larval settlement on small spatial scales is mediated by biotic and abiotic ... many marine polychaetes require specific cues to settle and metamorphose.
Hydrobiologia 402: 239–253, 1999. A.W.C. Dorresteijn & W. Westheide (eds), Reproductive Strategies and Developmental Patterns in Annelids. © 1999 Kluwer Academic Publishers. Printed in the Netherlands.
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Larval settlement of polychaetes Pei-Yuan Qian Department of Biology, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong Key words: larval settlement, chemical settlement cues, larval settlement inducers, juveniles, metamorphosis
Abstract Many benthic marine invertebrate species have a dispersive larval stage in their life histories. Larvae typically spend hours, weeks, or months developing in plankton before they become competent to settle and metamorphose. Recruitment to benthic populations depends on the numbers of competent larvae transported to sites and/or the interaction between larvae and the surface of substratum. While there is considerable evidence that on large spatial scales, the number of competent larvae transported to sites is determined primarily by hydrodynamics, success of larval settlement on small spatial scales is mediated by biotic and abiotic characteristics of substratum. Larvae of many marine polychaetes require specific cues to settle and metamorphose. Cues can originate from conspecific or congeneric individuals, microbial films, sympatric species, food items, or habitat. Larval settlement in an individual species can be controlled by a single cue or a mixture of cues. Larval settlement of multiple species can be mediated by a common cue or a mixture of cues. Although a variety of chemicals, including proteins, free fatty acids, polysaccharides, inorganic ions, and neurotransmitters, have been suggested as inducing larval settlement of marine polychaetes, few natural cues have been isolated and structurally identified.
Introduction Most benthic marine invertebrate species have dispersal larval stages in their life cycles (Thorson, 1950: Figure 1). Larvae spend weeks or months developing in plankton before they become competent to settle and metamorphose. Recruitment to benthic populations then depends on the numbers of competent larvae transported to appropriate sites (Roughgarden et al., 1988). Availability of competent larvae is affected by biological parameters such as adult reproductive output and reproductive cycles (Connell, 1985; Roughgarden et al., 1988), abundance of adult population (Davis et al., 1989), larval mortality (Thorson, 1950; Gaines & Roughgarden, 1985), and hydrodynamics such as water current and flow velocity (Butman, 1987). On large spatial scales, larvae of marine invertebrates in a water column behave as passive particles that are transported to a given site. Local current pattern, flow velocity, and, particularly, nearbottom flow dynamics play very important roles in larval retention and larval settlement (Hannan, 1984). It is becoming apparent that the process of larval settlement is a dynamic event. Larvae can reject
one site and select another for settlement. Such active site selection is often mediated by environmental cues (Wilson, 1932, 1937, 1948, 1952, 1955; DeSilva, 1962; Williams, 1964; Crisp, 1974; Woodin, 1986; Chia, 1989; Pawlik, 1992; and many other reviews). Therefore, interaction between competent larvae and the substratum determines the site of larval settlement on small spatial scales and may determine postsettlement mortality. This interaction can be affected by biological, physical or chemical parameters, such as community structures, presence or absence of natural inducers released by conspecific individuals, biofilms, prey species, or sympatric species. In the last three decades, a number of papers have reviewed larval settlement and metamorphosis of marine invertebrates (Meadows & Campbell, 1972; Crisp, 1974; Schroeder & Hermans, 1975; Eckelbarger, 1978; Chia & Rice, 1978; Hermans, 1978; Potswald, 1978; Keough & Downes, 1982; Connell, 1985; Hadfield, 1986; Jackson, 1986; Trapido-Rosenthal & Morse, 1986; Chia, 1989; Davis et al., 1989; Jensen & Morse, 1990; Morse, 1990; Morse, 1991; Woodin, 1991; Pawlik, 1992; Rodrîguez et al., 1993; Woodin et al., 1995). Few of these papers,
240 Larval settlement patterns in marine polychaetes
Figure 1. Summary of life history of sessile marine invertebrates. Modified from Chia (1974) and Crisp (1984).
however, reviewed specifically the larval settlement of marine polychaetes as a whole. In the papers cited above, the terminology relating to ‘larval settlement’ and ‘metamorphosis’ is discussed extensively. This paper will discuss only larval settlement of polychaetes, not the details of larval metamorphosis. ‘Settlement’ is the process by which a planktonic larva moves toward the substratum, explores, attaches to the substratum, and begins its benthic life. ‘Metamorphosis’ is the process by which a planktonic larva goes through morphological and physiological changes to complete the transition from planktonic larva to benthic juvenile. Metamorphosis can commence even before larval settlement or occur concurrently with larval settlement or right after larval attachment to the substratum (Fenaux & Pedrotti, 1988). The two different processes, larval settlement and larval metamorphosis in marine invertebrates may require different environmental cues (Chia, 1989). Larvae of many marine invertebrates settle and metamorphose in response to a very specific cue from a unique source such as a conspecific or prey item (Burke, 1983; reviewed by Pawlik 1992; Rodriguez et al., 1993). Others respond to cues originating from a variety of sources (Pearce & Scheibling, 1990, 1991). The location of a site inhabited by conspecific adults may possess both positive and negative characteristics. This paper reviews the general settlement patterns of polychaete larvae and discusses possible mechanisms involved in their formation. Common cues for larval settlement and metamorphosis of marine polychaetes are reviewed and some well-documented cases are briefly discussed.
Although post-settling differential mortality can give rise to aggregative distribution patterns (Commito, 1982; Woodin, 1985; Watzin, 1986; Luckenbach, 1987), aggregative larval settlement near or on conspecifics due to active substratum selection is the determinant factor in most cases. Gregarious larval settlement and metamorphosis in response to conspecfic cues leads to the patchy distribution of the adult population in Phragmatopoma californica (Sabellariidae) (Jensen & Morse, 1984, 1990; Jensen, 1992; Nishi, 1992; Morse et al., 1993), Spirorbis borealis (Mackay & Doyle, 1978), Spirobranchus polycerus var. augeneri (Marsden, 1991), Janua (Dexiospira) brasiliensis (Nelson, 1979), Hydroides dianthus (Scheltema et al. 1981; Toonen & Pawlik, 1994), Capitella spp. (Grassle & Butman, 1989), Hydroides elegans (Hadfield et al., 1994; Bryan et al., 1997a; Qiu & Qian, 1997), and many other species (Knight-Jones, 1951; Wilson, 1968, 1970, 1974; Eckelbarger, 1978) (see review by Burke, 1986). The advantages of gregariousness include: 1. ensuring location in a habitat that will support postlarval growth better than other locations or at least have less of a predation threat to juveniles; 2. increasing fertilization success for both internally fertilizing and freely spawning species; 3. increasing life-span and thus increasing fecundity, and 4. increasing resistance to crushing as indicated in aggreated Phragmatopoma californica (Barry, 1989; Thomas, 1996). Comparatively speaking, only limited studies have demonstrated random distribution in marine polychaetes such as Eupolymnia nebulosa (Bhaud, 1990b). This has also been related to random larval settlement (no larval settlement preference). In some cases, negative-individual interaction among newly-settled juveniles and among young juveniles and adults may distance the individuals from each other, resulting in an even distribution pattern (Woodin, 1976, 1985; Tamaki, 1985; Bhaud, 1990a).
Larval settlement in response to specific cues from conspecific organisms Larvae of many polychaetes are attracted to conspecifics as sites for settlement and metamorphosis (re-
241 viewed by Pawlik, 1992; Rodriguez et al., 1993). The most intriguing works last decade focused on inductive substances for larval settlement and metamorphosis of several reef-building, gregarious sabellariid polychaete species (Sabellariidae). Larvae of Phragmatopoma lapidosa californica attached and metamorphosed upon contact with conspecific tubes but often delayed metamorphosis in the absence of such contact (Jensen & Morse, 1984). Jesnsen & Morse (1984) proposed that DOPA residues were responsible for settlement of this species. Pawlik (1986) found however, that the stimulative substance for metamorphosis was inactivated by boiling and the bioactivity of stimulus was diminished by organic solvent extraction. The bioactive material purified by HPLC (High Performance Liquid Chromatography) and chemically characterized by NMR (Nuclear Magnetic Resonance) spectroscopy and GC (Gas Chromatography) consisted of a mixture of free fatty acids ranging from 14 to 22 carbon atoms in length (Pawlik, 1986). The active larval settlement inducers in another subspecies, P. l. lapidosa, from the Atlantic coast of North America, were also a mixture of free fatty acids (Pawlik, 1988a). The same set of free fatty acids was separated from the tube sand of both the California and Atlantic subspecies and induced larval settlement of both subspecies at about the same concentrations. These free fatty acids, however, did not induce larval settlement of Sabellaria alveolata, a reef-building sabellariid from European waters (Pawlik, 1988a). Although larvae of Sabellaria alveolata also settled in response to conspecific tube sand, bioactivity of larval settlement inducers from the tube sand was diminished by organic solvent extraction. Inductive compounds could not be isolated and purified (Pawlik, 1988a). Free fatty acids were found in the tube sand of S. alveolata at much lower concentrations than those found in the tube sand of P. l. lapidosa and P. l. californica. Free fatty acids did not induce larval settlement of non-gregarious sabellarid species such as Sabellaria floridensis from the Caribbean (Pawlik, 1988b) and Sabellaria cementarium from the eastern Pacific Ocean (Pawlik & Chia, 1991). Inducers involved in larval settlement of different species of gregarious sabellariids and larval response to free fatty acids is species-specific (Pawlik, 1992). Concurrent studies on larval settlement of P. lapidosa californica produced findings different from that of Pawlik (Jensen & Morse, 1984, 1990; Yool et al., 1986; Jensen et al., 1990). In these experiments, larvae of P. l. californica settled and metamorphosed readily upon contact with 2,6-di-(1,1-dimethyl)-4-methyl-
phenol (DBMP) (Jensen & Morse, 1984, 1990), which has no chemical similarity to DOPA (see Pawlik, 1990). Jensen et al. (1990) partially purified the natural inducer and concluded that the tube cement of this species is largely proteinaceous and highly resistant to deterioration. They believe that free fatty acids are not responsible for the natural gregarious attachment and metamorphosis of larvae in P. l. californica and P. l. lapidosa because: 1. free fatty acids were not detected on glass beads used by adult worms to build tubes in their experiments (the source of natural inducers defined in Pawlik’s studies); 2. freeze-drying and stirring reduced the inductive activity of the natural inducer but not of glass beads coated with free fatty acids; 3. induction by free fatty acids was temperaturedependent while induction by the natural inducer was not; and 4. induction by natural inducer was taxon-specific but induction by free fatty acids was not (Jensen et al., 1990). Pawlik (1992) argued that 1. replication of the experiments detailed in Jensen et al. (1990) failed to confirm that freeze-drying or stirring decrease the inductive activity of the natural inducer as compared with substrata coated with free fatty acids, or that induction by free fatty acids is temperature dependent; 2. the high degree of specificity of larval response within the Sabellariidae can not be ignored; 3. if free fatty acids are not the natural inducers of settlement, then why are larval responses to these compounds restricted to the genus Phragmatopoma within the family Sabellariidae and why do these compounds occur at high concentrations in the natural tube sand of species that respond to them but not in the tube sand of species that do not? and 4. if free fatty acids are absent from inductive, uncontaminated tube sand, where does the natural cue come from? Pawlik (1992) could not rule out of the possibility that the natural tube sand used in his experiment, however, might have been contaminated with organic material containing free fatty acids. There are more individual cases of chemosensory recognition by larvae of newly built conspecific tube material or presumptive chemical analogs of the tube cement in other sabellariid species. For example, lar-
242 vae of Sabellaria alveolata can be stimulated to settle upon contact with adult tubes, tube remnants, or the mucoid tubes of juvenile worms. Surface contour and roughness, sediment type, water motion, and presence of surface microorganisms have only a minor influence on larval settlement. The larval settlement inducers in the tubes were water insoluble and unaffected by drying, but were destroyed by cold concentrated acid (Wilson, 1968, 1970). Isolation and purification of inducers of this species have not been successful. Similarly, larvae of Polydora ciliata (Lagadeuc, 1991), gregarious serpulid Spirobranchus polycerus var. augeneri (Marsden, 1991), serpulid Pomatoceros lamarckii (Chan & Walker, 1998), and Capitella sp. I of the Capitella complex (Biggers & Laufer, 1992) settle in response to chemical substances originated from conspecific adults or juveniles. The inducers of these species have not been isolated or purified. Hydroides elegans is a gregarious tube-building polychaete that occurs in tropical and sub-tropical waters. Metamorphosis includes secretion of the primary tube, resorption of the prototroch cilia, formation of the collar which secretes the secondary, calcified tube, and emergence of the adult feeding tentacles. In less than 12 hours, larvae that have been exposed to cues can settle and metamorphose into juveniles residing in an attached calcareous tube (Hadfield, 1998). H. elegans populations from India undergo metamorphosis when competent, regardless of the presence of external cues (Mary et al., 1994) but larvae from Hawaii and Hong Kong do not settle until provided with appropriate environmental cue(s) and settle on aged biofilms preferentially over unfilmed surfaces (Hadfield et al., 1994; Bryan et al., 1997a). Larvae of H. elegans settle in response to cues from various sources, including cues from conspecific juveniles and adults (Bryan et al., 1997a,b; Lau & Qian, 1997; Bryan et al., 1998; Pechenik & Qian, 1998). In the laboratory, aqueous homogenates of adult worms in tubes, worms alone, tubes alone, and worm tubes baked in a muffle furnace (at 525 ◦ C) were assayed at similar concentrations for settlement and metamorphic induction of H. elegans larvae (Bryan et al., 1997a). No larvae metamorphosed in the control after a 14 day incubation; at a concentration of 0.2 mg homogenate ml−1 seawater, homogenized worms with tubes or without tubes induced higher percentages of metamorphosis while tubes alone and baked tubes induce 2% and 0% metamorphosis within 2 days. These results indicate that the larval settlement inducer originates from the worm body (Bryan
et al., 1997a). Larval settlement response to adult homogenate (without tube) is concentration-dependent. Although the percentage of worms metamorphosed in dishes containing concentrations of 0.2, 0.1 and 0.04 mg homogenate ml−1 seawater were all significantly higher than the control dishes (FSW only), the concentration of 0.2 mg homogenate ml−1 seawater induced the highest levels of metamorphosis after a 2 and 4 day incubation while the highest concentrations assayed, 0.6 and 1.4 mg ml−1 , were toxic to larvae (Bryan et al., 1997a). Size fractions of 30 kd were assayed along with unfractioned homogenate and the results indicated that the
