Sea Urchin Larvae Research Articles

Induction to maturation of the sea urchin Paracentrotus lividus (Lamarck, 1816) under laboratory conditions

Laboratory experiments were carried out over two different periods, with the aim of investigating and utilizing the induction to gonadal maturation of the sea urchin Paracentrotus lividus. The final objective was to obtain viable gametes outside the period of natural spawning in the environment; which would allow the utilization of ecotoxicological bioassays with sea urchin larvae at any time of the year. The experiment consisted of maintaining some sea urchins in tanks and providing them with a natural photoperiod, unlimited food and a constant temperature of 20 °C. During days 0, 30, 60 and 90, gonads from 15 of these sea urchins were compared with those collected simultaneously in the natural environment. The gametes obtained were used to carry out fecundations, in order to check their viability. The final results obtained were clearly influenced by the gonadal state of the sea urchins at the initial stage of the experiment. The best results were obtained within a time period of 60 days and when the initial gonad index was low.

Dose-dependent effects of chlorpyriphos, an organophosphate pesticide, on metamorphosis of the sea urchin, Paracentrotus lividus

The effect of exposures to the insecticide chlorpyrifos on the larval stages of Paracentrotus lividus (Echinodermata, Euechinoidea) up to metamorphosis was investigated with the aim to identify novel risk biomarkers and a new promising model for toxicity tests. The planktonic sea urchin larvae have the ability to undergo a variable exploratory period, up to the choice of a suitable substrate for adult benthonic life. The juvenile bud (called rudiment) is built inside the larval body that, on environmental cues represented by a variety of signal molecules, is reabsorbed by apoptosis and releases the juvenile on the substrate. In this dialogue between larvae and environment, contaminants interfere with the signals reception, and may alter in dose-dependent way the correct regulation of environment-larva-rudiment interaction. Such interaction is shown by larval plasticity, i.e. the ability of the larva to change body proportions according to the environmental conditions. When exposed to low doses of chlorpyriphos (10(-7) to 10(-10) M) since 2-days after fertilization, the larvae showed altered size and shape, but all reached the metamorphosis at the same time as controls, and in the same percentage. Exposures to high concentrations such as 10(-4) to 10(-6) M since 2-days after fertilization did not allow larval growth and differentiation. Exposures at later stages caused reabsorption of larval structures within a few hours and precocious release of the immature rudiments, followed by death of the juveniles. Although the mechanism of chlorpyriphos toxicity in sea urchin larvae is still rather unclear, the measurable stress biomarkers can constitute the basis for new toxicity tests.

Evolutionary and experimental change in egg volume, heterochrony of larval body and juvenile rudiment, and evolutionary reversibility in pluteus form

Heterochronic developmental plasticity of the juvenile rudiment and larval body of sea urchin larvae occurs in response to supply of food. Evolutionary increase in egg size can also be associated with earlier development of the juvenile rudiment. We examined effects of egg volume of feeding larvae on this heterochrony and other changes in larval form. (1) Evolutionary and experimental enlargements of egg volume did not accelerate formation of the rudiment relative to the larval body. Development of the larval body and juvenile rudiment was compared for the echinoids Strongylocentrotus purpuratus (with an egg of 78-82 microm) and Strongylocentrotus droebachiensis (with an egg of 150-160 microm diameter). Development of both larval body and rudiment were accelerated in S. droebachiensis relative to S. purpuratus but with greater acceleration of the larval body, so that the rudiment of S. droebachiensis was initiated at a later larval stage even though at an earlier age. Also, experimentally doubling the egg volume of S. purpuratus did not accelerate development of the juvenile rudiment relative to the larval body. (2) Both species exhibited similar plasticity in timing of rudiment development in response to food supplies. (3) Doubling egg volume of S. purpuratus produced a larval form more similar to that of S. droebachiensis. This result mirrors previous experiments in which larvae from half embryos of S. droebachiensis were more similar to larvae of S. purpuratus. Many of the effects of egg volume on larval form are similar against either species' genetic background and are thus evolutionarily reversible effects on larval form.

Endocrine interactions between plants and animals: Implications of exogenous hormone sources for the evolution of hormone signaling

Hormones are central to animal physiology, metabolism and development. Details on signal transduction systems and regulation of hormone synthesis, activation and release have only been studied for a small number of animal groups, notably arthropods and chordates. However, a significant body of literature suggests that hormonal signaling systems are not restricted to these phyla. For example, work on several echinoderm species shows that exogenous thyroid hormones (THs) affect larval development and metamorphosis and our new data provide strong evidence for endogenous synthesis of THs in sea urchin larvae. In addition to these endogenous sources, these larvae obtain THs when they consume phytoplankton. Another example of an exogenously acquired hormone or their precursors is in insect and arthropod signaling. Sterols from plants are essential for the synthesis of ecdysteroids, a crucial group of insect morphogenic steroids. The availability of a hormone or hormone precursor from food has implications for understanding hormone function and the evolution of hormonal signaling in animals. For hormone function, it creates an important link between the environment and the regulation of internal homeostatic systems. For the evolution of hormonal signaling it helps us to better understand how complex endocrine mechanisms may have evolved.

Standardisation du développement larvaire de l'oursin, Paracentrotus lividus, pour l'évaluation de la qualité d'une eau de mer

All stages of development of the sea urchin are of interest in ecotoxicology; the largest number of prior works concerns studies on gametes. Previous studies indicated that the use of sea urchin larvae was difficult because of the need to obtain the parent generations and good quality gametes. Progress in sea urchin culture has allowed one to standardize the method. The proposed technique is based on an evaluation of the number of non-developing fertilized eggs, on the frequency of malformations, and on the length of the larvae at 96 hours, using parents raised under well-controlled conditions. Temperature (18–22 °C), salinity (28–34 ppt) and pH (8–8.4) have been fixed to standardize the proposed biological test. Thirty micrograms per litre of copper reduce significantly the length of the larvae and could be used as a positive control. On the other hand, reconstituted sea water permits an optimal development of the larvae and may be used as negative control. A seasonal follow-up of water quality has been achieved to validate the use of this technique in a surveillance network of water quality. The method may be used whatever the salinity, including fresh and brackish waters. To cite this article: S. Pétinay et al., C. R. Biologies 332 (2009).

Predicting water toxicity: Pairing passive sampling with bioassays on the Great Barrier Reef

Many coral reefs worldwide occur adjacent to urban or agricultural land which places these ecosystems at threat of exposure to complex mixtures of pollutants. In this study, the pairing of passive sampler extracts with bioassays is proposed as a tool for predicting effects of organic pollutant mixtures on key biota within coral reef ecosystems. Passive samplers, SDB-RPS Empore™ disks, which sequester a mixture of the contaminants present in the environment, were deployed at three sites in the Great Barrier Reef (GBR). Extracts from these samplers were analysed for herbicides and applied to bioassays targeting integral life stages or functions of coral reef biota. Biota included scleractinian coral larvae, sea urchin larvae, a marine diatom and marine bacteria. Photosynthesis in the marine diatom Phaeodactylum tricornutum was inhibited at the sampled environmental concentration while an environmental concentration factor of 15 times inhibited luminescence in the marine bacterium Vibrio fischeri. Concentrations of 50 times sampled environmental levels of organic pollutants inhibited >90% of Acropora millepora settlement and 100-fold environmental enrichment inhibited 100% Heliocidaris tuberculata larval development. These results demonstrate the utility of pairing passive sampling with bioassays and reveal that mixtures of organic pollutants in the GBR have the potential to cause detrimental effects to coral reef biota.

Transcriptomic response of sea urchin larvaeStrongylocentrotus purpuratusto CO2-driven seawater acidification

Ocean acidification from the uptake of anthropogenic CO(2) is expected to have deleterious consequences for many calcifying marine animals. Forecasting the vulnerability of these marine organisms to climate change is linked to an understanding of whether species possess the physiological capacity to compensate for the potentially adverse effects of ocean acidification. We carried out a microarray-based transcriptomic analysis of the physiological response of larvae of a calcifying marine invertebrate, the purple sea urchin, Strongylocentrotus purpuratus, to CO(2)-driven seawater acidification. In lab-based cultures, larvae were raised under conditions approximating current ocean pH conditions (pH 8.01) and at projected, more acidic pH conditions (pH 7.96 and 7.88) in seawater aerated with CO(2) gas. Targeting expression of approximately 1000 genes involved in several biological processes, this study captured changes in gene expression patterns that characterize the transcriptomic response to CO(2)-driven seawater acidification of developing sea urchin larvae. In response to both elevated CO(2) scenarios, larvae underwent broad scale decreases in gene expression in four major cellular processes: biomineralization, cellular stress response, metabolism and apoptosis. This study underscores that physiological processes beyond calcification are impacted greatly, suggesting that overall physiological capacity and not just a singular focus on biomineralization processes is essential for forecasting the impact of future CO(2) conditions on marine organisms. Conducted on targeted and vulnerable species, genomics-based studies, such as the one highlighted here, have the potential to identify potential ;weak links' in physiological function that may ultimately determine an organism's capacity to tolerate future ocean conditions.

Sea urchin arylsulfatase, an extracellular matrix component, is involved in gastrulation during embryogenesis

Arylsulfatases (Arses) have been regarded as lysosomal enzymes because of their hydrolytic activities on synthetic aromatic substrates and their lysosomal localization of their enzymatic activities. Using sea urchin embryos, we previously demonstrated that the bulk of Hemicentrotus Ars (HpArs) does not exhibit enzyme activity and is located on the apical surface of the epithelial cells co-localizing with sulfated polysaccharides. Here we show that HpArs strongly binds to sulfated polysaccharides and that repression of the synthesis by HpArs-morpholino results in retardation of gastrulation in the sea urchin embryo. Accumulation of HpArs protein and sulfated polysaccharides on the apical surface of the epithelial cells in sea urchin larvae is repressed by treatment with beta-aminopropionitrile (BAPN), suggesting that deposition of HpArs and sulfated polysaccharides is dependent on the crosslinking of proteins such as collagen-like molecules. We suggest that HpArs functions by binding to components of the extracellular matrix.

Overview of the amorphous precursor phase strategy in biomineralization

It was assumed for a long time that organisms produce minerals directly from a saturated solution. A few exceptions were known, including the well documented mineralized teeth of the chiton. In 1997 it was demon-strated that sea urchin larvae form their calcitic spicules by first depositing a highly unstable mineral phase called amorphous calcium carbonate. This strategy has since been shown to be used by animals from other phyla and for both aragonite and calcite. Recent evidence shows that vertebrate bone mineral may also be formed via a precursor phase of amorphous calcium carbonate. This strategy thus appears to be widespread. The challenge now is to understand the mechanisms by which these unstable phases are initially formed, how they are temporarily stabilized and how they are destabilized and transform into a crystalline mature product.

Fluorescent in situ hybridization reveals multiple expression domains for SpBrn1/2/4 and identifies a unique ectodermal cell type that co-expresses the ParaHox gene SpLox

Here we report on the expression of two transcription factors previously reported to be involved in endodermal patterning, SpLox and SpBrn1/2/4. We describe three distinct domains of expression of the pou-domain gene SpBrn1/2/4. Endodermal expression of this gene is restricted to the foregut. SpBrn1/2/4 is also expressed in two distinct ectodermal domains: throughout the stomodeal ectoderm, and within cells scattered throughout the ciliated band. The ParaHox gene SpLox is demonstrated to also be expressed in ectodermal bilateral cell pairs in prism and early pluteus stage larvae. Double fluorescent in situ hybridization reveals that these cell pairs co-express both SpLox and SpBrn1/2/4, thus marking a novel cell type within the ciliary band of the sea urchin larvae.

Predicted impact of ocean acidification on a marine invertebrate: elevated CO2 alters response to thermal stress in sea urchin larvae

Ocean acidification (OA) and the biological consequences of altered seawater chemistry have emerged as a significant environmental threat to healthy marine ecosystems. Because a more acidic ocean interferes with fixation of calcium carbonate to form shells or calcified skeletons, future ocean chemistry may significantly alter the physiology of calcifying marine organisms. These alterations may manifest themselves directly in the calcification process, or have synergistic effects with other environmental factors such as elevated temperatures. New tools permit us to explore subtle changes in gene expression patterns in response to environmental conditions. We raised sea urchins (Strongylocentrotus franciscanus) under conditions simulating future atmospheric CO2 levels of 540 and 970 ppm. When larvae raised under elevated CO2 conditions were subjected to 1-h acute temperature stress, their ability to mount a physiological response (as measured by expression of the molecular chaperone hsp70) was reduced relative to those raised under ambient CO2 conditions. These results represent the first use of gene expression assays to study the effects of OA on sea urchin development. They highlight the importance of looking at multiple environmental factors simultaneously as this approach may reveal previously unsuspected biological impacts of atmospheric changes.

Using functional genomics to explore the effects of ocean acidification on calcifying marine organisms

As the research community attempts to forecast the effects of ocean acidification on marine ecosystems, a critical element is a clear understanding of the effects of ocean acidification on an individual organism’s physiology. This article explores how the use of genomics-based tools that measure gene expression — DNA microarrays and quantitative PCR — can assist in this effort and reveal aspects of how calcifying marine organisms will respond to ocean acidification. More specifically, what stands to be gained from this approach is an understanding of the direct effects of ocean acidification and whether organisms have sufficient physiological plasticity to adapt to the altered CO2 conditions. We provide a brief overview of biomineralization processes in corals and sea urchin larvae, and then link these pathways to ways in which gene expression analysis can reveal physiological responses and mechanisms, and further, can define new testable hypotheses. In addition, we review the resources available and strategies that might be taken for each of 2 study organisms, stony corals and sea urchins. Finally, we suggest strategies for gene expression profiling in organisms that differ in availability of genomic resources.

Development of nervous systems to metamorphosis in feeding and non-feeding echinoid larvae, the transition from bilateral to radial symmetry

The development of nervous system (NS) in the non-feeding vestibula larva of the sea urchin, Holopneustes purpurescens, and the feeding echinopluteus larva of Hemicentrotus pulcherrimus was examined by focusing on fate during metamorphosis. In H. purpurescens, the serotonergic NS (SerNS) appeared simultaneously and independently in larval tissue and adult rudiment, respectively, from 3-day post-fertilization. In 4-day vestibulae, an expansive aboral ganglion (450 x 100 mum) was present in the larval mid region that extended axons toward the oral ectoderm. These axons diverged near the base of the primary podia. An axonal bundle connected with the primary podia and the rim of vestopore on the oral side. Thus, the SerNS of the larva innervated the rudiment at early stage of development of the primary podia. This innervation was short-lived, and immediately before metamorphosis, it disappeared from the larval and adult tissue domains, whereas non-SerNS marked by synaptotagmin remained. The NS of 1-month post-fertilization plutei of H. pulcherrimus comprised an apical ganglion (50 x 17 mum) and axons that extended to the ciliary bands and the adult rudiment (AR). A major basal nerve of serotonergic and non-serotonergic axons and a minor non-serotonergic nerve comprised the ciliary band nerve. In 3-month plutei, axonal connection among the primary podia in the neural folds completed. The SerNS never developed in the AR. Thus, there was distinctive difference between feeding- and non-feeding larvae of the above sea urchins with respect to SerNS and the AR.

Salinity sensitivity of early embryos of the Antarctic sea urchin, Sterechinus neumayeri

Embryos and larvae of the Antarctic sea urchin, Sterechinus neumayeri, have received considerable experimental attention assessing impacts of low temperature on development; however, salinity effects are not well documented because heretofore, the Antarctic coastal marine environment has been remarkably stenohaline. In this study, subtle decreases of 2 and 4 parts per 1,000 in standard salinity were tested to see if the developmental rate of S. neumayeri embryos would be impacted by a potential hyposmotic stress. At 30 psu, significantly fewer embryos (2 individuals out of 198 tested) reached morula stage by 36 h post-fertilization in comparison embryos in control treatments at 34 psu. Antarctic sea urchins are an important component of marine environments due to their grazing activities. Reductions in larval recruitment success due to the influx of freshwater from melting ice shelves resulting from global climate change could have far-reaching impacts on benthic ecosystem structure in Antarctica.

Meroditerpenoids and derivatives from the brown alga Cystoseira baccata and their antifouling properties.

The brown alga Cystoseira baccata harvested along the Atlantic coasts of Morocco yielded seven new meroditerpenoids (1-4) and derivatives (5-7), whose chemical structures were elucidated mainly by 2D NMR and mass spectrometry. Surprisingly, for all these compounds, which possess a bicyclo[4.3.0]nonane ring system, a trans fusion of the bicyclic system was deduced by stereochemical studies even though such compounds isolated from Cystoseira species are known to have a typical cis orientation for the bridgehead methyls. The antifouling and antibacterial activities of compounds 1-5 and 7 were evaluated, as well as their toxicity toward nontarget species. Compounds 4, 5, and 7 showed antifouling activities against growth of microalgae, macroalgal settlement, and mussel phenoloxidase activity, while being nontoxic to larvae of sea urchins and oysters.

Bioassays of water-accommodated fractions of No.0 diesel oil using embryos and larvae of sea urchin, Hemicentrotus pulcherrimus

Bioassays of water-accommodated fractions of No.0 diesel oil using embryos and larvae of sea urchin, Hemicentrotus pulcherrimus

Inhibitory effect of 2,4-dibromophenol and 2,4,6-tribromophenol on larval survival and metamorphosis of the sea urchinStrongylocentrotus nudus

As a possible factor leading to the low recruitment level of sea urchins in kelp forests, the inhibitory effect of 2,4-dibromophenol (DBP) and 2,4,6-tribromophenol (TBP) released from the large perennial brown algae Ecklonia kurome and Eisenia bicyclis on survival and metamorphosis of eight-armed larvae of the sea urchin Strongylocentrotus nudus was examined. The percentage of larvae that underwent metamorphosis in filtered sea water after 1 h exposure to one-half dilution of saturated dibromomethane solution (≈60 ppm) as a chemical inducer reached approximately 100% after 1 h, while that in filtered sea water containing 1 ppm TBP was reduced to 73%. This was further reduced to less than 40% in the presence of 10 and 20 ppm TBP after 2 h. In filtered sea water containing 1 and 10 ppm DBP, the proportion of metamorphosed larvae was reduced markedly to 43 and 5% after 2 h, respectively. All larvae exposed to 50 ppm TBP and to 20 and 50 ppm DBP died after 1 h. These findings suggest that DBP is more toxic than TBP for sea urchin larvae, strongly inhibiting their metamorphosis.

Nutritional ecology of sea urchin larvae: influence of endogenous and exogenous nutrition on echinopluteal growth and phenotypic plasticity in Tripneustes gratilla

Summary 1 Marine invertebrates use egg nutrients to develop the functional feeding larva and then enter a facultative feeding period (FFP) when development can proceed without food because larvae are supported by maternal reserves. Facultative feeding reduces starvation risk and so is important for larval success. It may also influence egg size evolution because FFP length correlates with egg size. 2 We quantified energetic lipid (triglyceride, TG) utilisation in unfed and fed larvae as an index of larval nutritive condition in the tropical echinoid Tripneustes gratilla during the FFP to determine if fed larvae would use the buffer provided by endogenous provisions to improve their condition or accelerate development. We predicted that: (i) the condition of unfed and fed larvae should diverge before egg TG is exhausted and/or (ii) that the size of fed larvae should outstrip that of unfed larvae. 3 Temperate echinoplutei exhibit phenotypic plasticity, increasing the length of their food capture apparatus (arms) in nutrient-poor conditions. We examined the generality of this phenomenon in a morphometric analysis of growth in the tropical larva of T. gratilla. We hypothesized that plastic arm growth in starved larvae would occur before the FFP ends as a bet hedging strategy to prepare for hard times ahead when lack of reserves may render this response impossible. 4 We found that fed larvae diverged in condition (higher TG) but not in size compared with starved larvae before egg energetic lipids were exhausted. In addition, unfed larvae showed plastic arm growth before the end of the FFP. 5 Lack of divergence in growth of unfed and fed larvae suggests there may be an imperative to maintain a nutrient storage buffer against starvation due to unpredictable future planktonic food supply. The FFP of T. gratilla exceeds 8 days, considerably longer than that of other echinoids with similarly sized eggs, emphasizing that egg quality may be more important than egg size as a predictor of the FFP in fecundity-time models of egg evolution.

Specification process of animal plate in the sea urchin embryo

The most animal part of the ciliated band of sea urchin larvae, the animal plate, is a specialized region in which elongated cells form long and non-beating cilia. To learn how this region is specified, animal halves were isolated from the early cleavage to pregastrulation stages. As is well known, the animal half that is isolated at the eight-cell stage develops into a 'dauerblastula', which forms long and non-beating cilia all around the surface. The region with long cilia, however, became restricted toward the animal pole when separation was delayed. If separated before primary mesenchyme ingression, even a small animal-pole-side fragment formed a normal-sized animal plate. Thus, the prospective animal plate region is gradually restricted by some signal from the vegetal hemisphere, and the specification process terminates before the mesenchyme blastula stage. It was also known that a normal-sized animal plate was formed in micromere-less embryos, indicating that the signal does not depend on micromeres or their descendants. Further, the animal-pole-side fragments were isolated from embryos in which the third cleavage plane was shifted toward the vegetal pole. They formed a normal-sized animal plate, containing more than 75% of the egg volume from the animal pole. This indicates that the egg cytoplasm delivered to veg1-lineage blastomeres plays an important role in the animal plate specification. Interestingly, the an1-less embryo formed long and non-beating cilia at its top region, but thickening did not occur. The cytoplasm near the animal pole might contain some factors necessary for the animal plate to become thick.

The effect of copper ions on behavior of larvae of the sea urchin Strongylocentrotus intermedius at different temperatures of sea water

We investigated the behavior of larvae of the sea urchin Strongylocentrotus intermedius at the blastula and early pluteus stages in the water column at a temperature of 20, 22, and 23.5°C and in the presence of copper in concentrations of 0.01 and 0.02 mg/l. In clean sea water, at all tested temperatures, sea urchin larvae preferred the subsurface layer to where they rose from the bottom of the vessel to form dense aggregations. In water supplemented with copper, the behavior of larvae at 20 and 22°C was not different from that in clean water. An increase in temperature to 23.5°C and the addition of copper altered the behavior of the larvae; they did not leave near-bottom water, did not rise to the surface, although they continued moving.