Marine Invertebrate Larvae Research Articles

Host-specificity of monogenean (platyhelminth) parasites: a role for anterior adhesive areas?

Monogeneans (flatworms) are among the most host-specific of parasites in general and may be the most host-specific of all fish parasites. Specificity, in terms of a restricted spatial distribution within an environment, is not unique to parasites and is displayed by some fungi, insects, birds, symbionts and pelagic larvae of free-living marine invertebrates. The nature of cues, how “habitats” are recognised and how interactions between partners are mediated and maintained is of interest across these diverse “associations”. We review some experiments that demonstrate important factors that contribute to host-specificity at the level of infective stages (larvae of oviparous monogeneans; juveniles of viviparous gyrodactylids) and adult parasites. Recent research on immune responses by fish to monogenean infections is considered. We emphasise the critical importance of host epidermis to the Monogenea. Monogeneans live on host epidermis, they live in its products (e.g. mucus), monopisthocotyleans feed on it, some of its products are “attractants” and it may be an inhospitable surface because of its immunological activity. We focus attention on fish but reference is made to amphibian hosts. We develop the concept for a potential role in host-specificity by the anterior adhesive areas, either the specialised tegument and/or anterior secretions produced by monogeneans for temporary but firm attachment during locomotion on host epithelial surfaces. Initial contact between the anterior adhesive areas of infective stages and host epidermis may serve two important purposes. (1) Appropriate sense organs or receptors on the parasite interact with a specific chemical or chemicals or with surface structures on host epidermis. (2) A specific but instant recognition or reaction occurs between component(s) of host mucus and the adhesive(s) secreted by monogeneans. The chemical composition of fish skin is known to be species-specific and our preliminary analysis of the chemistry of some monogenean adhesives indicates they are novel proteins that display some differences between parasite families and species.

Variation in attachment of the barnacle Balanus amphitrite: sensation or something else?

Settlement, attachment, and metamorphosis of larvae of marine invertebrates is extremely variable in the laboratory and in nature. The relative contributions of genetic, ontogenetic and environmental effects to this variation are not well understood. We tested for genetic, maternal and ontogenetic effects on permanent attachment of cyprids of the barnacle Balanus amphitrite (Darwin). Larvae were assayed for attachment in response to a control treatment and 3 artificial inducers: 20-hydroxyecdysone (20-HE), 3-isobutyl-1-methylxanthine (IBMX), and phorbol 12,13- dibutyrate (PDB). We observed strong family effects on attachment in the control, 20-HE, and IBMX treatments, suggesting that attachment in response to these treatments is influenced by genetic or maternal factors. In addition, attachment responses were correlated, indicating that the same genetic or maternal factors may affect attachment in each treatment. Family effects on attachment in the control treatments decreased with cypris age, suggesting the loss of genetically- or maternally- influenced discriminatory behavior in older larvae. The mechanisms underlying genetic or maternal effects on attachment may act via the external receptors and signal-transduction pathways regulat- ing larval responses. Alternatively, there may be variation in larval 'vigor' or the rate at which cyprids attain metamorphic competence.

Cellular analyses of sea urchin metamorphosis

Larvae of marine invertebrates undergo metamorphosis in response to environmental cues (Chia & Burke, 1978). In sea urchins, free fatty acids (Kitamura et al., 1993), dibromomethane (Taniguchi et al., 1994), pheromonal peptides (Burke, 1984) and L-glutamine (Yazaki & Harashima, 1994; Yazaki, 1995) have been known as metamorphosis-inducing substances. The mechanisms by which cells respond to these cues and how the larval tissues are absorbed have not been clear, however. In the present study, we used L-glutamine (Gln) and a natural cue, green algae (Ulvella sp.), to induce metamorphosis of Hemicentrotus pulcherrimus and Anthocidaris crassispina, and investigated the intracellular changes during metamorphosis.After being subjected to 10−5–10−3 M Gln for 10–24 h, larvae cease swimming, settle, begin to retract their larval arms, extrude the primary podia and finally evert their echinus rudiment (ER). In H. pulcherrimus, larvae retracted their arms from 6 h to 24 h after the start of Gln treatment and then everted the ER. A. crassispina larvae underwent similar processes to those of H. pulcherrimus. The larval surface is composed of squamous epithelium and columnar epithelium. The epithelium of the ciliary bands or epaulets is columnar.In the squamous epithelium, the nuclear chromatin in the larval arms and body, and in the oesophagus, markedly condensed after treatment with Gln for 24 h. Electron microscopy revealed swelling of both nuclei and mitochondria, while their membranes seemed to be intact. In the cytoplasm, lipid-like structures and electron-dense substances appeared. A further 24 h after Gln treatment, the chromatin condensation had progressed. Most nuclei in which chromatin had condensed were positive to the TUNEL assay, which detects DNA fragmentation. These results suggest that cell death in the squamous epithelium is apoptotic rather than necrotic.

Description of planktonic polychaete larvae from Terre Ad�lie and the Ross Sea (Antarctica)

An analysis was carried out on 51 samples from 32 stations, obtained during the I.C.O.T.A. cruise between 18 January 1996 and 7 February 1996 in the Pointe Geologie Archipelago, Terre Adelie, and on samples obtained by 2 sediment traps deployed in the Ross Sea, within the framework of the Italian C.L.I.M.A. Project. These samples showed the unexpected presence of numerous larvae of benthic marine invertebrates. In addition, larvae from the Ross Sea were examined. Seven phyla with complex life-cycles and at least 18 species of benthic polychaetes were represented. Morphological descriptions by scanning electron microscopy of the most frequent larvae and identification of larval types are provided. Descriptions from the literature of larvae collected in the Antarctic Peninsula were used for comparison. Morphological descriptions do not always lead to identification at the species level; however, these descriptions are indispensable for establishing the geographic distribution of larvae. It has been demonstrated that indirect identification methods relying on the premise that the benthic fauna is entirely known and that sampled larvae belong only to species already known through their benthic adult stage may lead to obvious absurdities. As an ecological consequence, Thorson's rule, according to which indirect development should not be represented and direct development should be dominant at high latitudes, is not supported by our data. A short comparison of the local reproductive pattern with a subantarctic area, the Kerguelen Archipelago, strongly suggests that the absence of planktonic development in this area cannot be explained by Thorson's rule. This rule has been invalidated in more austral zones; the prevalent reproductive pattern in the Kerguelen Archipelago is linked to the very marked advective strength of water flow.

The distribution, abundance and seasonality of pelagic marine invertebrate larvae in the maritime Antarctic

A survey of pelagic larvae was undertaken between November 1992 and February 1995 at Signy Island, Antarctica (60° 43′ S, 45° 36′ W). A diver–towed net and hand–held plankton pump were used at five sites of varying depths (6–30 m) and benthic substrata, in a combination of monthly and fortnightly samples. Overall larval ecological diversity was much higher than expected, with 131 morphologically distinct larval forms collected, apparently representing most of the benthic phyla present. The species richness observed is comparable with levels recorded at temperate latitudes, and higher than Arctic data and the implications for Thorson's rule (the inferred cline of reduced pelagic larval diversity towards high latitudes) is discussed. Larval abundances were low (mean 2.6 individuals per m 3 ) which were two to six orders of magnitude lower than peaks in comparable data from temperate and tropical zones. We suggest that the low abundances recorded are a reflection of both slow developmental rates and a high dilution of larvae, reducing synchrony and spreading larvae over larger distances. Three seasonal periods, during which different larval types occur, have been identified. Summer, late summer and winter spawning strategies were discernable, and in some groups larvae occurred throughout the year.

Ontogenies of Phototactic Behavior and Metamorphic Competence in Larvae of Three Species of Bugula (Bryozoa)

The free swimming larvae of many marine invertebrates actively respond to light. Light cues can be used to regulate position in the water column and to facilitate encountering sites suitable for metamorphosis. We examined the ontogeny of larval phototaxis and the ontogeny of metamorphic competency in larvae from three congeneric species of bryozoans. Larvae of Bugula neritina are positively phototactic on emergence from the brood chamber, whereas larvae of B. simplex and B. stolonifera appear initially photoneutral when populations of larvae are examined. Larvae of all three species become photonegative with time. Temporally coincident with this change to negative phototaxis is an increase in the competency of larvae to initiate metamorphosis. This observation suggests that these events are either physiologically linked or co-occurring, but independent developmental processes. We tested these hypotheses by artificially changing the sign of phototaxis from positive to negative using 10-5 M bathapplied 5-hydroxytryptamine (5HT) in larvae of B. neritina that were swimming for 1 h. Larvae that were photopositive and 1-h-old did not metamorphose at levels significantly different from larvae that were 1-h-old and treated with 5HT (i.e., young, photonegative larvae). Additionally, photopositive larvae which were swimming for 4 h initiated metamorphosis at rates nearly identical to photonegative larvae of the same age. Our data document that in larvae of B. neritina the changes in sign of phototaxis and levels of metamorphic competency are independent developmental events that occur in temporal coincidence. The concurrent timing of these two pathways may have been synchronized through selective processes resulting in a tight coupling between arrival at potentially suitable sites for metamorphosis and ability to respond to metamorphic cues. Additional key words: bryozoans, phototaxis Many benthic marine invertebrates have as part of their life cycle a planktonic larval stage. Successful completion of the life cycle requires return to a suitable benthic habitat. Larvae utilize a variety of physical and biological cues to facilitate location of sites favorable for subsequent adult life (e.g., Pawlik 1992; Young 1995). Light is one such physical signal. Light is a ubiquitous vector in relatively shallow waters that exhibits both spatial and temporal variation. The temporal variation can be highly predictable (sunrise and sunset) or highly variable (changes in cloud cover). Responses to light are one potentially useful means of regulating vertical orientation and position in the water column (e.g., Thorson 1964; Clarke 1970; Cronin & Forward 1979; Sulkin 1984; Forward 1988; Barile et al. 1994; Young 1995). Additionally, in the case of meroplanktonic larvae, phototactic responses can condu tribute to delivery of larvae to benthic sites suitable for metamorphosis (e.g., McDougall 1943; Ryland 1960; Crisp 1974; Young & Chia 1982; Olson 1985; Dirnberger 1993). Thorson's (1964) seminal work was the first detailed comparative study of the ontogeny of phototaxis across several phyla of benthic marine invertebrates. He observed that 82% of 141 species from 11 phyla had early-stage larvae that were initially photopositive. Of these, 76% became photonegative before the conclusion of larval life. Thorson suggested that earlystage positive phototaxis increases the chances that larvae will be transported into the water column and hence away from conspecific adults and benthic predators. In the water column, larvae can be passively dispersed by currents, and feeding larvae can have access to more abundant populations of phytoplankton. Larvae that change to photonegative with time will then likely move to benthic sites, which is essential for completion of the life cycle. ies tactic ior t rphic tence e i s la ( a)

A method for accurate measurements of the respiration rates of marine invertebrate embryos and larvae

Measurements of respiration rates are essential to quantify the energy requirements of embryos and larvae. Here we describe a ')_rBOD' method that employs small (~1 ml) Biological Oxygen Demand (BOD) glass vials in which embryos and larvae are incubated. A decrease in oxygen concen- tration is measured by injecting seawater from each vial into the measurement chamber of a standard polarographic oxygen sensor. This pBOD method was used to measure respiration rates of larvae of the sea urchin Strongylocentrotus purpuratus (at 15°C) and embryos and larvae of 2 Antarctic echino- derms, the sea urchin Sterecbinus neumayeri and the seastar Odontasfer valiclus (both at -1SC). For validation, a comparison of different methods was performed with embryos and larvae of S. neumayeri. The nBOD method gave results for respiration rates during development that were equivalent to those obtained with either coulometric capacitance respirometry or standard Winkler's titrations of large (300 ml) BOD bottles. Currently, the most common method for measuring respiration rates of inverte- brate embryos and larvae is to place them in small respiration chambers and continuously monitor oxygen tension with a polarographic oxygen sensor (POS). However, respiration rates for embryos and larvae of S. neumayeri were underestimated when standard POS measurements were compared to the measurements made with either pBOD, Winkler's titrations or coulometric capacitance respirometry. A comparison of the pBOD and POS methods during early development in S. neumayeri resulted in different estimates of 920 and 343 pJ, respectively, for the total energetic cost of embryogenesis to gastrulation, illustrating that the POS error can be as great as 63 %. The yBOD method is accurate for micro-respiration measurements of invertebrate embryos and larvae as well as being simple to operate and appropriate for field work.

Metamorphosis Is Not a New Beginning

M any marine invertebrate species have complex life cycles, in which one or more free-living developmental stages eventually metamorphoses to a morphologically-and often ecologically and physiologically-distinct juvenile stage. Such life cycles are also common among insects, amphibians, and marine fishes. Among marine invertebrates, complex life cycles are widely distributed among such diverse animals as sponges; turbellarian and trematode flatworms; gastropod and bivalved mollusks; polychaete worms; lobsters, crabs, barnacles, and other crustaceans; bryozoans; and echinoderms (Thorson 1950). Marine invertebrate larvae may feed on phytoplankton and other particulates or subsist entirely on yolk or other nutrients provided by the mother. They may spend as little as a few minutes or as long as several to many months in the plankton before metamorphosing to adult form and habitat (Pechenik 1990). Marine invertebrate larvae are

Seaweed secondary metabolites as antifoulants: effects of Dictyota spp. diterpenes on survivorship, settlement, and development of marine invertebrate larvae

A recent investigation showed that the brown seaweed Dictyota menstrualis was unfouled relative to co-occurring seaweeds, and that larvae of fouling invertebrates avoided settling on D. menstrualis due to chemicals on its surface. The secondary metabolites dictyol E and pachydictyol A are among the compounds found on this alga's surface. In the present study, we tested the effects of specific diterpenes from Dictyota on the survivorship, growth, and development of invertebrate larvae and developing juveniles that could foul seaweeds. Exposure to dictyol E, dictyol B acetate, pachydictyol A, and dictyodial from Dictyota menstrualis and D. ciliolata caused significant larval mortality, abnormal development, and reduce growth rates for three species of co-occurring invertebrates when their larvae were forced into contact with these metabolites. Larvae were damaged at metabolite concentrations as low as 5% of maximum possible surface concentrations of these compounds for the populations of Dictyota we studied. The negative effects of these secondary metabolites on potential foulers, in conjunction with data demonstrating larval avoidance of dictyol-covered surfaces, suggest that these compounds could function as chemical defenses against fouling, and could select for larvae that avoid hosts producing these metabolites.

The genetic structure of intertidal populations of two species of nudibranch molluscs with planktotrophic and pelagic lecithotrophic larval stages: are pelagic larvae “for” dispersal?

Planktonic larvae of benthic marine invertebrates may confer larval dispersal and colonization potential on the reproductive adult. Analyses of genetic heterogeneity of local populations provide an indirect means of assessing gene flow and the scale of realized dispersal of larval migrants. Here we compared populations of two species of nudibranchs with long term planktotrophic ( Goniodoris nodosa) and shorter term pelagic lecithotrophic ( Adalaria proxima) veliger stages. Both species have annual life cycles, population turnover is absolute and free-swimming larvae hatch from benthic spawn masses. G. nodosa larvae are pelagic for up to ∼3 months, whereas those of A. proxima can metamorphose within 1–2 d of release although, at least in the laboratory, they may delay metamorphosis for several weeks. The broad ranges of pelagic larval transport potential were estimated at 10 2–10 3 km for G. nodosa and 10 1–10 2 km for A. proxima: significant differentiation of populations of G. nodosa was predicted to occur on a scale of 10 3–10 4 km, whereas for A. proxima differentiation was expected on a scale of perhaps only 10 2–10 3 km. For both species, analyses were undertaken of allozyme variation among sites throughout the 1600 km British Isles range of A. proxima. F-statistic analyses showed no evidence of population differentiation for G. nodosa (11 populations). By contrast, comparable data for 19 populations of A. proxima showed very high levels of differentiation, but no obvious pattern of variation or significant isolation by distance. More detailed sampling of nine populations of A. proxima over a 26.18 km range in W Scotland – in a location characterised by highly dispersive tidal currents – revealed (1) similar levels of differentiation to the 1600 km range, (2) clear distance-related clinal patterns of variation, and (3) a significant inverse relationship between genetic differentiation and geographic distance. For Adalaria proxima there was evidence of significant differentiation of populations on a scale of ≤10 1 km. This was counter to our expectations of larval transport potential for A. proxima and implies that these larvae are behaviourally adapted to avoid dispersal in the water column and thereby recruit locally. Whilst the planktotrophic species, G. nodosa, conformed to our expectations, the clear implication of the data for A. proxima is that possession of a pelagic larval stage does not necessarily facilitate dispersal and colonisation potential for this sedentary invertebrate. Extrapolations and predictions of dispersal potential, and gene flow in natural populations, based on laboratory observations of the larval phase duration may be markedly error-prone.

The role of GLWamides in metamorphosis of Hydractinia echinata.

The metamorphosis of many marine invertebrate larvae is induced by environmental signals. Upon reception of the cues, internal signals have to be set in motion to convey information to all cells of the larvae. For hydrozoan larvae it was hypothesised that ectodermal neurosensory cells at the anterior part are those cells receptive of the inducer. Recently, it was shown that novel peptides with a common GLWamide terminus are found in Cnidaria. These peptides are located in a specific subset of the anterior sensory cells. It was hypothesised that the neuropeptides represent an internal signal coordinating the metamorphic process. In the current study we present further evidence for this hypothesis. Induction of metamorphosis is very specific for the GLWamide terminus and amidation is essential. The potency to metamorphose is strongly correlated with the presence of GLWamide-immunoreactive cell bodies. Our data fit our hypothesis about a very important role of GLWamides in the initiation of the morphogenetic processes very well.

Metamorphic processes in the soft corals Heteroxenia fuscescens and Xenia umbellata: The effect of protein kinase C activators and inhibitors

Summary During the life cycle of marine invertebrates, including corals with benthic and planktonic phases, embryogenesis leads to a free swimming larva which has to find a suitable habitat to settle and metamorphose. Initial settlement and induction of metamorphosis of benthic marine invertebrate larvae are generally determined by interaction of external biochemical and/or physical factors. In a preliminary study we showed that the phorbol ester 12-o-tetra-decanoyl-phorbol-13-acetate (TPA) induces metamorphosis in six Red Sea coral species. Phorbol esters are known to activate the enzyme protein kinase C (PKC) and therefore play an important role in studying the phosphatidy linositol signal (PI) transduction system. The potency of several phorbol esters to induce metamorphosis in planulae of the Red Sea soft coral species Heteroxenia fuscescens and Xenia umbellata was examined to find further indications for the involvement of PKC in anthozoan metamorphosis. All tested phorbol esters, 12-o-tetra-decanoyl-phorbol-13-acetate, phorbol-12, 13-didecanoate, 12-retinoyl-phorbol-13-acetate and phorbol-12, 13-dibutyrate triggered metamorphosis in planulae of both species in a concentration-dependent way. The maximum levels of metamorphosis achieved varied from 30% to 100%, depending on both the species tested and the phorbol ester applied, using concentrations ranging from 10−7 to 10−9 mol/l except for 12-retinoyl-phorbol-13-acetate. In addition, the response of planula fragments to artificial inducers was studied. Our results indicate that the different cnidarian taxa not only respond to a variety of metamorphic cues, but also possess divergent mechanisms for reception and transmission of metamorphic inducers. Phloretin, an inhibitor of mammalian PKC, completely blocked phorbol ester-induced metamorphosis in Heteroxenia fuscescens planulae. Data which indicate the presence of PKC-related enzymes as the biochemical base for metamorphosis are now documented for species of the cnidarian classes Hydrozoa, Scyphozoa and Anthozoa. Based on our results, and the work of other authors, we therefore suggest that the phosphatidy linositol-signaling pathway might represent the general mechanism which regulates metamorphosis in cnidarians.

Attachment of barnacle ( Balanus amphitrite Darwin) larvae: responses to bacterial films and extracellular materials

Bacterial films on submerged substrata can affect settlement of marine invertebrate larvae. We examined the effects of three species of bacteria, Deleya marina (Cobet et al.; Baumann et al.), Alteromonas macleodii Baumann et al. and Pseudomonas fluorescens Migula, on the attachment of cypris larvae of the barnacle Balanus amphitrite Darwin in laboratory assays. Bacterial cells or culture supernatant containing extracellular materials were allowed to adsorb to replicate sterile polystyrene petri dishes or borosilicate glass vials, and B. amphitrite cyprids were introduced to filmed and unfilmed (control) containers. Cells and supernatant from both log- and stationary-phase bacterial cultures were used in assays, which were repeated using different batches of larvae. Attachment of cyprids to control and filmed containers was monitored for several days. Cells and supernatant from stationary-phase cultures of D. marina consistently retarded cyprid attachment compared to attachment onto control polystyrene petri dishes, but not glass vials. Assays using films derived from log-phase D. marina cultures, and all assays with both log- and stationary-phase A. macleodii and P. fluorescens films, yielded weak and inconsistent effects on cyprid attachment. Despite variability among assays, bacterial effects on larval attachment within a particular assay generally were consistent for the duration of the assay (up to a week). Settlement was independent of the number of cyprids introduced to a container, within the range employed in assays. However, attachment of cyprids to control containers varied significantly among batches of barnacle larvae. Variation in results among assays may be related to variability in larval behavior, possibly explainable by larval diet.

Role of Chemical Inducers in Larval Metamorphosis of Queen Conch, Strombus gigas Linnaeus: Relationship to Other Marine Invertebrate Systems.

Chemical cues are important in the exogenous and endogenous control of metamorphosis in many marine invertebrate larvae. In the queen conch, Strombus gigas Linnaeus, larval metamorphosis is induced by low molecular weight compounds associated with dominant species of red algae found in conch nursery grounds; these species include the foliose rhodophyte Laurencia poitei (Lamouroux). The responses of conch larvae to the algal-associated cues are dependent on concentration and length of exposure, with the initial events of metamorphosis occurring within 10 min of treatment with an aqueous extract of L. poitei. The free amino acids valine and isoleutine mimic the effects of the natural inducer, and they may bind to and be recognized by the same sites on the larvae as the algal cues. Hydrogen peroxide, vanadate, and {gamma}-aminobutyric acid (GABA), as well as elevated K+ concentrations (i.e., above ambient seawater levels), also induce larval metamorphosis. Acetylsalicylic acid decreases the responses of conch larvae to the algal-associated cues and to the free amino acids, but it has no effect on the induction triggered by hydrogen peroxide. The chemical induction of metamorphosis in conch larvae shares many general features with chemoreception in aquatic invertebrates. The natural inducers of metamorphosis, like the cues involved in olfactory responses in other marine organisms, are of low molecular weight and water soluble. In addition, the results of the experiments with hydrogen peroxide, vanadate, and GABA suggest that second messenger pathways are involved in conch metamorphosis.

Effects of substratum and conspecific adults on the metamorphosis of Chasmagnathus granulata (Dana) (Decapoda: Grapsidae) megalopae

The ability of marine invertebrate larvae to delay their metamorphosis in the absence of adequate environmental cues has been reported for numerous sedentary and sessile species. In the present study, the effect of various substrata and the presence of conspecific adults on the metamorphosis of a mobile species, the crab Chasmagnathus granulata, was evaluated. The duration of the megalopa stage in experiments with six different substrata and in the presence or absence of conspecific adults was compared in a laboratory study. In addition, the influence of natural substrata was compared with that of artificial substrata of similar grain size or texture. In a further experiment, the two most effective cues (natural mud and conspecific adults) were tested as single vs. combined factors. Natural mud and unidentified chemical cues from conspecific adults had the strongest accelerating effects on development duration to metamorphosis. With the exception of nylon threads (artificial filamentous substratum), none of the artificial substrata had a significant effect on the duration of the megalopa stage. Simultaneous exposure to natural mud and water containing chemical cues from conspecific adults accelerated metamorphosis more than each of these factors separately. Megalopae that were reared without a substratum (control) delayed their metamorphosis by 29% (about 3 days) compared with those in simultaneous contact with natural mud and rearing water of adult conspecifics. The results indicate that the metamorphosis of the megalopa of C. granulata is influenced by the presence or absence of environmental stimuli that are associated with the preferred adult habitat.

The D P Wilson Lecture. Research on settlement and metamorphosis of marine invertebrate larvae: Past, present and future

In 1952, D P Wilson remarked that invertebrate larvae, “... are indeed not helpless, for they are endowed with the power of choice,...”; at settlement. Subsequent efforts have greatly enhanced understanding of the conditions under which larvae metamorphose, altering the conceptual view of this process from “larval choice”; to “developmental induction,”; and aligning studies of invertebrate metamorphosis with those of modern developmental biology. While the precise chemical nature of most metamorphic inducers evades us, elucidation of the phenomena of reception and transduction appears imminent for many larvae. The evidence for receptor cells and molecules specific for inducer substances is convincing for larvae in several phyla, and a generalized picture of stimulation of electrically excitable cells has emerged; larvae from at least eight invertebrate phyla metamorphose in response to augmented K + . To be determined is whether genes specifying receptor proteins for metamorphic inducers share homology and whether metamorphic signals are transduced by the same intracellular pathways across invertebrate phyla. Metamorphic competence can be a developmentally unique condition where non‐growing, non‐feeding larvae are primed to metamorphose when signaled; a genetic explanation for competence merits intense exploration. The acquisition for marine invertebrate larvae of knowledge comparable to that for the genetic basis of timing of photic responses and pupal eclosion in Drosophila will provide material for exciting research in the years ahead. Many species (e.g. the polychaete Hydroides elegans) offer excellent models for laboratory genetic studies of invertebrate settlement and metamorphosis.

Inhibition and facilitation of settlement of epifaunal marine invertebrate larvae by microbial biofilm cues

The methodologies applied in laboratory and field studies of the effects of biofilm cues on marine invertebrate larval settlement on hard substrata are briefly reviewed and some of the general problems (such as experimental repeatability, handling effects) that may be encountered, and the ecological representativeness of the species utilized are discussed. The review focuses also on the range of bioassay approaches and highlights the wide range of larval densities, substrata and biofilm sources that have been variously applied. It is important to consider all of these features in any attempt at formulating general principles and trends relating to larval settlement responses in natural assemblages. This review also highlights the prevalence of reports of facilitatory, rather than neutral or inhibitory, effects amongst the various species investigated and considers the possible role(s) of larva‐larva interactions, cue perception distances, hierarchical effects and whether or not biofilm cues are surface‐bound or waterborne in determining and affecting these responses. With specific reference to natural hard substratum assemblages, the putative adaptive significance of biofilm‐mediated facilitation and inhibition of larval settlement is assessed and additional possible lines of future research are suggested.

Phototoxicity of individual polycyclic aromatic hydrocarbons and petroleum to marine invertebrate larvae and juveniles

AbstractPhototoxicity resulting from photoactivated polycyclic aromatic hydrocarbons (PAHs) has been reported in the literature for a variety of freshwater organisms. The magnitude of increase in PAH toxicity often exceeds a factor of 100. In the marine environment phototoxicity to marine organisms has not been reported for individual or complex mixtures of PAHs. In this study, larvae and juveniles of the bivalve, Mulinia lateralis, and juveniles of the mysid shrimp, Mysidopsis bahia, were exposed to individual known phototoxic PAHs (anthracene, fluoranthene, pyrene), as well as the water‐accommodated fractions of several petroleum products (Fuel Oil #2, Arabian Light Crude, Prudhoe Bay Crude, Fuel Oil #6) containing PAHs. Phototoxicity of individual PAHs was 12 to >50,000 times that of conventional toxicity. Three of the petroleum products demonstrated phototoxicity while the lightest product, Fuel Oil #2, was not phototoxic at the concentrations tested. The phototoxicity of petroleum products appears to be dependent on the composition and concentrations of phototoxic PAHs present: lighter oils have fewer multiple aromatic ring, phototoxic compounds while heavier oils have higher levels of these types of molecules. This study shows that phototoxicity can occur in marine waters to marine species. Further, the occurrence of oil in marine waters presents the additional risk of phototoxicity not routinely assessed for during oil spills.

Life on the edge: conspecific attraction and recruitment of populations to disturbed habitats.

Variation in the recruitment of benthic marine invertebrates is often attributed to the interaction of the supply of new individuals to a habitat and the availability of space for colonisation when they arrive. Also important in determining variation in recruitment is the response of the larvae to the characteristics of the habitat. Larvae of many benthic marine invertebrates have shown great specificity of requirements in setting their limits of distribution at the time of selection of a habitat. The tubeworm Galeolaria caespitosa shows great variation in recruitment from place to place on rocky intertidal seashores and is a gregarious animal with larvae showing directed responses to conspecific adults on the substratum. I hypothesised that, if variation in recruitment of G. caespitosa were independent of conditions on the substratum, the magnitude of recruitment in patches of the same shape but different sizes cleared within continuous mats of conspecific adults would be directly related to the area available for colonisation in the patch. Alternatively, if variation in recruitment were due to the response of larvae to conspecific adults on the substratum, the magnitude of recruitment would be a function of the perimeter of the patch, which, given patches of the same shape, is a measure of the influence of conspecific adults in that patch. To distinguish between these alternatives, small (area = 25 cm2; perimeter = 20 cm) and large (area = 225 cm2; perimeter = 60 cm) square patches were cleared within continuous mats of conspecific adults at four sites and recruitment of G. caespitosa was monitored over two seasons of recruitment. The density of recruits per unit area was, on average, almost three times greater in small than in large patches, indicating that recruitment of G. caespitosa is not directly related to the area of the patch. In contrast, the density of recruits per unit perimeter was not significantly different between small and large patches, indicating that recruitment of G. caespitosa is related to the proximity of conspecific adults in the patch. Therefore, at a given site, the perimeter of patches within mats of G. caespitosa is a better predictor of the relative magnitude of recruitment among patches than that provided by their areas. These results are contrary to many models of invertebrate recruitment that assume close linkage between available space on the substratum and settlement. Moreover, they highlight the importance of behavioural responses of animals at the time of selection of habitat in accounting for variation in recruitment. For populations of organisms that display gregarious behaviour at settlement, or conspecific attraction, this direct relationship between the perimeter of patches and recruitment could be used as a tool in restoring populations to disturbed habitats. The added benefit of such facilitative interactions in restoring populations is that they provide increasing returns to the population for a given supply of potential colonists to a habitat.

Inhibition and facilitation of bryozoan and ascidian settlement by natural multi-species biofilms: effects of film age and the roles of active and passive larval attachment

Sessile marine invertebrate larvae can recognize suitable settlement substrata by using various environmental cues, including organic/microbial biofilms. In laboratory choice assays, the effect of biofilms of varying ages on the settlement behaviour of two fouling organisms was assessed. The species included the arborescent cheilostome bryozoan Bugula flabellata (Thompson) and the solitary ascidian Ciona intestinalis (L.), both of which are characteristic of temperate sublittoral hard substratum assemblages in northwest Europe. Experiments were carried out using polystyrene petri dish substrata preconditioned with multispecies biofilms from natural laboratory-aquarium seawater for 1, 3, 6 or 12 d. Unfilmed (new, initially sterile) dishes were used as control substrata. Whereas the coronate larvae of B. flabellata generally were inhibited by biofilming, irrespective of film age, the settlement of tadpole larvae of C. intestinalis was facilitated on biofilmed substrata, and numbers of settled larvae generally increased with biofilm age: the highest mean numbers were counted on 12 d-old biofilms. In C. intestinalis, settlement and metamorphosis are processes which can be temporally separate and are possibly induced by different environmental cues. This study therefore distinguished between C. intestinalis larvae which were attached to the biofilm surface by the anterior, and those larvae entrapped by the biofilm but not settled in the conventional meaning of the term. As reported in previous studies, we did observe that such entrapped larvae could subsequently attach and develop successfully into sessile juveniles. Both the numbers of “attached” and “trapped” tadpoles increased with biofilm age. Assuming that “settlement” is essentially a process involving the active behavioural response of larvae to environmental cues, it seems that the facilitated attachment of C. intestinalis onto biofilmed substrata is due to the combined effect of active habitat selection and passive deposition/“entrapment” of larvae onto the “sticky” substratum.