez-Parallé ML (2005) Effects of GABA and epinephrine on the settlement and metamorphosis of the larvae of four species of bivalve molluscs. J Exp Mar Biol Ecol 316:149–156CrossRefGoogle Scholar
  6. Huang S, Hadfield MG (2003) Composition and density of bacterial biofilms determine larval settlement of the polychaete Hydroides elegans. Mar Ecol Prog Ser 260:161–172CrossRefGoogle Scholar
  7. Huggett MJ, Williamson JE, de Nys R, Kjelleberg S, Steinberg PD (2006) Larval settlement of the common Australian sea urchin Heliocidaris erythrogramma in response to bacteria from the surface of coralline algae. Oecologia 149:604–619CrossRefGoogle Scholar
  8. Johnson CR, Sutton DC (1994) Bacteria on the surface of crustose coralline algae induce metamorphosis of the crown-of-thorns starfish Acanthaster planci. Mar Biol 120:305–310CrossRefGoogle Scholar
  9. Johnson CR, Muir DG, Reysenbach AL (1991) Characteristic bacteria associated with surfaces of coralline algae: a hypothesis for bacterial induction of marine invertebrate larvae. Mar Ecol Prog Ser 74:281–294CrossRefGoogle Scholar
  10. Khandeparker L, Anil AC, Raghukumar S (2003) Barnacle larval destination: piloting possibilities by bacteria and lectin interaction. J Exp Mar Biol Ecol 289:1–13CrossRefGoogle Scholar
  11. Kirchman D, Graham S, Reish D, Mitchell R (1982a) Bacteria induce settlement and metamorphosis of Janua (Dexiospira) brasiliensis Grube (Polychaeta: Spirorbidae). J Exp Mar Biol Ecol 56:153–163 CrossRefGoogle Scholar
  12. Kirchman D, Graham S, Reish D, Mitchell R (1982b) Lectins may mediate in the settlement and metamorphosis of Janua (Dexiospira) brasiliensis Grube (Polychaeta: Spirorbidae). Mar Biol Lett 3:131–142Google Scholar
  13. Kogure K, Simidu U, Taga N (1979) A tentative direct microscopic method for counting living marine bacteria. Can J Microbiol 25:415–420CrossRefGoogle Scholar
  14. Kogure K, Simidu U, Taga N (1984) An improved direct viable count method for aquatic bacteria. Arch Hydrobiol 102:117–122Google Scholar
  15. Lau SCK, Qian PY (2001) Larval settlement in the serpulid polychaete Hydroides elegans in response to bacterial films: an investigation of the nature of putative larval settlement cue. Mar Biol 138:321–328 CrossRefGoogle Scholar
  16. Lau SCK, Harder T, Qian PY (2003a) Induction of larval settlement in the serpulid polychaete Hydroides elegans (Haswell): role of bacterial extracellular polymers. Biofouling 19(3):197–204CrossRefGoogle Scholar
  17. Lau SCK, Thiyagarajan V, Qian PY (2003b) The bioactivity of bacterial isolates in Hong Kong waters for the inhibition of barnacle (Balanus amphitrite Darwin) settlement. J Exp Mar Biol Ecol 282:43–60CrossRefGoogle Scholar
  18. Leitz T, Wagner T (1993) The marine bacterium Alteromonas espejiana induces metamorphosis of the hydroid Hydractinia echinata. Mar Biol 115:173–178CrossRefGoogle Scholar
  19. Maki JS, Mitchell R (1985) Involvement of lectins in the settlement and metamorphosis of marine invertebrate larvae. Bull Mar Sci 37:675–683Google Scholar
  20. Maki JS, Rittschof D, Schmidt AR, Snyder AG, Mitchell R (1989) Factors controlling attachment of bryozoan larvae: a comparison of bacterial films and unfilmed surfaces. Biol Bull 177:295–302CrossRefGoogle Scholar
  21. Negri AP, Webster NS, Hill RT, Heyward AJ (2001) Metamorphosis of broadcast spawning corals in response to bacteria isolated from crustose algae. Mar Ecol Prog Ser 223:121–131CrossRefGoogle Scholar
  22. Neumann R (1979) Bacterial induction of settlement and metamorphosis in the planula larvae of Cassiopea andromeda (Cnidaria: Scyphozoa, Rhizostomeae). Mar Ecol Prog Ser 1:21–28 CrossRefGoogle Scholar
  23. Olivier F, Tremblay R, Bourget E, Rittschof D (2000) Barnacle settlement: field experiments on the influence of larval supply, tidal level, biofilm quality and age on Balanus amphitrite cyprids. Mar Ecol Prog Ser 199:185–204CrossRefGoogle Scholar
  24. Rahim SAKA, Li JY, Kitamura H (2004) Larval metamorphosis of the sea urchins, Pseudocentrotus depressus and Anthocidaris crassispina in response to microbial films. Mar Biol 144:71–78CrossRefGoogle Scholar
  25. Ramírez SC, Cáceres-Martínez J (1999) Settlement of the blue mussel Mytilus galloprovincialis Larmark on artificial substrates in Bahía de Todos Santos B.C., México. J Shellfish Res 18(1):33–39Google Scholar
  26. Rittschof D (1990) Peptide-mediated behaviors in marine organisms: evidence for a common theme. J Chem Ecol 16(1):261–272CrossRefGoogle Scholar
  27. Satuito CG, Natoyama K, Yamazaki M, Fusetani N (1995) Induction of attachment and metamorphosis of laboratory cultured mussel Mytilus edulis galloprovincialis larvae by biofilm. Fisheries Sci 61(2):223–227CrossRefGoogle Scholar
  28. Satuito CG, Natoyama K, Yamazaki M, Shimizu K, Fusetani N (1999) Induction of metamorphosis in the pediveliger larvae of the mussel Mytilus galloprovincialis by neuroactive compounds. Fisheries Sci 65(3):384–389CrossRefGoogle Scholar
  29. Satuito CG, Bao WY, Yang JL, Kitamura H (2005) Survival, growth, settlement and metamorphosis of refrigerated larvae of the mussel Mytilus galloprovincialis Lamarck and their use in settlement and antifouling bioassays. Biofouling 21(3/4):217–225CrossRefGoogle Scholar
  30. Steinberg PD, de Nys R, Kjelleberg S (2001) Chemical mediation of surface colonization. In: McClintock JB, Baker JB (eds) Marine chemical ecology. CRC Press, Boca Raton, pp 431–460Google Scholar
  31. Szewzyk U, Holmström C, Wrangstadh M, Samuelsson MO, Maki JS, Kjelleberg S (1991) Relevance of the exopolysaccharide of marine Pseudomonas sp. Strain S9 for the attachment of Ciona intestinalis larvae. Mar Ecol Prog Ser 75:259–265CrossRefGoogle Scholar
  32. Unabia CRC, Hadfield MG (1999) Role of bacteria in larval settlement and metamorphosis of the polychaete Hydroides elegans. Mar Biol 133:55–64CrossRefGoogle Scholar
  33. Weiner RM, Walch M, Labare MP, Bonar DB, Colwell RR (1989) Effect of biofilms of the marine bacterium Alteromonas colwelliana (LST) on set of the oysters Crassostrea gigas (Thunberg, 1793) and C. virginica (Gmelin, 1791). J Shellfish Res 8(1):117–123Google Scholar
  34. Wieczorek SK, Clare AS, Todd CD (1995) Inhibitory and facilitatory effect of biofilms on settlement of Balanus amphitrite larvae. Mar Ecol Prog Ser 119:221–228CrossRefGoogle Scholar
  35. Zimmer-Faust RK, Tamburri MN (1994) Chemical identity and ecological implications of a waterborne, larval settlement cue. Limnol Oceanogr 39(5):1075–1087CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Wei-Yang Bao
    • 1
    Email author
  • Jin-Long Yang
    • 1
  • Cyril Glenn Satuito
    • 1
  • Hitoshi Kitamura
    • 2
  1. 1.Graduate School of Science and TechnologyNagasaki UniversityNagasakiJapan
  2. 2.Faculty of FisheriesNagasaki UniversityNagasakiJapan

Personalised recommendations