Demonstration of the granular layer and the fate of the hyaline layer during the development of a sea urchin (Lytechinus variegatus)

Summary Employing electron-microscopic methods that help retain polyanionic materials, we describe the extracellular coverings of a sea urchin (Lytechinus variegatus) throughout ontogeny. The surface of the embryo is covered by a two-layered cuticle (commonly called the hyaline layer), which in turn is covered by a granular layer. The granular layer is retained after addition of alcian blue to the fixative solutions, and has not been previously described for any sea urchin. After hatching, the granular layer disappears, but the hyaline layer continues to cover most of the larval surface until settlement and metamorphosis. A few days before metamorphosis, the hyaline layer lining the vestibular invagination of the competent pluteus larva is replaced by a three-layered cuticle resembling that of the adult sea urchin. The hyaline layer covering the rest of the larva is evidently lost at metamorphosis during the involution of the general epidermis.     

Embrionary and larval development of Lytechinus variegatus (Echinoidea: Toxopneustidae) in laboratory conditions at Isla de Margarita-Venezuela

The sea urchin Lytechinus variegatus is a promissory species for aquaculture activities in tropical countries. In Venezuela, this species has some economical importance but their embryonic and larval development had not been studied. We collected specimens from seagrass beds in Margarita Island (Venezuela) and kept them in the laboratory, where they spawned naturally. With filtered sea water (temperature 28 degrees C, salinity 37 psu) and moderate aeration, the eggs and sperm were mixed (relation 1:100) and reached a 90% fertilization rate. The fertilization envelope was observed after two minutes, the first cellular division after 45 minutes and the prism larval stage after 13 hours. The echinopluteus larval stage was reached after 17 hours and metamorphosis after 18 days of planktonic life, when the larvae start their benthic phase.     

A switch in the cellular basis of skeletogenesis in late-stage sea urchin larvae

Primary mesenchyme cells (PMCs) are solely responsible for the skeletogenesis during early larval development of the sea urchin, but the cells responsible for late larval and adult skeletal formation are not clear. To investigate the origin of larval and adult skeletogenic cells, I first performed transplantation experiments in Pseudocentrotus depressus and Hemicentrotus pulcherrimus, which have different skeletal phenotypes. When P. depressus PMCs were transplanted into H. pulcherrimus embryos, the donor phenotype was observed only in the early larval stage, whereas when secondary mesenchyme cells (SMCs) were transplanted, the donor phenotype was observed in late and metamorphic larvae. Second, a reporter construct driven by the spicule matrix protein 50 (SM50) promoter was introduced into fertilized eggs and their PMCs/SMCs were transplanted. In the resultant 6-armed pluteus, green fluorescent protein (GFP) expression was observed in both PMC and SMC transplantations, suggesting SMC participation in late skeletogenesis. Third, transplanted PMCs or SMCs tagged with GFP were analyzed by PCR in the transgenic chimeras. As a result, SMCs were detected in both larval and adult stages, but GFP from PMCs was undetectable after metamorphosis. Thus, it appears that SMCs participate in skeletogenesis in late development and that PMCs disappear in the adult sea urchin, suggesting that the skeletogenesis may pass from PMCs to SMCs during the late larval stage.     

Larvae and postlarvae production of the green-white sea urchin Lytechinus variegatus (Echinodermata: Echinoidea) in culture conditions

We evaluated the biological feasibility of massive larvae and postlarvae production of the Caribbean green-white urchin Lytechinus variegatus. Experiments were designed to choose the initial larval density and microalgae diets under culture, to study metamorphosis, postlarval and juvenile growth. Massive production of competent larvae 650 microm long at 12-13 days is possible using larval densities of 0.25 to 1 larva/ml. The microalgae Rhodomonas sp. was suitable for the optimization of larval growth and survival. Metamorphosis of 100% of the larvae can be induced with films of bentic diatoms (Navicula sp. and Amphora sp.), after 96 h; however, diatoms are not adequate for postlarval development and a food supply of Ulva lactuta is necessary for proper growth. For juveniles, a diet of macroalgae (U. lactuta) and/or commercial marine shrimp culture pellet food is enough for growth, but the best results were obtained with shrimp or U. lactuta used alone (85-86%, against 46% with the mixed diet). We recommend future experiments on nutritional requirements to optimize growth of these and subsequent stages.     

A PROPOSAL FOR A NEW RED ALGAL ORDER, THE THOREALES

Two abalone species: green Haliotis fulgens and yellow Halioti corrugata represent nearly 97% of the total production in the Mexican abalone fishery. It has been assumed that abalone feed on the kelp algae Macrocystis pyrifera. Regional hatcheries use this species as a main source of natural food. M. pyrifera does not occur at the southern limit of the distribution of abalone species along the Baja California Peninsula. In this study, growth rates of juveniles H. fulgens , 17.3 ± 2.2 mm shell length and 0.4 ± 0.2 g body weight, were evaluated. Juveniles were fed with common species in the benthic environments inhabited by abalone along the western coast of Baja California during 191 days. Three diets were based on algae: palm kelp, Eisenia arborea , giant kelp, M. pyrifera and Gelidium robustum , and one on seagrass, Phyllospadix torreyi. Shell length and body growth rates varied between 21.5 μm day⁻¹ and 2.2 mg day⁻¹ for E. arborea and between 45.9 μm day⁻¹ and 6.7 mg day⁻¹ for M. pyrifera. Higher specific growth rates (SGR) in length and weight were determined for M. pyrifera : 0.2% and 0.7% day⁻¹. Significant differences between values of juveniles fed M. pyrifera with the rest of the diets were found. The highest mortality (21%) was in juveniles fed the red algae G. robustum.     

Trophic Cascades Induced by Lobster Fishing Are Not Ubiquitous in Southern California Kelp Forests

Fishing can trigger trophic cascades that alter community structure and dynamics and thus modify ecosystem attributes. We combined ecological data of sea urchin and macroalgal abundance with fishery data of spiny lobster (Panulirus interruptus) landings to evaluate whether: (1) patterns in the abundance and biomass among lobster (predator), sea urchins (grazer), and macroalgae (primary producer) in giant kelp forest communities indicated the presence of top-down control on urchins and macroalgae, and (2) lobster fishing triggers a trophic cascade leading to increased sea urchin densities and decreased macroalgal biomass. Eight years of data from eight rocky subtidal reefs known to support giant kelp forests near Santa Barbara, CA, USA, were analyzed in three-tiered least-squares regression models to evaluate the relationships between: (1) lobster abundance and sea urchin density, and (2) sea urchin density and macroalgal biomass. The models included reef physical structure and water depth. Results revealed a trend towards decreasing urchin density with increasing lobster abundance but little evidence that urchins control the biomass of macroalgae. Urchin density was highly correlated with habitat structure, although not water depth. To evaluate whether fishing triggered a trophic cascade we pooled data across all treatments to examine the extent to which sea urchin density and macroalgal biomass were related to the intensity of lobster fishing (as indicated by the density of traps pulled). We found that, with one exception, sea urchins remained more abundant at heavily fished sites, supporting the idea that fishing for lobsters releases top-down control on urchin grazers. Macroalgal biomass, however, was positively correlated with lobster fishing intensity, which contradicts the trophic cascade model. Collectively, our results suggest that factors other than urchin grazing play a major role in controlling macroalgal biomass in southern California kelp forests, and that lobster fishing does not always catalyze a top-down trophic cascade.     

Changes in transferrin during the red cell replacement in amphibia

Transferrin, a plasma glycoprotein, carries iron from storage sites to immature erythroid cells for hemoglobin synthesis. The replacement of larval red cells by adult red cells, which occurs during metamorphosis in bullfrogs, requires extensive formation of hemoglobin and new red cells. Large changes in red cell iron storage also occur during the red cell replacement. Both the concentration and the level of iron saturation of plasma transferrin were measured during metamorphosis to determine if there were changes in plasma transferrin which coincided with the changes in red cell iron storage and ferritin content. Plasma transferrin concentrations increased from 0.96 to 2.6 mg/ml during the period when red cell storage iron and ferritin decreased. Plasma iron concentrations also increased when the transferrin concentration increased, suggesting that the additional transferrin may be involved in moving iron from the larval red cell stores. At the end of metamorphosis, the plasma iron concentration decreased to premetamorphic levels but the transferrin concentration remained high, resulting in a decrease in saturation to 18% compared to 45% in the larvae. In addition to differences in iron saturation, adult transferrin had different electrophoretic properties from larval transferrin. The results support the hypotheses that during early ontogeny plasma transferrin and red cell iron storage are coordinated to provide iron for the formation of the first generation of adult red cells and that transferrin may participate in the control of red cell ferritin synthesis.     

Lunar phase and catch success of the striped marlin (Tetrapturus audax) in sport fishing at Los Cabos, Baja California Sur, Mexico

The influence of the lunar phases on the catch-per-unit effort (CPUE) of the striped marlin (Tetrapturus audax) captured by the Cabo San Lucas, Baja California Sur, Mexico sport-fishing fleet from October 1987 to June 1989 was analyzed. The information is from 3,377 fishing trips by 13 vessels that represent about 10% of the fleet. The analysis of the CPUE showed a maximum in January 1988 and a minimum in February 1989. Taking into account the knowledge of the factors that had influence on the fishing success is important in the resource management. No significant difference during the full moon compared with results during other lunar phases was found.     

Early events in sea urchin metamorphosis,description and analysis

The larval epithelium of the sea urchin, Lytechinus pictus, consists of squamous cells and bands of columnar epithelial cells bearing cilia. During metamorphosis this tissue undergoes a series of rapid, complex changes. Through the scanning and transmission electron microscope, we describe and analyse these changes. The changes can be divided into three steps. (1) The larval arms bend away from the left side of the larva, exposing the urchin rudiment. Cells which are identical to smooth muscle cells are in a position to bring about this bending. (2) The squamous epithelial cells assume a cuboidal shape. This change in shape results in the collapse of the larval epithelium onto the presumptive aboral surface. These cells possess a subapical band of microfilaments. The cellular shape change but not the bending of the arms is reversibly inhibited by Cytochalasin B. These observations suggest a mechanism for this change. (3) The former lining of the vestibule of the urchin rudiment comes to lie over the collapsed larval tissue and forms the adult epithelium. At this point, after only one hour, the larva has assumed the external shape of an adult sea urchin.     

Kelp and sea urchin settlement mediated by biotic interactions with benthic coralline algal species

Species interactions can influence key ecological processes that support community assembly and composition. For example, coralline algae encompass extensive diversity and may play a major role in regime shifts from kelp forests to urchin-dominated barrens through their role in inducing invertebrate larval metamorphosis and influencing kelp spore settlement. In a series of laboratory experiments, we tested the hypothesis that different coralline communities facilitate the maintenance of either ecosystem state by either promoting or inhibiting early recruitment of kelps or urchins. Coralline algae significantly increased red urchin metamorphosis compared with a control, while they had varying effects on kelp settlement. Urchin metamorphosis and density of juvenile canopy kelps did not differ significantly across coralline species abundant in both kelp forests and urchin barrens, suggesting that recruitment of urchin and canopy kelps does not depend on specific corallines. Non-calcified fleshy red algal crusts promoted the highest mean urchin metamorphosis percentage and showed some of the lowest canopy kelp settlement. In contrast, settlement of one subcanopy kelp species was reduced on crustose corallines, but elevated on articulated corallines, suggesting that articulated corallines, typically absent in urchin barrens, may need to recover before this subcanopy kelp could return. Coralline species differed in surface bacterial microbiome composition; however, urchin metamorphosis was not significantly different when microbiomes were removed with antibiotics. Our results clarify the role played by coralline algal species in kelp forest community assembly and could have important implications for kelp forest recovery.     

Myostatin precursor is present in several tissues in teleost fish: a comparative immunolocalization study

In this study, the distribution of myostatin was investigated during larval and postlarval developmental stages of Sparus aurata(sea bream), Solea solea(sole) and Brachydanio rerio(zebrafish) by immunohistochemistry using antisera raised against a synthetic peptide located within the precursor region of sea bream myostatin. All the three species examined showed the strongest immunoreactivity in red skeletal muscle in juveniles and adults. During larval development of sea bream, strong staining was detected in skin and brain. Immunoreactivity was also found in muscle, pharynx, gills, pancreas and liver. From metamorphosis, immunoreactivity was identifiable in the oesophagus, in the apical portion of the stomach epithelium, in the intestinal epithelium and in renal tubules. In larval zebrafish at hatching, the most intense myostatin immunoreactivity was evident in the skin epithelium. Immunoreactivity was also found in the retina and brain. In the adult, an intense immunostaining occurred in the gastrointestinal tract as well as in the ovary. In sole larvae, immunoreactivity was found in liver and intestine. Our results support the hypothesis suggested earlier that myostatins in fish have retained a different partition (compared with mammals) of the expression patterns and functions which characterized the ancestral gene before the duplication event that gave rise to growth differentiation factor-11 (GDF-11) and GDF-8 (myostatin).     

Larval survival and growth of Arbacia punctulata (Echinodermata: Echinoidea) fed with five micro-algae at two salinities

Fertilized eggs from an spontaneously spawn of thirty sexually mature sea urchins (Arbacia punctulata) were incubated to complete embryonic development. The echinopluteus larvae (3 ind/ml) were distributed into 50 plastic containers (25 containers at 30 psu and 25 containers at 40 psu) and fed on Tetraselmis chuii, Nannochloropsis oculata, Isochrysis galbana, Chaetoceros gracilis and C. calcitrans under a natural photoperiod. The water of the containers was partially renewed (75%) everyday. Larval anatomic development aspects, daily survival and growth were determined. The growth was determined through postoral arms and body length measurement, and body diameter of twelve larvae during metamorphosis. During the planktonic larval phase, only the I. galbana diet produced similar results for both salinities. The relative growth of larvae was isometric (I) for larvae fed on I. galbana at two salinities and positive allometric for those fed on C. gracilis and C. calcitrans at both salinities. In this study A. punctulata started metamorphosis at day 14 and was completed 30 days after fecundation. Significant differences were detected in post-settlement body growth between the two salinities (F = 23.58, p < 0.05): growth was better for larvae at 30 psu (final body diameter was 3.14 +/- 0.44 mm). The final rate of planktonic larvae was highest with I. galbana (58.33%). For juveniles the rate was 6.48% for those fed on C. gracilis (40 psu in both larvae and juveniles). We recommend the use of this diet and 40 psu for survival or 30 psu for growth.     

Development of serotonin-like and SALMFamide-like immunoreactivity in the nervous system of the sea urchin Psammechinus miliaris.

The present immunocytochemical study utilizes serotonin and SALMFamide antisera, together with confocal laser scanning microscopy, to provide new information about the development of the nervous system in the sea urchin Psammechinus miliaris (Echinodermata: Echinoidea). Special attention is paid to the extent of the nervous system in later larval stages (6-armed pluteus to metamorphic competency), a characteristic that has not been well described in this and other species of sea urchin. An extensive apical ganglion appears by the 6-armed pluteus stage, forming a complex of 10-20 cells and fibers, including discrete populations of both serotonin-like and SALMF-amide-like immunoreactive cells. At metamorphosis this complex is large, comprising at least 40 cells in distinct arrays. Serotonin-like immunoreactivity is also particularly apparent in the lower lip ganglion of 6- to 8-armed plutei; this ganglion consists of 15-18 cells that are distributed around the mouth. The ciliary nerves that lie beneath the ciliary bands in the larval arms, the esophagus, and a hitherto undescribed network associated with the pylorus all show SALMFamide-like immunoreactivity. The network of cells and fibers in the pyloric area develops later in larval life. It first appears as one cell body and fiber, then increases in size and complexity through the 8-armed pluteus stage to form a complex of cells that encircles the pylorus. SALMFamide-like, but not serotonin-like, immunoreactivity is seen in the vestibule wall, tube feet, and developing radial nerve fibers of the sea urchin adult rudiment as the larva gains metamorphic competency.     

SEA OTTERS AND BENTHIC PREY COMMUNITIES IN WASHINGTON STATE

An August 1987 benthic survey of otter-free and otter-occupied areas along the outer coast of Washington State's Olympic Peninsula confirms that this area has been as profoundly influenced by sea otters as other rocky, nearshore communities studied in California, Canada, and Alaska. Prey density, size, and biomass were found to be negatively correlated with sea otter abundance, suggesting that the re-introduction of sea otters to this area in 1969–1970 has profoundly affected invertebrate prey abundance and distribution, particularly that of the red sea urchin, Strongylocentrotus franciscanus. Red urchin distribution appears to influence algal groups differently and in a manner consistent with current otter/urchin/kelp theory. Foliose red algal abundance was negatively related to urchin numbers and coralline crusts were positively correlated. Aerial photographs of Macrocystis integrifolia cover at Cape Alava suggest an increase since the introduction of sea otters. Given the present distribution of prey along the Olympic Peninsula coast, we conclude that as the sea otter population continues to grow, range expansion is more likely to occur to the north, which may also lead to possible conflicts with an increasing sea urchin fishery and Native American set net activity.     

Uranium-series ages of marine terraces,La Paz Peninsula,Baja California Sur,Mexico

Uranium-series dating of coral samples from raised marine terrace deposits between 1.5 and 10 m above sea level in the La Paz Peninsula area, Baja California Sur, yielded ages between 123 ka and 138 ka that are in agreement with previously reported results. The stratigraphy and ages of marine units near the El Coyote Arroyo indicate the presence of two high stands of the sea during the last interglacial or oxygen isotope substage 5e at about 140 ka and 123 ka. Accepting 5 m for the sea level during the last interglacial transgression, we calculate average uplift rates for the marine terraces of about 70 mm/ka and 40 mm/ka. These slow rates of uplift indicate a relative stability of the La Paz peninsula area for the past 140 000 years. In contrast, areas of Baja California affected by major faultf experienced higher rates of uplift. Rockwell et al. (1987) reported vertical uplift rates of 180 to 300 mm/ka at Punta Banda within the Aqua Blanea fault zone in northern Baja California.     

Lower thermal limits to larval development do not predict poleward range limits of the introduced tropical barnacle Megabalanus coccopoma

As the earth's climate has warmed, many tropical species have expanded their ranges poleward and encountered high‐latitude seasonal temperature regimes, in which further permanent expansion is limited by physiological vulnerability to cold temperatures. The barnacle Megabalanus coccopoma is native to shorelines from Baja California to Peru and has been introduced to many locations worldwide, including the southeastern USA. The ability of larvae to develop successfully at local temperatures can be an important factor limiting the spread of invasive species. To determine if cold temperatures limited larval success near the northern range limit of M. coccopoma along the Atlantic southeastern USA coast, we measured lower temperature limits to larval development, examined the effects of temperature on larval growth and energy accumulation, and calculated a larval energy budget to estimate the extent of potential larval dispersal in this region. Larvae were able to develop through metamorphosis at 16°C, which is much colder than sea surface temperatures during the spawning season in their invasive range, making it unlikely northern range limits are set by a lower temperature limit to larval development. Energy budgets suggest that for larvae produced at the northern end of the invasive range, long distance dispersal to sites far poleward of the current range limit is possible. Similar to the findings of the handful of other studies on cold tolerances of tropical marine invertebrate larvae, larvae should be successful far poleward of current adult distributions.     

Apoptosis in early development of the sea urchin, Strongylocentrotus purpuratus

Apoptosis provides metazoans remarkable developmental flexibility by (1) eliminating damaged undifferentiated cells early in development and then (2) sculpting, patterning, and restructuring tissues during successive stages thereafter. We show here that apoptotic programmed cell death is infrequent and not obligatory during early embryogenesis of the purple sea urchin, Strongylocentrotus purpuratus. During the first 30 h of urchin development, fewer than 20% of embryos exhibit any cell death. Cell death during the cleavage stages consists of necrotic or pathological cell death, while cell death during the blastula and gastrula stages is random and predominantly caspase-mediated apoptosis. Apoptosis remains infrequent during the late blastula stage followed by a gradual increase in frequency during gastrulation. Even after prolonged exposure during the cleavage period to chemical stress, apoptosis occurs in less than 50% of embryos and always around the pre-hatching stage. Embryonic suppression of apoptosis through caspase inhibition leads to functionally normal larvae that can survive to metamorphosis, but in the presence of inducers of apoptosis, caspase inhibition leads to deformed larvae and reduced survival. Remarkably, however, pharmacological induction of apoptosis, while reducing overall survival, also significantly accelerates development of the survivors such that metamorphosis occurs up to a week before controls.     

Relationships between larval nutritional experience, larval growth rates, juvenile growth rates, and juvenile feeding rates in the prosobranch gastropod Crepidula fornicata

Planktonic larvae experiencing short periods of starvation or reduced food supply often grow and develop more slowly, have poor survival, fail to metamorphose, metamorphose at smaller sizes, or grow slowly as juveniles. In this study, we examined the impact of short periods of food limitation at various stages of larval development on larval and juvenile growth in Crepidula fornicata. In addition, we considered whether juveniles that were stressed as larvae grew poorly because of reduced rates of food collection due to impaired gill function. For 5 experiments, larvae were either starved for several days beginning within 12 h of hatching or were starved for the same number of days following 1 or more days of feeding at full ration (cells of the naked flagellate Isochrysis galbana, clone T-ISO, at 18×10⁴ cells ml⁻¹). In one experiment, larvae were transferred for 2 or 4 days to seawater with extremely low phytoplankton concentration (1×10⁴ cells ml⁻¹). In all experiments, larvae were returned to full ration following treatment. Control larvae were fed full ration from hatching to metamorphosis. When larvae reached shell lengths of about 900 μm they were induced to metamorphose and then reared individually at full ration in glass bowls, with phytoplankton suspension replenished daily. Larval and juvenile growth rates were determined by measuring changes in shell length (longest dimension) over time. Juvenile feeding rates were determined by monitoring changes in phytoplankton concentration over 2–3 h at the end of the growth rate determinations. In general, larval growth rates for the first 2 days after the resumption of feeding were inversely proportional to the length of time that larvae were starved. However, larval growth rates ultimately recovered to control levels in most treatments. Starving the larvae caused a significant reduction in initial juvenile growth rates (first 3–4 days post-metamorphosis) in most experiments even when larval growth rates had recovered to control levels prior to metamorphosis. Juvenile growth rates were not significantly reduced when larvae were subjected to reduced food availability (1×10⁴ cells ml⁻¹), even for treatments in which larval growth rates were compromised. Mean weight-specific filtration rates for juveniles were significantly reduced (p<0.05) following larval feeding experience in only one or possibly 2 of the 4 experiments conducted. Our data suggest that although larvae of C. fornicata may fully recover from early nutritional stress, the resulting juveniles may exhibit poor initial growth due to impaired gill function, reduced digestive capability, or reduced assimilation efficiency.