Metamorphic Competence, a Major Adaptive Convergence in Marine Invertebrate Larvae

Larvae from diverse marine-invertebrate phyla are able to respondrapidly to environmental cues to settlement and to undergo veryrapid metamorphic morphogenesis because they share the developmentaltrait of metamorphic competence. The competent state, characteristicof larvae as diverse as those of cnidarian planulae, molluscanveligers, and barnacle cyprids, is one in which nearly all requisitejuvenile characters are present in the larva prior to settlement.Thus metamorphosis, in response to more or less specific environmentalcues (inducers), is mainly restricted to loss of larva-specificstructures and physiological processes. Competent larvae oftwo "model marine invertebrates" studied in the authors' laboratory,the serpulid polychaete Hydroides elegans and the nudibranchPhestilla sibogae, complete metamorphosis in about 12 and 20hr, respectively. Furthermore, little or no de novo gene actionappears to be required during the metamorphic induction processin these species. Contrasting greatly with the slow, hormonallyregulated metamorphic transitions of vertebrates and insects,competence and consequent rapid metamorphosis in marine invertebratelarvae are conjectured to have arisen in diverse phylogeneticclades because they allow larvae to continue to swim and feedin the planktonic realm while simultaneously permitting extremelyfast morphological transition from larval locomotory and feedingmodes to a different set of such modes that are adaptive tolife on the sea bottom.     

Interspecific variation in metamorphic competence in marine invertebrates: the significance for comparative investigations into the timing of metamorphosis

Metamorphosis in marine invertebrate larvae is a dynamic, environmentallydependent process that integrates ontogeny with habitat selection.The capacity of many marine invertebrate larvae to survive andmaintain metamorphic competence in the absence of environmentalcues has been hypothesized to be an adaptive convergence (Hadfieldand others 2001). A survey of the literature reveals that asingle generalized hypothesis about metamorphic competence asan adaptive convergence is not sufficient to account for interspecificvariation in this character. In an attempt to capture this variation,we discuss the "desperate larva hypothesis" and propose twoadditional hypotheses called the "variable retention hypothesis"and the "death before dishonor hypothesis." To validate theseadditional hypotheses we collected data on taxa from the publishedliterature and performed a contingency analysis to detect correlationsbetween spontaneous metamorphosis, habitat specificity and/orlarval life-history mode, three characters relevant to environmentallyinduced settlement and metamorphosis. In order to account forphylogenetic bias in these correlations, we also constructeda phylogeny of these taxa and again performed a character-correlationanalysis. Both these tests suggest that juvenile habitat specificityis correlated to the capacity of individuals to retain the competentlarval state in the absence of substrate cues and thereforevalidate the existence of more than one hypothesis about metamorphiccompetence. We provide new data from the sea urchin Lytechinuspictus that suggest that nitric oxide (NO) and thyroxine hormonesignaling interact to determine the probability of settlementin response to a settlement cue. Similarly, we provide evidencethat thyroxine signaling in the sand dollar Dendraster excentricusincreases spontaneous metamorphosis in the absence of cues fromadult conspecifics in a manner that is independent of larvalage.     

Implantation Serine Proteinases heterodimerize and are critical in hatching and implantation

Background

Metamorphosis is a complex, highly conserved and strictly regulated development process that involves the programmed cell death of obsolete larval organs. Here we show a novel functional role for the aspartic proteinase cathepsin D during insect metamorphosis.

Results

Cathepsin D of the silkworm Bombyx mori (BmCatD) was ecdysone-induced, differentially and spatially expressed in the larval fat body of the final instar and in the larval gut of pupal stage, and its expression led to programmed cell death. Furthermore, BmCatD was highly induced in the fat body of baculovirus-infected B. mori larvae, suggesting that this gene is involved in the induction of metamorphosis of host insects infected with baculovirus. RNA interference (RNAi)-mediated BmCatD knock-down inhibited programmed cell death of the larval fat body, resulting in the arrest of larval-pupal transformation. BmCatD RNAi also inhibited the programmed cell death of larval gut during pupal stage.

Conclusion

Based on these results, we concluded that BmCatD is critically involved in the programmed cell death of the larval fat body and larval gut in silkworm metamorphosis.     

The initiation of metamorphosis as an ancient polyphenic trait and its role in metazoan life-cycle evolution

Comparative genomics of representative basal metazoans leaves little doubt that the most recent common ancestor to all modern metazoans was morphogenetically complex. Here, we support this interpretation by demonstrating that the demosponge Amphimedon queenslandica has a biphasic pelagobenthic life cycle resembling that present in a wide range of bilaterians and anthozoan cnidarians. The A. queenslandica life cycle includes a compulsory planktonic larval phase that can end only once the larva develops competence to respond to benthic signals that induce settlement and metamorphosis. The temporal onset of competence varies between individuals as revealed by idiosyncratic responses to inductive cues. Thus, the biphasic life cycle with a dispersing larval phase of variable length appears to be a metazoan synapomorphy and may be viewed as an ancestral polyphenic trait. Larvae of a particular age that are subjected to an inductive cue either maintain the larval form or metamorphose into the post-larval/juvenile form. Variance in the development of competence dictates that only a subset of a larval cohort will settle and undergo metamorphosis at a given time, which in turn leads to variation in dispersal distance and in location of settlement. Population divergence and allopatric speciation are likely outcomes of this conserved developmental polyphenic trait.     

Ecological regulation of development: induction of marine invertebrate metamorphosis

In the marine environment a wide range of invertebrates have a pelagobenthic lifecycle that includes planktonic larval and benthic adult phases. Transition between these morphologically and ecologically distinct phases typically occurs when the developmentally competent larva comes into contact with a species-specific environmental cue. This cue acts as a morphogenetic signal that induces the completion of the postlarval/juvenile/adult developmental program at metamorphosis. The development of competence often occurs hours to days after the larva is morphologically mature. In the non-feeding--lecithotrophic--larvae of the ascidian Herdmania curvata and the gastropod mollusc Haliotis asinina, gene expression patterns in pre-competent and competent stages are markedly different, reflecting the different developmental states of these larval stages. For example, the expression of Hemps, an EGF-like signalling peptide required for the induction of Herdmania metamorphosis, increases in competent larvae. Induction of settlement and metamorphosis results in further changes in developmental gene expression, which apparently is necessary for the complete transformation of the larval body plan into the adult form.     

Tissue-specific profile of DNA replication in the swimming larvae of Ciona intestinalis

The cell cycle is strictly regulated during development and its regulation is essential for organ formation and developmental timing. Here we observed the pattern of DNA replication in swimming larvae of an ascidian, Ciona intestinalis. Usually, Ciona swimming larvae obtain competence for metamorphosis at about 4-5 h after hatching, and these competent larvae initiate metamorphosis soon after they adhere to substrate with their papillae. In these larvae, three major tissues (epidermis, endoderm and mesenchyme) showed extensive DNA replication with distinct pattern and timing, suggesting tissue-specific cell cycle regulation. However, DNA replication did not continue in aged larvae which kept swimming for several days, suggesting that the cell cycle is arrested in these larvae at a certain time to prevent further growth of adult organ rudiments until the initiation of metamorphosis. Inhibition of the cell cycle by aphidicolin during the larval stage affects only the speed of metamorphosis, and not the formation of adult organ rudiments or the timing of the initiation of metamorphosis. However, after the completion of tail resorption, DNA replication is necessary for further metamorphic events. Our data showed that DNA synthesis in the larval trunk is not directly associated with the organization of adult organs, but it contributes to the speed of metamorphosis after settlement.     

Three‐dimensional fate mapping of larval tissues through metamorphosis in the glass sponge Oopsacas minuta

The tissue of glass sponges (Class Hexactinellida) is unique among metazoans in being largely syncytial, a state that arises during early embryogenesis when blastomeres fuse. In addition, hexactinellids are one of only two poriferan groups that already have clearly formed flagellated chambers as larvae. The fate of the larval chambers and of other tissues during metamorphosis is unknown. One species of hexactinellid, Oopsacas minuta, is found in submarine caves in the Mediterranean and is reproductive year round, which facilitates developmental studies; however, describing metamorphosis has been a challenge because the syncytial nature of the tissue makes it difficult to trace the fates using conventional cell tracking markers. We used three‐dimensional models to map the fate of larval tissues of O. minuta through metamorphosis and provide the first detailed account of larval tissue reorganization at metamorphosis of a glass sponge larva. Larvae settle on their anterior swimming pole or on one side. The multiciliated cells that formed a belt around the larva are discarded during the first stage of metamorphosis. We found that larval flagellated chambers are retained throughout metamorphosis and become the kernels of the first pumping chambers of the juvenile sponge. As larvae of O. minuta settle, larval chambers are enlarged by syncytial tissues containing yolk inclusions. Lipid inclusions at the basal attachment site gradually became smaller during the six weeks of our study. In O. minuta, the flagellated chambers that differentiate in the larva become the post‐metamorphic flagellated chambers, which corroborate the view that internalization of these chambers during embryogenesis is a process that resembles gastrulation processes in other animals.     

Analysis of molecular markers for metamorphic competency and their response to starvation or feeding in the mosquito, Aedes aegypti (Diptera: Culicidae)

The nutritional condition of fourth instar larvae of the yellow fever mosquito, Aedes aegypti, governs female longevity and egg production, both are key determinants of pathogen transmission. As well, nutrition provisions larval growth and development and attains its greatest pace in the last larval instar in preparation for metamorphosis to an adult. These developmental processes are regulated by a complex endocrine interplay of juvenile hormone, neuropeptides, and ecdysteroids that is nutrition sensitive. We previously determined that feeding for only 24 h post-ecdysis was sufficient for fourth instar Ae. aegypti larvae to reach critical weight and accumulate sufficient nutritional stores to commit to metamorphosis. To understand the genetic basis of metamorphic commitment in Ae. aegypti, we profiled the expression of 16 genes known to be involved in the endocrine and nutritional regulation of insect metamorphosis in two ways. The first set is a developmental profile from the beginning of the fourth instar to early pupae, and the second set is for fourth instars starved or fed for up to 36 h. By comparing the two sets, we found that seven of the genes (AaegCYP302, AaegJHE43357, AaegBrCZ4, AaegCPF1-2, AaegCPR-7, AaegPpl, and AaegSlif) were expressed during metamorphic commitment in fourth instars and in fed but not starved larvae. Based on these results, the seven genes alone or in combination may serve as molecular indicators of nutritional and metamorphic status of fourth instar Ae. aegypti larvae and possibly other mosquito species in field and laboratory studies to gauge sub-lethal effects of novel and traditional cultural or chemical controls.     

Phosphoproteomes of <Emphasis Type="Italic">Strongylocentrotus purpuratus</Emphasis> shell and tooth matrix: identification of a major acidic sea urchin tooth phosphoprotein,phosphodontin

Background  

Sea urchin is a major model organism for developmental biology and biomineralization research. However, identification of proteins involved in larval skeleton formation and mineralization processes in the embryo and adult, and the molecular characterization of such proteins, has just gained momentum with the sequencing of the Strongylocentrotus purpuratus genome and the introduction of high-throughput proteomics into the field.     

Evolution of direct-developing larvae: selection vs loss

Observations of a sea urchin larvae show that most species adopt one of two life history strategies. One strategy is to make numerous small eggs, which develop into a larva with a required feeding period in the water column before metamorphosis. In contrast, the second strategy is to make fewer large eggs with a larva that does not feed, which reduces the time to metamorphosis and thus the time spent in the water column. The larvae associated with each strategy have distinct morphologies and developmental processes that reflect their feeding requirements, so that those that feed exhibit indirect development with a complex larva, and those that do not feed form a morphologically simplified larva and exhibit direct development. Phylogenetic studies show that, in sea urchins, a feeding larva, the pluteus, is the ancestral form and the morphologically simplified direct-developing larva is derived. The current hypothesis for evolution of the direct-developing larval form in sea urchins suggests that major developmental changes occur by neutral loss of larval features after the crucial transition to a nonfeeding life history strategy. We present evidence from Clypeaster rosaceus, a sea urchin with a life history intermediate to the two strategies, which indicates that major developmental changes for accelerated development have been selected for in a larva that can still feed and maintains an outward, pluteus morphology. We suggest that transformation of larval form has resulted from strong selection on early initiation and acceleration of adult development.     

Lecithotrophic development and metamorphosis in the Indo-West Pacific brittle star Ophiomastix venosa (Echinodermata: Ophiuroidea)

Summary

The larval development of the ophiocomid ophiuroid Ophiomastix venosais described using SEM. The gastrula transforms into a uniformly ciliated early larva which progressively changes into a lecithotrophic late premetamorphic larva with a continuous bilateral ciliated band. This stage is short-lived and equivalent to a highly reduced ophiopluteus. Comparisons between O. venosa and other ophiuroid species whose development has been investigated suggest that, whatever the developmental mode (lecithotrophic or planktotrophic), a pluteus stage always occurs in ophiuroids with planktonic development. Two metamorphic stages were identified, the late metamorphic larva differing from the early one by the closure of the larval mouth. The appearance of the permanent mouth marks the end of the metamorphosis. The postlarva still possesses remnants of larval features. The transformation of the reduced ophiopluteus into a barrel-shaped metamorphic larva with transverse ciliated bands, a vitellaria larva, is followed. The possible occurrence of a unique type of metamorphic larva in non-brooding ophiuroids is discussed. Verification of this, however, needs further SEM investigations on metamorphic larva from species having “regular” planktotrophic development.     

Consumption and utilization of a soyflour-wheat germ diet by larvae of the tobacco budworm parasitized by the tachinidEucelatoria sp

The number ( \(\bar X\) =2.4) ofEucelatoria sp. maggots that completed development in 4th- or 5th-instar larvae of the tobacco budworm (TBW),Heliothis virescens (F.), was significantly greater (P<0.05) than the number ( \(\bar X\) =1.2) that completed development in 3rd-instar larvae. Maggot development time decreased with increasing number of maggots per host larva. It also decreased with advancing larval instars. The range was 6.9±1.1 days in early 3rd-instar TBW larvae and 5.0±0.8 days in early 5th-instar TBW larvae. Unparasitized 3rd- or 4th-instar TBW larvae consumed significantly more food than did similar aged larvae parasitized byEucelatoria sp., but larvae parasitized during the early 5th-instar consumed more food than did similar aged unparasitized larvae. Consumption by 4th- or 5th-instar larvae increased significantly as maggot densities increased from 1 to 3 per host larva, but decreased at a density of 4 or more maggots per host larva. Although body weight gain and consumption were both significantly reduced 48 and 120 h after parasitization of late 3rd-instar larvae (6 days old), the approximate digestibility (AD) value was significantly greater for parasitized than for unparasitized larvae. Unparasitized larvae were more efficient in converting digested food to body substance (ECD) than parasitized larvae, but the efficiency in conversion of ingested food to body substance (ECI) was similar for both parasitized and unparasitized larvae.     

Streambed microstructure predicts evolution of development and life history mode in the plethodontid salamander Eurycea tynerensis

Background

Habitat variation strongly influences the evolution of developmentally flexible traits, and may drive speciation and diversification. The plethodontid salamander Eurycea tynerensis is endemic to the geologically diverse Ozark Plateau of south-central North America, and comprises both strictly aquatic paedomorphic populations (achieving reproductive maturity while remaining in the larval form) and more terrestrial metamorphic populations. The switch between developmental modes has occurred many times, but populations typically exhibit a single life history mode. This unique system offers an opportunity to study the specific ecological circumstances under which alternate developmental and life history modes evolve. We use phylogenetic independent contrasts to test for relationships between a key microhabitat feature (streambed sediment) and this major life history polymorphism.

Results

We find streambed microstructure (sediment particle size, type and degree of sorting) to be highly correlated with life-history mode. Eurycea tynerensis is paedomorphic in streams containing large chert gravel, but metamorphoses in nearby streams containing poorly sorted, clastic material such as sandstone or siltstone.

Conclusion

Deposits of large chert gravel create loosely associated streambeds, which provide access to subsurface water during dry summer months. Conversely, streambeds composed of more densely packed sandstone and siltstone sediments leave no subterranean refuge when surface water dries, presumably necessitating metamorphosis and use of terrestrial habitats. This represents a clear example of the relationship between microhabitat structure and evolution of a major developmental and life history trait, and has broad implications for the role of localized ecological conditions on larger-scale evolutionary processes.     

Linking larval history to juvenile demography in the bicolor damselfish Stegastes partitus (Perciformes: Pomacentridae)

Otolith-based reconstructions of daily larval growth increments were used to examine the effect of variation in larval growth on size and age at settlement and post-settlement growth, survival and habitat preferences of juvenile bicolor damselfish (Stegastes partitus Poey). During August 1992 and 1994, newly settled S. partitus were collected from Montastraea coral heads and Porites rubble piles in Tague Bay, St. Croix, U.S. Virgin Islands (17 degrees 45'N, 64 degres 42' W). Daily lapillar otolith increments from each fish were counted and measured with Optimas image analysis software. S. partitus pelagic larval duration was 23.7 d in 1992 (n = 70) and 24.6 d in 1994 (n = 38) and larval age at settlement averaged 13.0 mm total length both years. Analysis of daily otolith increments demonstrated that variation in larval growth rates and size and age at settlement had no detectable effect on post-settlement survivorship but that larger larvae showed a preference for Montastraea coral at settlement. Late larval and early juvenile growth rates showed a significant positive relationship indicating that growth patterns established during the planktonic stage can span metamorphosis and continue into the benthic juvenile phase. Larval growth rates during the first two weeks post-hatching also had a strong effect on age to developmental competence (ability to undergo metamorphosis) in both 1992 and 1994 with the fastest growing larvae being 8 d younger and 0.8 mm smaller at settlement than the slowest growing larvae. These differential growth rates in early stage larvae established trajectories toward larval developmental competence and may prove important in biogeographical studies of larval dispersal.     

Induction of larval settlement and metamorphosis in the donkey-ear abalone,Haliotis asinina Linnaeus,by chemical cues

The effects of the chemical inducers, gamma-aminobutyric acid (GABA) and potassium chloride (KCl), on the larval settlement and metamorphosis of the donkey-ear abalone, Haliotis asinina, was investigated. H. asinina larvae (5–6 h post-hatch) were exposed to a range of GABA (0.125–2.00 μM) and KCl (1.00–12.00 mM) concentrations for 72 h. Results of the dose response experiments showed that settlement and metamorphosis vary according to the dose levels of the inducer compounds. Under controlled laboratory conditions, 0.45–0.50 μM and 6.0 mM seemed to be the optima for GABA and KCl, respectively, as these concentrations elicited the greatest number of postlarvae that metamorphosed, settled or survived. However, GABA generally promoted better attachment and metamorphic response as well as survival than KCl in H. asinina postlarvae.     

Neural system reorganization during metamorphosis in the planula larva of Clava multicornis (Hydrozoa, Cnidaria)

The planula larva of the hydroid Clava multicornis (Forskål, 1775) has a complex nervous system, characterized by the presence of distinct, anteriorly concentrated peptidergic populations of amidated neurons, presumably involved in the detection of environmental stimuli and metamorphic signals. Differently from other hydrozoan larvae in C. multicornis planulae GLW-positive cells with putative sensory role have a peculiar dome-shaped forefront organization, followed by a belt of RF-positive nerve cells. By immunohistochemistry, we investigated the transformation of the peptidergic (GLW-amide and RF-amide) larval neuroanatomy at different stages of metamorphosis and the subsequent development of the primary polyp nervous system. By terminal transferase-mediated dUTP nick end-labeling assay, apoptotic nuclei were first identified in the anterior pole of the settled larva, in the same region occupied by GLW-amide positive putative sensory cells. In primary polyps, GLW-amide positive signals first encircled the hypostome area, later extending downwards along the polyp column or upwards over the hypostome dome, whereas RF-amide positive sensory cells initially appeared at the tentacles base to later extend in the tentacles and the polyp column. In spite of the possession of distinct neuroanatomies, different cnidarian planulae may share common developmental mechanisms underlying metamorphosis, including apoptosis and de novo differentiation. Our data confirm the hypothesis that the developmental dynamics of tissue rearrangements may be not uniform across different taxa.