Toxic Effects of Biogenic and Synthesized Silver Nanoparticles on Sea Urchin Echinometra lucunter Embryos.

Due to their broad-spectrum antimicrobial action and ease of synthesis, silver nanoparticles (AgNP) are one of the most widely used nanomaterials in different industrial and ecological areas. AgNP are released into marine ecosystems, nevertheless, their ecotoxicological effects have been overlooked. In this study, we evaluated the toxic effects of biogenic and synthesized AgNP (AgNPIBCLP11 and AgNPSINT) on sea urchin Echinometra lucunter embryos and compared them with the metal precursor silver nitrate (AgNO3). Fertilized eggs were exposed to five concentrations of the test compounds and a negative control for 48 h under controlled conditions. The IC50-48 h of AgNPIBCLP11, AgNPSINT and AgNO3 were 0.31, 4.095, and 0.01 µg L-1, evidencing that both AgNP are less toxic than AgNO3, and that AgNPSINT is less toxic than the AgNPIBCLP11. Toxicity to E. lucunter embryos could be explained by the fact that Ag affects DNA replication and induces the formation of pores in the cellular wall, leading to apoptosis.

Sterol Composition of Sponges, Cnidarians, Arthropods, Mollusks, and Echinoderms from the Deep Northwest Atlantic: A Comparison with Shallow Coastal Gulf of Mexico.

Triterpenoid biosynthesis is generally anaerobic in bacteria and aerobic in Eukarya. The major class of triterpenoids in bacteria, the hopanoids, is different to that in Eukarya, the lanostanoids, and their 4,4,14-demethylated derivatives, sterols. In the deep sea, the prokaryotic contribution to primary productivity has been suggested to be higher because local environmental conditions prevent classic photosynthetic processes from occurring. Sterols have been used as trophic biomarkers because primary producers have different compositions, and they are incorporated in primary consumer tissues. In the present study, we inferred food supply to deep sea, sponges, cnidarians, mollusks, crustaceans, and echinoderms from euphotic zone production which is driven by phytoplankton eukaryotic autotrophy. Sterol composition was obtained by gas chromatography and mass spectrometry. Moreover, we compared the sterol composition of three phyla (i.e., Porifera, Cnidaria, and Echinodermata) collected between a deep and cold-water region and a shallow tropical area. We hypothesized that the sterol composition of shallow tropical benthic organisms would better reflect their photoautotrophic sources independently of the taxonomy. Shallow tropical sponges and cnidarians from environments showed plant and zooxanthellae sterols in their tissues, while their deep-sea counterparts showed phytoplankton and zooplankton sterols. In contrast, echinoids, a class of echinoderms, the most complex phylum along with hemichordates and chordates (deuterostomes), did not show significant differences in their sterol profile, suggesting that cholesterol synthesis is present in deuterostomes other than chordates.

Volume regulation of intestinal cells of echinoderms: Putative role of ion transporters (Na(+)/K(+)-ATPase and NKCC).

Echinoderms are exclusively marine osmoconformer invertebrates. Some species occupy the challenging intertidal region. Upon salinity changes, the extracellular osmotic concentration of these animals also varies, exposing tissues and cells to osmotic challenges. Cells and tissues may then respond with volume regulation mechanisms, which involve transport of ions and water into and/or out of the cells, through ion transporters, such as the Na(+)/K(+)-ATPase and NKCC. The goal of this study was to relate the cell volume regulation capacity of echinoderm intestinal cells Na(+)/K(+)-ATPase and NKCC activities, in three echinoderm species: Holothuria grisea, Arbacia lixula, and Echinometra lucunter. Isolated cells of these species displayed some control of their cell volume upon exposure to anisosmotic media (isolated intestinal cells, calcein fluorescence as indicator of volume change), with a distinct higher capacity shown by H. grisea, which did not swell even upon 50% hyposmotic shock. The holothuroid cells showed indirect evidence (effect of furosemide) of the participation of NKCC in this process, with a secretory function, and of a secondary role by the NKA (effect of ouabain). Other mechanisms are probably responsible for this function in the urchins. Variable expression of these transporters, and others not examined here, may to some extent account for the variability in cell volume regulation capacity in echinoderm cells.

Near-Chromosomal-Level Genome Assembly of the Sea Urchin Echinometra lucunter, a Model for Speciation in the Sea.

Echinometra lucunter, the rock-boring sea urchin, is a widely distributed echinoid and a model for ecological studies of reproduction, responses to climate change, and speciation. We present a near chromosome-level genome assembly of E. lucunter, including 21 scaffolds larger than 10 Mb predicted to represent each of the chromosomes of the species. The 760.4 Mb assembly includes a scaffold N50 of 30.0 Mb and BUSCO (benchmarking universal single-copy orthologue) single copy and a duplicated score of 95.8% and 1.4%, respectively. Ab-initio gene model prediction and annotation with transcriptomic data constructed 33,989 gene models composing 50.4% of the assembly, including 37,036 transcripts. Repetitive elements make up approximately 39.6% of the assembly, and unresolved gap sequences are estimated to be 0.65%. Whole genome alignment with Echinometra sp. EZ revealed high synteny and conservation between the two species, further bolstering Echinometra as an emerging genus for comparative genomics studies. This genome assembly represents a high-quality genomic resource for future evolutionary and developmental studies of this species and more broadly of echinoderms.

Preliminary molecular characterization of a proinflammatory and nociceptive molecule from the Echinometra lucunter spines extracts.

BACKGROUND: Sea urchins are animals commonly found on the Brazilian shoreline, being Echinometra lucunter the most abundant species. Accidents caused by E. lucunter have been reported as one of the most frequent in Brazil, and are characterized by intense pain and inflammation, consequence of spine puncture in the skin. In order to characterize such toxic effects, we isolated one molecule that caused inflammatory and nociceptive effects. METHODS: E. lucunter specimens were collected without gender distinction. Spines were removed and molecules were extracted, fractionated by RP-HPLC and assayed for inflammatory and nociceptive activity, in a biological-driven fractionation way, until the obtainment of one active molecule and its subsequent analysis by mass spectrometry (MS and MS/MS). For inflammation, intravital microscopy was performed on the mouse cremaster muscle, in order to evaluate rolled, adherent and migrating leukocytes. Paw edema was also evaluated. For the nociceptive activity, the paw pressure test was performed in rats. RESULTS: One molecule could be isolated and related to the inflammatory and nociceptive activity. Regarding inflammation, increase in adherent and migrating cells was observed in the cremaster muscle after the administration of the molecule. Corroborating the inflammatory response, paw edema was also observed, although only in 20% of controls and 20 min after injection. Additionally, this molecule was able to decrease significantly the pain threshold, characterizing hyperalgesia. This molecule was analyzed by mass spectrometry, and according to the exact molecular mass, isotopic distribution and fragmentation profile, it was possible to propose the molecular formula C29H48N3O10. CONCLUSIONS: One isolated molecule from the spine extract of E. lucunter is able to elicit inflammation and hypernociception in animal models, which is in agreement with the effects observed in sea urchin accidents.

Ocean acidification affects parameters of immune response and extracellular pH in tropical sea urchins Lytechinus variegatus and Echinometra luccunter.

The rising concentration of atmospheric CO2 by anthropogenic activities is changing the chemistry of the oceans, resulting in a decreased pH. Several studies have shown that the decrease in pH can affect calcification rates and reproduction of marine invertebrates, but little attention has been drawn to their immune response. Thus this study evaluated in two adult tropical sea urchin species, Lytechinus variegatus and Echinometra lucunter, the effects of ocean acidification over a period of 24h and 5days, on parameters of the immune response, the extracellular acid base balance, and the ability to recover these parameters. For this reason, the phagocytic capacity (PC), the phagocytic index (PI), the capacity of cell adhesion, cell spreading, cell spreading area of phagocytic amebocytes in vitro, and the coelomic fluid pH were analyzed in animals exposed to a pH of 8.0 (control group), 7.6 and 7.3. Experimental pH's were predicted by IPCC for the future of the two species. Furthermore, a recovery test was conducted to verify whether animals have the ability to restore these physiological parameters after being re-exposed to control conditions. Both species presented a significant decrease in PC, in the pH of coelomic fluid and in the cell spreading area. Besides that, Echinometra lucunter showed a significant decrease in cell spreading and significant differences in coelomocyte proportions. The recovery test showed that the PC of both species increased, also being below the control values. Even so, they were still significantly higher than those exposed to acidified seawater, indicating that with the re-establishment of the pH value the phagocytic capacity of cells tends to restore control conditions. These results demonstrate that the immune system and the coelomic fluid pH of these animals can be affected by ocean acidification. However, the effects of a short-term exposure can be reversible if the natural values ​​are re-established. Thus, the effects of ocean acidification could lead to consequences for pathogen resistance and survival of these sea urchin species.

Echinometrin: a novel mast cell degranulating peptide from the coelomic liquid of Echinometra lucunter sea urchin.

Echinometra lucunter is an abundant sea urchin found in Brazilian waters. Accidents caused by this animal are common and are characterized by the penetration of the spines in the skin, which raises an inflammatory reaction through mechanical trauma as well as by the presumable action of toxins. Additionally, there have been reports of inflammatory reaction after the consumption of raw sea urchin eggs. In this work, we have isolated a peptide from E. lucunter coelomic fluid that could elicit inflammatory reactions, such as paw edema, leukocyte recruitment and diminishment of the pain threshold. This peptide was termed Echinometrin. Moreover, the peptide administration was able to produce in vivo degranulation of mouse mast cells, in a dose-response manner. The peptide was 'de novo' sequenced by mass spectrometry and its synthetic analog could reproduce all the observed effects. Sequence alignment indicates that this peptide is comprised in vitellogenin, an abundant nutrient protein present in the gametogenic cells of sea urchins, making it possible that echinometrin would be a cryptide with pro-inflammatory effects.

Geochemical and ecotoxicological assessment for estuarine surface sediments from Southern Brazil.

Sediments from Guaratuba Bay (PR, Brazil), a marine protected area, were collected and evaluated for geochemistry and toxicity. High levels of P and acute toxicity were observed in some samples. Concentrations of Cu, Cd, Pb and Zn were relatively low; however, Cd levels eventually exceeded Threshold Effect Level. Toxicities were associated to nutrients and metals enrichment. Results suggest that impacts are incipient and occur only at specific sites, associated to multiple contamination sources. Despite sediments quality seems to range between good and fair, attention is required to land-use planning around Guaratuba Bay and controlling local pollution sources.

The impact of rising sea temperature on innate immune parameters in the tropical subtidal sea urchin Lytechinus variegatus and the intertidal sea urchin Echinometra lucunter.

Ocean temperatures are rising throughout the world, making it necessary to evaluate the impact of these temperature changes on sea urchins, which are well-known bioindicators. This study evaluated the effect of an increase in temperature on the immune response of the subtidal Lytechinus variegatus and the intertidal Echinometra lucunter sea urchins. Both species were exposed to 20 (control), 25 and 30 °C temperatures for 24 h, 2, 7 and 14 days. Counting of coelomocytes and assays on the phagocytic response, adhesion and spreading of coelomocytes were performed. Red and colorless sphere cells were considered biomarkers for heat stress. Moreover, a significant decrease in the phagocytic indices and a decrease in both cell adhesion and cell spreading were observed at 25 and 30 °C for L. variegatus. For E. lucunter, the only alteration observed was for the cell proportions. This report shows how different species of sea urchins respond immunologically to rising temperatures.

Preliminary molecular characterization of a proinflammatory and nociceptive molecule from the Echinometra lucunter spines extracts

Abstract Background Sea urchins are animals commonly found on the Brazilian shoreline, being Echinometra lucunter the most abundant species. Accidents caused by E. lucunter have been reported as one of the most frequent in Brazil, and are characterized by intense pain and inflammation, consequence of spine puncture in the skin. In order to characterize such toxic effects, we isolated one molecule that caused inflammatory and nociceptive effects. Methods E. lucunter specimens were collected without gender distinction. Spines were removed and molecules were extracted, fractionated by RP-HPLC and assayed for inflammatory and nociceptive activity, in a biological-driven fractionation way, until the obtainment of one active molecule and its subsequent analysis by mass spectrometry (MS and MS/MS). For inflammation, intravital microscopy was performed on the mouse cremaster muscle, in order to evaluate rolled, adherent and migrating leukocytes. Paw edema was also evaluated. For the nociceptive activity, the paw pressure test was performed in rats. Results One molecule could be isolated and related to the inflammatory and nociceptive activity. Regarding inflammation, increase in adherent and migrating cells was observed in the cremaster muscle after the administration of the molecule. Corroborating the inflammatory response, paw edema was also observed, although only in 20% of controls and 20 min after injection. Additionally, this molecule was able to decrease significantly the pain threshold, characterizing hyperalgesia. This molecule was analyzed by mass spectrometry, and according to the exact molecular mass, isotopic distribution and fragmentation profile, it was possible to propose the molecular formula C29H48N3O10. Conclusions One isolated molecule from the spine extract of E. lucunter is able to elicit inflammation and hypernociception in animal models, which is in agreement with the effects observed in sea urchin accidents.

Preliminary molecular characterization of a proinflammatory and nociceptive molecule from the Echinometra lucunter spines extracts

Background Sea urchins are animals commonly found on the Brazilian shoreline, being Echinometra lucunter the most abundant species. Accidents caused by E. lucunter have been reported as one of the most frequent in Brazil, and are characterized by intense pain and inflammation, consequence of spine puncture in the skin. In order to characterize such toxic effects, we isolated one molecule that caused inflammatory and nociceptive effects. Methods E. lucunter specimens were collected without gender distinction. Spines were removed and molecules were extracted, fractionated by RP-HPLC and assayed for inflammatory and nociceptive activity, in a biological-driven fractionation way, until the obtainment of one active molecule and its subsequent analysis by mass spectrometry (MS and MS/MS). For inflammation, intravital microscopy was performed on the mouse cremaster muscle, in order to evaluate rolled, adherent and migrating leukocytes. Paw edema was also evaluated. For the nociceptive activity, the paw pressure test was performed in rats. Results One molecule could be isolated and related to the inflammatory and nociceptive activity. Regarding inflammation, increase in adherent and migrating cells was observed in the cremaster muscle after the administration of the molecule. Corroborating the inflammatory response, paw edema was also observed, although only in 20% of controls and 20 min after injection. Additionally, this molecule was able to decrease significantly the pain threshold, characterizing hyperalgesia. This molecule was analyzed by mass spectrometry, and according to the exact molecular mass, isotopic distribution and fragmentation profile, it was possible to propose the molecular formula C29H48N3O10. Conclusions One isolated molecule from the spine extract of E. lucunter is able to elicit inflammation and hypernociception in animal models, which is in agreement with the effects observed in sea urchin accidents.(AU)

Dieta del erizo negro Echinometra lucunter (Echinometra: Echinoidea) en el Nororiente de Venezuela / Diet of the black sea urchin Echinometra lucunter (Echinometra: Echinoidea) in the northeast of Venezuela

Resumen Mensualmente entre junio 2010 y agosto 2011 en la ensenada de Turpialito, Golfo de Cariaco, Venezuela, se recolectaron macroalgas e identificaron, y ejemplares de Echinometra lucunter con diámetro de testa 41.04 ± 6.60 mm para examinar el contenido estomacal y el índice de llenado. Se identificaron 65 especies de macroalgas, pertenecientes a los Phylla Rhodophyta, Heterokontophyta y Chlorophyta. La mayor biomasa anual la presentaron Halimeda opuntia (56.5 g.m-2), Acanthophora spicifera (15.94 g.m-2), Sargassum vulgare (15.28 g.m-2) y Centroceras clavulatum (12.26 g.m-2). Los ítems alimenticios más frecuentes fueron las macroalgas filamentosas: Sphacelaria sp. (546) y Herposiphonia sp. (441) y la foliácea Dictyota sp. (410), y entre los invertebrados esponjas (155), moluscos bivalvos (70) y crustáceos cirrípedos (65). El ID osciló entre 0.8 y 6.0% (2.6 ± 1.21 %), con máximos en diciembre 2010 (6.0 ± 2.21 %), abril (4.3 ± 0.34 %) y junio 2011 (4.0 ± 1.79 %), con oscilaciones mensuales asociadas a los períodos de surgencia y estratificación. El conocimiento de las características principales de la dieta, es necesaria para proponer planes de cultivo y de conservación de la especie.

Abstract Echinometra lucunter is relatively abundant in the Venezuela coast, however no details are known about its diet. We compare the stomach contents and repletion index of E. lucunter with macroalgae available in their environment (June 2010-August) and the nutrient cycle at the Golfo de Cariaco. Seventy-one species of macroalgae were identified: Rhodophyta (45), Heterokontophyta (11) and Chlorophyta (15). The mean testa diameter was 41.04 ± 6.60 mm. The food preferences included filamentous, siphonal and calcaeous macroalgae; main food items were the filamentous macroalgae Sphacelaria sp. and Herposiphonia sp., and foliose Dictyota sp., including invertebrates such as sponges, bivalves and barnacle crustaceans. The repletion index was 0.8 - 6.0 % (2.6 ± 1.21 %), maximum values were associated to primary productivity cycle, i.e. December 2010 (6.0 ± 2.21 %), April (4.3 ± 0.34 %), June 2011 (4.0 ± 1.79 %). Food preferences could be associated to the upwelling season and productivity cycle. Rev. Biol. Trop. 63 (Suppl. 2): 233-242. Epub 2015 June 01.

Cathepsin B/X is secreted by Echinometra lucunter sea urchin spines, a structure rich in granular cells and toxins

Background Echinometra lucunter is a common American sea urchin responsible for the majority of the marine accidents in Brazil. Although not lethal, these accidents are reported to be extremely painful. Recently, our group described the presence of toxins in its spines that contribute to the pathological reactions. Additionally, we have observed that the E. lucunter spines can regenerate when broken. In the present work we evaluated the enzymatic activities of sea urchin spine extracts in order to identify an enzyme that could contribute not only to the toxicity, but also participate in the spine growth and regeneration. Results The spine aqueous extract was tested for peptidase activity, with synthetic substrates, in the presence and absence of inhibitors and activators. For proper enzyme classification, the FRET-substrate cleavage pattern, pH-dependency activity and Western-blot analyses were performed. The spine extract was able to cleave Z-R-MCA and Abz-GIVRAK(Dnp)-OH following pre-incubation with DTT, and was inhibited by E-64. Furthermore, the double-peaked pH curve (5 and 7) and the cleavage site proportion (4:6, R,A:A,K) indicate the presence of both mono and dicarboxypeptidase activities. Moreover, in Western-blot analysis, the spine extract was positive for anti-cathepsin B antibody. Conclusions E. lucunter spines extracts presented a cysteine peptidase activity that was identified as cathepsin B/X that would participate in the remodeling and growth processes of the spine, as well as in the inflammatory response to the accident.

Cathepsin B/X is secreted by Echinometra lucunter sea urchin spines, a structure rich in granular cells and toxins

Background Echinometra lucunter is a common American sea urchin responsible for the majority of the marine accidents in Brazil. Although not lethal, these accidents are reported to be extremely painful. Recently, our group described the presence of toxins in its spines that contribute to the pathological reactions. Additionally, we have observed that the E. lucunter spines can regenerate when broken. In the present work we evaluated the enzymatic activities of sea urchin spine extracts in order to identify an enzyme that could contribute not only to the toxicity, but also participate in the spine growth and regeneration. Results The spine aqueous extract was tested for peptidase activity, with synthetic substrates, in the presence and absence of inhibitors and activators. For proper enzyme classification, the FRET-substrate cleavage pattern, pH-dependency activity and Western-blot analyses were performed. The spine extract was able to cleave Z-R-MCA and Abz-GIVRAK(Dnp)-OH following pre-incubation with DTT, and was inhibited by E-64. Furthermore, the double-peaked pH curve (5 and 7) and the cleavage site proportion (4:6, R-A:A-K) indicate the presence of both mono and dicarboxypeptidase activities. Moreover, in Western-blot analysis, the spine extract was positive for anti-cathepsin B antibody. Conclusions E. lucunter spines extracts presented a cysteine peptidase activity that was identified as cathepsin B/X that would participate in the remodeling and growth processes of the spine, as well as in the inflammatory response to the accident.(AU)

Observation of initial clinical manifestations and repercussions from the treatment of 314 human injuries caused by black sea urchins (Echinometra lucunter) on the southeastern Brazilian coast / Observação das manifestações clínicas iniciais e sua influência no tratamento de 314 acidentes causados por ouriços-do-mar pretos (Echinometra lucunter) na região sudeste do Brasil

INTRODUCTION: Injuries caused by sea urchins are the most common caused by marine animals in humans in Brazil, with the black sea urchin (Echinometra lucunter) causing the most injuries to bathers. METHODS: This study observed 314 human wounds with emphasis on the early observation of clinical signs and symptoms and their implications on the recommended treatment. RESULTS: All the injuries were caused by black sea urchins and were observed in bathers. The lesions and the pain were associated with penetration of the spines; there was no early inflammation or pain without pressure on the wounded places. Complications arising from this kind on injury, including infections and foreign body granulomas, are associated with the permanence of the spines in the wounds. CONCLUSIONS: The study confirmed that this kind of injury is the most common accident caused by aquatic animals in Brazil. The main therapeutical recommendation is early removal of the spines to prevent late complications, such as infections and the formation of foreign body granulomas.

INTRODUÇÃO: Os acidentes causados por ouriços-do-mar são as ocorrências por animais marinhos mais comuns no país. O ouriço-do-mar preto (Echinometra lucunter) é a espécie que mais causa ferimentos em banhistas. MÉTODOS: Este trabalho registrou e estudou 314 agravos com ênfase nas manifestações clínicas iniciais observadas e suas implicações na terapêutica recomendada. RESULTADOS: Todos os acidentes foram causados pelo ouriço-do-mar preto e aconteceram em banhistas. As lesões e a dor foram associadas ao trauma causado pela penetração das espículas (não ocorreu inflamação ou dor imediata sem pressão sobre os pontos comprometidos). As complicações deste tipo de acidente, incluindo infecções e granulomas de corpo estranho, estão associadas com a permanência das espículas nos ferimentos. CONCLUSÕES: Foi confirmado o fato do acidente causado por esta espécie ser o mais comum no Brasil e apresentar caráter traumático, sendo a principal recomendação a retirada precoce dos espinhos para prevenção de complicações tardias como as infecções e formação de granulomas de corpo estranho.

Estudos toxinológicos do ouriço-do-mar Echinometra lucunter / Toxinologic studies about Echinometra lucunter sea urchin

Echinometra lucunter (Linnaeus, 1758) e o ourico do mar mais conhecido da costa brasileira, responsavel por cerca de 50% dos acidentes com animais marinhos. A injuria inicial causada pelos ouricos e a penetracao dos espinhos na pele, seguida da retencao de seus fragmentos. Esses fragmentos causam reacoes inflamatorias, dor local, e ocasionalmente doenca sistemica,sintomas que eram atribuidos somente ao trauma mecanico. Acidentes apos a ingestao de ovas tambem ja foram descritos. O objetivo deste trabalho foi verificar a presenca de toxinas nos espinhos e no liquido celomico perivisceral do ourico do mar E. lucunter do litoral de São Paulo, isolar e caracterizar essas moleculas e avaliar a correlacao histologica entre as toxinas presentes e uma possivel estrutura sectora... .

Echinometra lucunter (Linnaeus, 1758) is the most spread the sea urchin of the Brazilian shore line and its responsible for circa 50% of all marine animals acidents. Initial sea urchin injury is caused by the spine penetration, followed by its fragmentation under the skin. This fragments can cause local pain and inflammatory reactions, initially atributed to the mechanical trauma of the spines penetration, and ocasionally systemic disorders. Few accidents were reported after the ingestion of raw sea urchin. The aim of this work was to asses the presence of toxins in the spines and perivisceral celomic fluid of E. lucunter sea urchun from São Paulo shore line, through the biological driven isolation and biochemical characterization of the toxins... .