Marine Biology

, Volume 151, Issue 4, pp 1417–1431 | Cite as

Influence of bacteria and diatoms in biofilms on metamorphosis of the marine slipper limpet Crepidula onyx

  • Jill Man-Ying Chiu
  • Vengatesen Thiyagarajan
  • Jan A. Pechenik
  • Oi-Shing Hung
  • Pei-Yuan QianEmail author
Research Article


Larvae of the slipper limpet Crepidula onyx metamorphose in response to marine biofilms. In this study, we investigated how the percentage of larval metamorphosis in this species was affected by biofilms that differed in certain attributes. To manipulate bacterial and diatom cell densities and community composition, we developed biofilms in the laboratory (1) at different temperatures (16, 23 and 30°C) and salinities (20, 27 and 34‰), (2) with or without addition of antibiotics, and (3) in the light or in the dark. We also allowed biofilms to develop at three field sites with different prevailing environmental conditions so as to generate biofilms with different, but natural, attributes. Bacterial and diatom community composition in biofilms were determined using a DNA fingerprinting technique and microscopic examination, respectively. The effects of biofilms on metamorphosis were investigated in laboratory assays. The percentage of larval metamorphosis correlated with bacterial and diatom cell densities in only one of the three experiments conducted, but was substantially affected by differences in bacterial and diatom community composition in all three experiments. It also appears that metamorphosis of C. onyx depends on the simultaneous presence of both bacterial and diatom communities in biofilms.


Bacterial Community Community Composition Onyx Bacterial Community Composition Diatom Community 
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.



This manuscript was greatly benefited by the comments and suggestions by two anonymous reviewers, especially those on the statistical analysis and interpretation. We also wish to thank Stanley Lau (National University of Singapore) for his advice on drafting this manuscript. We are grateful to Mandy Tsoi and Ki Tam for kindly assisting in the terminal-restriction fragment length polymorphism analysis and technical supports. This study was supported by the Area of excellence scheme of UGC (project#AoE/P–04/2004) and RGC grants (HKUST 6402/05M) to PY Qian.


  1. Anderson MJ (1995) Variations in biofilms colonizing artificial surfaces: seasonal effects and effects of grazers. Mar Biol 75:705–714Google Scholar
  2. Avelin SRM, Vitalina SRM, Rittschof D, Nagabhushanam R (1993) Bacterial–barnacle interaction: potential of using juncellins and antibiotics to alter structure of bacterial communities. J Chem Ecol 19:2155–2167CrossRefGoogle Scholar
  3. Bonar DB, Weiner RM, Colwell RR (1986) Microbial-invertebrate interactions and potential for biotechnology. Microb Ecol 12:101–110CrossRefGoogle Scholar
  4. Brandini FP, Da Silva ET, Pellizzari FM, Fonseca ALO, Fernandes LF (2001) Production and biomass accumulation of periphytic diatoms growing on glass slides during a 1-year cycle in a subtropical estuarine environment (Bay of Paranagua, southern Brazil). Mar Biol 138:163–171CrossRefGoogle Scholar
  5. Bryan PJ, Qian PY (1998) Induction of larval attachment and metamorphosis in the abalone Haliotis diversicolor (Reeve). J Exp Mar Biol Ecol 223:39–51CrossRefGoogle Scholar
  6. Clarke KR (1993) Non-parametric multivariate analyses of changes in community structure. Aust J Ecol 18:117–143CrossRefGoogle Scholar
  7. Clarke KR, Warwick RM (1994) Change in marine communities: an approach to statistical analysis and interpretation. Natural Environmental Research Council, PlymouthGoogle Scholar
  8. Coe WR (1935) Sexual phases in prosobranch mollusks of the genus Crepidula. Science 81:570–572CrossRefGoogle Scholar
  9. Daume S, Brand-Gardner S, Woelkerling W (1999) Preferential settlement of abalone larvae: diatom films vs. non-geniculate coralline red algae. Aquaculture 174:243–254CrossRefGoogle Scholar
  10. Ebert E, Houk J (1984) Elements and innovations in the cultivation of red abalone Haliotis diversicolor (Reeve). Aquaculture 39:375–392CrossRefGoogle Scholar
  11. Eilers H, Pernthaler J, Glockner FO, Amann R (2000) Culturability and in situ abundance of pelagic bacteria from the North Sea. Appl Environ Microbiol 66:3044–3051CrossRefGoogle Scholar
  12. Eyster LS, Pechenik JA (1988) Comparison of growth, respiration, and feeding of juvenile Crepidula fornicata (L.) following natural or KCl-triggered metamorphosis. J Exp Mar Biol Ecol 118:269–279CrossRefGoogle Scholar
  13. Griffith P, Shiah FK, Gloersen K, Ducklow HW, Fletcher M (1994) Activity and distribution of attached bacteria in Chesapeake Bay. Mar Ecol Prog Ser 108:1–10CrossRefGoogle Scholar
  14. Hadfield MG, Paul VJ (2001) Natural chemical cues for settlement and metamorphosis of marine invertebrate larvae. In: McClintock JB, Baker BJ (eds) Marine chemical ecology. CRC, Boca Raton Florida, pp 431–461CrossRefGoogle Scholar
  15. Harder T, Lam C, Qian PY (2002) Induction of larval settlement in the polychaete Hydroides elegans by marine biofilms: an investigation of monospecific diatom films as settlement cues. Mar Ecol Prog Ser 229:105–112CrossRefGoogle Scholar
  16. Holmstrom C, Kjelleberg S (2000) Bacterial interactions with marine fouling organisms. In: Evans LV (eds) Biofilms: recent advances in their study and control. Harwood Academic Publisher, Amsterdam, pp 101–115Google Scholar
  17. Holmstrom C, Rittschof D, Kjelleberg S (1992) Inhibition of settlement of larvae of Balanus amphitrite and Ciona intestinalis by a surface-colonizing bacterium. Appl Environ Microbiol 58:2111–2115PubMedPubMedCentralGoogle Scholar
  18. Huang ZG, Morton B, Yip YW (1984) The distribution and ecological and biological features of Crepidula onyx in Hong Kong. Acta Oceanol Sin 5:827–839Google Scholar
  19. Ito S, Kitamura H (1997) Induction of larval metamorphosis in the sea cucumber Stichopus japonicus by periphitic diatoms. Hydrobiologia 358:281–284CrossRefGoogle Scholar
  20. Kawamura T, Kikuchi S (1992) Effects of benthic diatoms on settlement and metamorphosis of abalone larvae. Suisanzoshoku 40:403–409Google Scholar
  21. Keough MJ, Raimondi PT (1995) Responses of settling invertebrate larvae to bioorganic films: effects of different types of films. J Exp Mar Biol Ecol 158:235–253CrossRefGoogle Scholar
  22. Keough MJ, Raimondi PT (1996) Responses of settling invertebrate larvae to bioorganic films: effects of large-scale variation in films. J Exp Mar Biol Ecol 207:59–78CrossRefGoogle Scholar
  23. Lam C, Harder T, Qian PY (2003) Induction of larval settlement in the polychaete Hydroides elegans by surface-associated settlement cues of marine benthic diatoms. Mar Ecol Prog Ser 263:83–92CrossRefGoogle Scholar
  24. Lau SCK, Thiyagarajan V, Cheung SCK, Qian PY (2005) A laboratory investigation of the role of bacterial community in biofilm as an indicator of local environmental conditions for the settling larvae of marine invertebrates. Aquat Microbial Ecol 38:41–51CrossRefGoogle Scholar
  25. Le Tourneux F, Bourget E (1988) Importance of physical and biological settlement cues used at different scales by the larvae of Semibalanus balanoides. Mar Biol 97:57–66CrossRefGoogle Scholar
  26. Lima GM, Pechenik JA (1985) The influence of temperature on growth rate and length of larval life of the gastropod Crepidula plana (Say). J Exp Mar Biol Ecol 90:55–71CrossRefGoogle Scholar
  27. Liu WT, Marsh TL, Cheng H, Forney LJ (1997) Characterization of microbial diversity by determining terminal restriction fragment length polymorphisms of genes encoding 16S rRNA. Appl Environ Microbiol 63:4516–4522PubMedPubMedCentralGoogle Scholar
  28. Maki JS, Rittschof D, Costlow JD, Mitchell R (1988) Inhibition of attachment of larval barnacles, Balanus amphitrite, by bacterial surface film. Mar Biol 97:199–206CrossRefGoogle Scholar
  29. Maki JS, Ding L, Stokes J, Kavouras JH, Rittschof D (2000) Substratum/bacterial interactions and larval attachment: Films and exopolysaccharides of Halomonas marina (ATCC 25374) and their effect on barnacle cyprid larvae, Balanus amphitrite Darwin. Biofouling 16:159–170CrossRefGoogle Scholar
  30. McGee BL, Targett NM (1989) Larval habitat selection in Crepidula (L.) and its effect on adult distribution patterns. J Exp Mar Biol Ecol 131:195–214CrossRefGoogle Scholar
  31. Miron G, Boudreau B, Bourget E (1999) Intertidal barnacle distribution: a case study using a multiple working hypothesis. Mar Ecol Prog Ser 189:205–219CrossRefGoogle Scholar
  32. Montiel YA, Chaparro OR, Segura CJ (2005) Changes in feeding mechanisms during early ontogeny in juveniles of Crepidula fecunda (Gastropoda: Calyptraeidae). Mar Biol 147:1333–1342CrossRefGoogle Scholar
  33. Navarro JM, Chaparro OR (2002) Grazing-filtration as feeding mechanisms in motile specimens of Crepidula fecunda (Gastropoda: Calyptraeidae). J Exp Mar Biol Ecol 270:111–122CrossRefGoogle Scholar
  34. 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
  35. Pawlik JR (1992) Chemical ecology of the settlement of benthic marine invertebrates. Oceanogr Mar Annu Rev 30:273–335Google Scholar
  36. Pearce CM, Scheibling RE (1991) Effects of macroalgae, microbial films and conspecifics on the induction of metamorphosis of the green sea urchin Strongylocentrotus droebrachiensis (Muller). J Exp Mar Biol Ecol 147:147–162CrossRefGoogle Scholar
  37. Pechenik JA (1999) On the advantages and disadvantages of larval stages in benthic marine invertebrate life cycles. Mar Ecol Prog Ser 177:269–297CrossRefGoogle Scholar
  38. Pechenik JA, Gee CC (1993) Onset of metamorphic competence in larvae of the gastropod Crepidula fornicata (L.), judged by a natural and an artificial cue. J Exp Mar Biol Ecol 167:59–72CrossRefGoogle Scholar
  39. Pechenik JA, Heyman WD (1987) Using KCl to determine size at competence for larvae of the marine gastropod Crepidula fornicata (L.). J Exp Mar Biol Ecol 112:27–38CrossRefGoogle Scholar
  40. Pechenik JA, Li W, Cochrane DE (2002) Timing is everything: the effects of putative dopamine antagonists on metamorphosis vary with larval age and experimental duration in the prosobranch gastropod Crepidula fornicata. Biol Bull 202:137–147CrossRefGoogle Scholar
  41. Qian PY, Thiyagarajan V, Lau SCK, Cheung SCK (2003) Relationship between bacterial community profile in biofilm and attachment of the acorn barnacle Balanus amphitrite. Aquat Microb Ecol 33:225–237CrossRefGoogle Scholar
  42. Quinn GP, Keough MJ (2002) Experimental design and data analysis for biologists. University Press, Cambridge, pp 199–201CrossRefGoogle Scholar
  43. Rittschof D, Branscomb ES, Costlow JD (1984) Settlement and behavior in relation to flow and surface in larval barnacles, Balanus amphitrite Darwin. J Exp Mar Biol Ecol 82:131–146CrossRefGoogle Scholar
  44. Slattery M (1992) Larval settlement and juvenile survival in the red abalone (Haliotis rufescens): an examination of inductive cues and substrate selection. Aquaculture 102:143–153CrossRefGoogle Scholar
  45. Sonak S, Bhosle NB (1995) Observations on biofilm bacteria isolated from aluminum panels immersed in estuarine waters. Biofouling 8:243–254CrossRefGoogle Scholar
  46. Steinberg PD, Nys RD, Kjelleberg S (2001) Chemical mediation of surface colonization. In: McClintock JB, Baker JB (eds) Marine chemical ecology. CRC, Boca Raton Florida, pp 355–387Google Scholar
  47. Strathmann RR, Branscomb ES, Vedder K (1981) Fatal errors in set as a cost of dispersal and the influence of intertidal flora on set of barnacles. Oecologia 48:13–18CrossRefGoogle Scholar
  48. Tani Y, Ito Y (1979) Effects of benthic diatoms on settlement and metamorphosis of the sea urchin, Pseudocentrotus depressus. Suisanzoshoku 27:148–150Google Scholar
  49. Thompson RC, Norton TA, Hawkins SJ (1998) The influence of epilithic microbial films on the settlement of Semibalanus balanoides cyprids—a comparison between laboratory and field experiments. Hydrobiologia 375:203–216CrossRefGoogle Scholar
  50. Thornton DCO, Dong LF, Underwood GJC, Nedwell DB (2002) Factors affecting microphytobenthic biomass, species composition and production in the Colne Estuary (UK). Aquat Microb Ecol 27:285–300CrossRefGoogle Scholar
  51. Thorson G (1950) Reproductive and larval ecology of marine bottom invertebrates. Biol Rev 25:1–45CrossRefGoogle Scholar
  52. Todd CD, Keough MJ (1994) Larval settlement in hard substratum epifaunal assemblages: a manipulative field study of the effects of substratum filming and the presence of incumbents. J Exp Mar Biol Ecol 181:159–187CrossRefGoogle Scholar
  53. Tomas CR (ed) (1996) Identifying marine diatoms and dinoflagellates. Academic, San DiegoGoogle Scholar
  54. Wieczorek SK, Todd CD (1998) Inhibition and facilitation of settlement of epifaunal marine invertebrate larvae by microbial biofilm cues. Biofouling 12:81–118CrossRefGoogle Scholar
  55. Wieczorek SK, Murray AWA, Todd CD (1996) Seasonal variation in the effects of hard substratum biofilming on settlement of marine invertebrate larvae. Biofouling 10:309–330CrossRefGoogle Scholar
  56. Zar JH (1999) Biostatistical analysis, 4th edn. Prentice Hall, Englewood CliffsGoogle Scholar
  57. Zhao B, Qian PY (2002) Larval settlement and metamorphosis in the slipper limpet Crepidula onyx (Sowerby) in response to conspecific cues and the cues from biofilm. J Exp Mar Biol Ecol 269:39–51CrossRefGoogle Scholar
  58. Zhao B, Qiu JW, Qian PY (2003) Effects of food availability on larval development in the slipper limpet Crepidula onyx (Sowerby). J Exp Mar Biol Ecol 294:219–233CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  • Jill Man-Ying Chiu
    • 1
  • Vengatesen Thiyagarajan
    • 1
  • Jan A. Pechenik
    • 2
  • Oi-Shing Hung
    • 1
  • Pei-Yuan Qian
    • 1
    Email author
  1. 1.Department of Biology/Coastal Marine LaboratoryThe Hong Kong University of Science and TechnologyHong Kong SARChina
  2. 2.Biology DepartmentTufts UniversityMedfordUSA

Personalised recommendations