Naturwissenschaften

, Volume 100, Issue 3, pp 285–289

Postembryonic development of the bone-eating worm Osedax japonicus

Authors

    • Institute of BiogeosciencesJapan Agency for Marine-Earth Science and Technology
  • Tomoko Yamamoto
    • Faculty of FisheriesKagoshima University
  • Yoichi Yusa
    • Faculty of ScienceNara Women’s University
  • Yoshihiro Fujiwara
    • Institute of BiogeosciencesJapan Agency for Marine-Earth Science and Technology
Short Communication

DOI: 10.1007/s00114-013-1024-7

Cite this article as:
Miyamoto, N., Yamamoto, T., Yusa, Y. et al. Naturwissenschaften (2013) 100: 285. doi:10.1007/s00114-013-1024-7

Abstract

Bone-eating worms of the genus Osedax exclusively inhabit sunken vertebrate bones on the seafloor. The unique lifestyle and morphology of Osedax spp. have received much scientific attention, but the whole process of their development has not been observed. We herein report the postembryonic development and settlement of Osedax japonicus Fujikura et al. (Zool Sci 23:733–740, 2006). Fertilised eggs were spawned into the mucus of a female, and the larvae swam out from the mucus at the trochophore stage. Larvae survived for 10 days under laboratory conditions. The larvae settled on bones, elongated their bodies and crawled around on the bones. Then they secreted mucus to create a tube and the palps started to develop. The palps of O. japonicus arose from the prostomium, whereas the anterior appendages of other siboglinids arose from the peristomium. The recruitment of dwarf males was induced by rearing larvae with adult females. Females started to spawn eggs 6 weeks after settlement.

Keywords

AnnelidSiboglinidaeWhale boneSettlementDwarf male

Introduction

Bone-eating Osedax spp. are enigmatic marine worms that belong to the family Siboglinidae, phylum Annelida (Rouse et al. 2004). They exclusively inhabit sunken vertebrate bones (Rouse et al. 2004, 2011; Jones et al. 2008). Since their discovery in Monterey Bay in 2004 (Rouse et al. 2004), five species have been described and at least a further 12 species were reported based on genetic evidence (Rouse et al. 2004, 2008; Glover et al. 2005; Dahlgren et al. 2006; Fujikura et al. 2006; Braby et al. 2007; Vrijenhoek et al. 2009; Schander et al. 2010). Because of their unique biological features, Osedax worms have received much scientific attention. They show distinct sexual dimorphism with vermiform females and dwarf males living in the tube of the female (Rouse et al. 2004). It has been hypothesised that their sex is determined environmentally (Rouse et al. 2004). They lack a digestive system and a repeated segmental structure, although they belong to the phylum Annelida (Rouse et al. 2004). Osedax females host heterotrophic bacteria in a branching root system that invades the bone to extract organic compounds (Goffredi et al. 2005, 2007; Verna et al. 2010). From an ecological viewpoint, they show rapid dispersal and colonisation. For example, a time-series observation of whale carcasses in Monterey Bay revealed that exposed bones are colonised as rapidly as 2 months following their deposition (Braby et al. 2007). Osedax worms are sessile as adults and are presumed to disperse at embryonic and larval stages. To elucidate the evolution and nature of the unique biological features mentioned above, an understanding of their development and maturation is necessary. Although the early development of Osedax worms has been reported (Rouse et al. 2009), information about postembryonic development is completely lacking. We succeeded in inducing larval settlement under laboratory conditions and herein reported the postembryonic development and sexual maturation of this worm.

Materials and methods

Whale bones were collected at a depth of 226 m off Cape Noma via the remotely operated vehicle (ROV) Hyper-Dolphin on research vessel (R/V) Natsushimai on 28 March and 13 April 2012. Osedax japonicus were kept in 100-L aquaria at 11 °C in the laboratory. Embryos were collected from the mucus of females and reared in 200-mL dishes. Larval settlement was induced by the addition of small pieces of whale bone to the dishes. Juvenile worms living on the whale bone were kept in 3-L aquaria. Metamorphosis into dwarf males was induced by culturing larvae with females. To observe the juvenile worms, we dissected the bones under a stereomicroscope. Fluorescence in situ hybridisation was performed according to Fujinoki et al. (2012). More detailed methodology is described in the electronic supplementary material (ESM_1).

Results

Mature females continuously spawn eggs into the mucus attached to their tubes and the embryos develop in the mucus up to the trochophore stage (c. 3 days after spawning) as previously described (Fujikura et al. 2006). Larvae start to swim in the water column in this stage. They exhibit a metameric body comprising the prototroch, telotroch and apical tuft (Fig. 1a). Larvae of O. japonicus swim actively for at least 10 days if settlement is not induced. When the larvae were reared with pieces of whale bone, some larvae settled on the bones. Their bodies elongated and they ceased swimming and started crawling on the bones (Fig. 1b). The prototroch, gut and yolk were still present (Fig. 1b). They have two pairs of chaetae at the posterior end of the body (Fig. 1c). The chaetae probably function as an anchor when worms attach to substrates. Juvenile worms started to secrete mucus to form the tube 1 day post-settlement (dps; Fig. 1d). Yolk was not observed in this stage. Two ventral palps started to develop on the dorsal side of the prostomium or cephalic lobe. The palps arose anterior to the prototroch (Fig. 1e). The palps slightly elongated and the heart started to beat at 2 dps (Fig. 1e). No chaetae were observed at this stage in which bones attached to the root began to be digested and the root dug into the bones (Fig. 1f). The trunk and ventral palps elongated at 4 dps (Fig. 1g, h, i). Buds of dorsal palps were formed on the dorsal side of the ventral palps (Fig. 1h, arrowhead). Symbiotic bacteria were also detected in the root (Fig. 1m, n, o, p). This result indicates that the infection of symbionts occurred before this stage. Pinnules projected from the ventral palps at 7 dps (Fig. 1j, k). The dorsal palps were developing, but no pinnule was present at this stage. The projection of the oviduct formed between the dorsal palps (Fig. 1k). Juvenile worms have four palps with pinnules, an oviduct and a distinct root system, as mature females 10 dps (Fig. 1l).
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Fig. 1

Postembryonic development of O. japonicus. a A trochophore larva. Anterior is left. b An early juvenile. The prototroch is still present. Juveniles of this stage crawl on bones. Arrowheads indicate the posterior constrict. c Posterior end of the juvenile shown in b. Four chaetae are present at the posterior end. d Dorsolateral view of a 1-dps (day post-settlement) juvenile showing the developing ventral palps. e Lateral view of a 2-dps juvenile. Anterior is top and ventral is left. The heart starts to beat. Inset showing the location of the prototroch. f Two days post-settlement juvenile attaching to a bone. g Lateral view of a 4-dps juvenile. Anterior is top and ventral is left. h The anterior region in f showing the dorsal palp bud (arrowhead). i Ventral view of the anterior region of a 4-dps juvenile showing the pigmented collar. j, k Dorsal views of a 7-dps juvenile showing ventral palps with pinnules and developing dorsal palps. The projection of the oviduct was observed between the dorsal palps. l A 10-dps juvenile showing the same body plan with adult worm. mq Bright field, DAPI, EUB338, Sym732 and merged images of 4-dps juvenile showing symbiotic bacteria in the root. dp dorsal palp, dv dorsal vessel, h heart, mu mucus, p prostomium, pi pinnule, po projection of oviduct, pt prototroch, r root, vp ventral palp, y yolk

Two weeks after settlement, worms extend their palps and trunk to the outside of bones (Fig. 2a, b). We found that if many trochophore larvae were added to aquaria with females, some larvae settled on the tube of the females and metamorphosed into dwarf males (ESM_3). Dwarf males crawled on the tubes of females and some males stayed within the tubes. Six weeks after settlement, the length of female palps was about 5 mm, and females started to spawn eggs into the mucus (Fig. 2c, d). ‘Harems’ of dwarf males were observed within the tube of females 9 weeks after settlement (Fig. 2e). The time course of O. japonicus development is presented in ESM_4.
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Fig. 2

a A bone piece on which O. japonicus colonised 2 weeks after settlement. b High magnification of the bone in a. Worms did not spawn at this stage. Arrowheads in a and b indicate worms of O. japonicus. c A colony of O. japonicus 6 weeks after settlement. The same bone as a. d At this stage, females start to spawn (arrow). e Numerous dwarf males in the tube of a 9-week-old female (arrowheads)

Discussion

We succeeded in observing the whole developmental process of O. japonicus and revealed larval duration, acquisition of dwarf males and maturation time in this species. Our findings provide essential information on the reproductive strategy of Osedax worms. We show that female O. japonicus spawn fertilised eggs 6 weeks after settlement. The rapid sexual maturation of females, together with male dwarfism, enables them to reproduce effectively in the food-rich but highly isolated habitat of whale bones. The dispersal mode and time are also important in their ecology. The present observations show that larvae can drift at least 10 days. Further studies about tolerance to temperature and water pressure should provide essential information on the dispersal of this species. Our present observation that newly settled females did not host males until larvae were added to aquaria with females is not incompatible with the environmental sex determination proposed previously (Rouse et al. 2004, 2008). However, further studies addressing the cues for female and male settlement are necessary.

Postembryonic development of O. japonicus shows some similarities and differences from that of other siboglinid worms (reviewed in Southward 1999). The newly settled juveniles of siboglinids show elongated bodies with chaetae in their posterior parts (Southward 1999). At the following stage, the juvenile of O. japonicus has two palps. Although this form, at first glance, is similar to the juveniles of other siboglinids (Southward 1999; Nussbaumer et al. 2006), there is a striking difference in the topology of the anterior appendages. The palps of O. japonicus arise from the prostomium and the appendages of other siboglinids arise from the peristomium (Southward 1999; Rouse 2001; Nussbaumer et al. 2006). In adult vestimentiferans, because of the uncertain delimitation between the prostomium and peristomium, it is not clear where the palps arise (Rouse 2001). At least the larval branchial filaments of vestimentiferans, which arise behind the prototroch, are not homologous to the palps of Osedax (Jones and Gardiner 1989; Nussbaumer et al. 2006).

The relationships among body regions in siboglinids have been discussed (Rouse et al. 2008). Our results show that the trunk of O. japonicus develops from the region between the prototroch and the posterior constriction. This suggests that the trunk of Osedax is homologous to the vestimentum of vestimentiferans and to the forepart and the anterior end of other siboglinids. The ovisac and roots of O. japonicus develop from the posterior part of the juvenile. Our observation of O. japonicus development supports the recently defined dorsoventral axis of Osedax worms (Huusgaard et al. 2012). To elucidate the evolution of the Osedax body plan, further investigations of the anteroposterior patterning and tissue development will be necessary. The present study shows that O. japonicus is a promising model organism with which to address the evolution and development of Osedax worms.

Acknowledgments

We are grateful to the captain and crew of the R/V Natsushima and the operation team of the ROV Hyper-Dolphin for animal collection. We thank Thornton Blair at the University of Tokyo for organising a cruise and allowing us to use the rock cutter. This work is supported by Grants-in-Aid for Research Activity Start-up 23870044 to NM.

Supplementary material

114_2013_1024_MOESM1_ESM.pdf (7.8 mb)
ESM 1(PDF 7982 kb)

Copyright information

© Springer-Verlag Berlin Heidelberg 2013