Biological activity in extracts of ascidians (Tunicata, Ascidiacea) from the northeastern Brazilian coast

Ascidians are marine animals with a great ability to synthesize bioactive substances. This study examined the cytotoxic potential of 10 ascidians found in the coastal waters of Northeast Brazil. Samples of the species Eudistoma vannamei Millar, 1977, Eudistoma sp., Didemnum ligulum Monniot F., 1983, Didemnum psammatodes (Sluiter, 1895), Didemnum sp., Polysyncraton sp., Trididemnum sp., Cystodytes dellechiajei (Della Valle, 1877), Euherdmania sp., and an unidentified species belonging to the Holozoidae family were extracted in methanol 5:1 (v/w). The extracts were tested for cytotoxicity using the brine shrimp lethality assay, sea urchin egg development assay, hemolysis assay, and MTT assay using tumor cell lines. The extract of E. vannamei showed the highest toxicity in brine shrimp (LD₅₀=34.7 μg/ml) and in all tumor cell lines tested, with an IC₅₀ of <2 μg/ml for CEM, 11.2 μg/ml for HL-60, 23.8 μg/ml for B16, and 14.3 μg/ml for HCT-8. In sea urchin eggs, it inhibited the cell cycle progression mainly at the blastula stage (IC₅₀=74.8 μg/ml). The extract of Euherdmania sp. also exhibited some toxicity in these assays, but at a lower potency than that of E. vannamei. The extracts of D. psammatodes and Polysyncraton sp. showed a strong inhibition of the sea urchin egg cell cycle during both phases examined, first cleavage and blastula, with a possible action on the cell microfilament apparatus. The extract of D. ligulum showed selective toxicity toward HCT-8 cells (IC₅₀=35.3 μg/ml). The extract from the Holozoidae was the only one that possessed a hemolytic effect, with an IC₅₀ of 175.2 μg/ml. Further studies are necessary for a better characterization of the active principles of these extracts and a possible elucidation of the mechanisms of action.     

Zooplankton biomass distribution patterns along the western Antarctic Peninsula (December 2002)

The phytoplankton [chlorophyll a (Chl a)], microzooplankton,mesozooplankton and macrozooplankton biomass and distributionwere studied as part of a multidisciplinary project (Tempano)along the Antarctic Peninsula during December 2002. Even thoughthe summer phytoplankton bloom was not yet developed in thearea, autotrophs dominated the plankton biomass. Phytoplanktonvertical distribution was, in general, homogeneous in the upper40–50 m of the water column, further decreasing with depth.Protozoans showed low biomass; their contribution to the totalplankton being one order of magnitude lower than that of autotrophs.The vertical distribution of protozoans was variable among stationswith marked peaks at depths ranging from 30 to 80 m. Mesozooplankton-integratedbiomass was generally low, although there was a notable increasesouthward near the ice marginal zone. Macrozooplankton distributionwas more variable without any clear zonal distribution pattern.The vertical distribution of meso- and macrozooplankton (>4mm) biomass showed clear peaks of abundance comprising differentspecies depending on the geographical area. Our biomass distributiondata suggest a food-web scenario in which macrozooplankton arepreying on mesozooplankton populations only in the northernerstations, and mesozooplankton are, in their turn, shaping theabundance of the emerging populations of microzooplankton. Phytoplankton,on the other hand, seem to be hardly controlled by grazing activity.     

Cupular sense organs in Ciona (Tunicata: Ascidiacea)

The cupular organs of the atrial (exhalent) siphon of Ciona have been studied with scanning and transmission microscopy and are shown to resemble those of the vertebrate acoustico-lateralis system in several respects. The sensory cells are ciliated, and their cilia are apparently non-motile, having a modified inner tubular array. These cells lie amongst supporting cells that probably secrete the cupula, which is composed of polysaccharide and proteins as is the test. Ciona is sensitive to near-field vibrations, even after the brain has been removed; the significance of this observation and of the arrangement of the cupular organs is discussed. It is concluded that the tunicates show a suitable morphological starting point for the vertebrate acoustico-lateralis system.     

Antileukemic effects of Didemnum psammatodes (Tunicata: Ascidiacea) constituents

Chemical investigation of the methanolic extract of the ascidian Didemnum psammatodes has led to the identification of fourteen known compounds: three methyl esters (methyl myristate, methyl palmitate and methyl stearate), four steroids (cholesterol, campesterol, stigmasterol and beta-sitosterol), two fatty acids (palmitic acid and stearic acid), three glyceryl ethers {(1,2-propanediol, 3-(heptadecyloxy), batyl alcohol and 1,2-propanediol, 3-[(methyloctadecyl)oxy]} and two nucleosides (thymidine and 2'-deoxyguanosine). Their structures were proposed by NMR and comparison with literature data and GC analysis in comparison with authentic sample. The cytotoxic activity of these compounds was evaluated against human leukemia cell line panel using the MTT assay. The mixture of the three methyl esters was the most active group of compounds, showing antiproliferative and cytotoxic effects. Further studies on their mode of action suggest that these activities are connected with inhibition of DNA synthesis and induction of both necrosis and apoptosis.     

Metabolism and elemental composition of aggregate and solitary forms of Salpa thompsoni (Tunicata: Thaliacea) in waters off the Antarctic Peninsula during austral summer 1999

Oxygen consumption, ammonia excretion and phosphate excretionrates were measured on the aggregate form (aggregated sexualblastozooid generation) and solitary form (solitary asexualoozooid generation) of Salpa thompsoni sampled from waters offthe Antarctic Peninsula from December 1999 to February 2000,in conjunction with body composition analysis (water, ash, carbon,nitrogen and phosphorus). The specific metabolic rates of S.thompsoni were weight-independent. No significant differencesbetween the aggregate and solitary forms were observed for theweight-specific rates with the exception of the oxygen consumptionrate on phosphorus weight. Metabolic loss estimated for theaggregate and solitary forms was 3.0 and 3.5% day^–1 forbody carbon, 1.6 and 1.8% for body nitrogen, 3.9 and 10.6% forbody phosphorus, respectively. Contents of carbon (6.0% of dryweight for the aggregate form and 5.1% for the solitary form),nitrogen (1.5 and 1.3%) and phosphorus (0.15 and 0.11%) tendedto decrease with the increase of dry weight. All mean valuesof each body constituent (water, ash, C, N and P) for the aggregateand solitary forms were not significantly different in the samedry weight ranges.     

Seabird ticks (Ixodes uriae) distribution along the Antarctic Peninsula

The distribution of the tick Ixodes uriae is studied in the South Shetlands and different locations along the Antarctic Peninsula. Ticks were found beneath stones close to penguin rookeries of chinstrap, gentoo and adelie penguin, although no individuals were found parasitized. Our results showed that ticks are not distributed evenly along the Antarctic Peninsula being more common and abundant in the northern part with relative abundances of ticks ranging from 1 to 57 individuals per stone and from 2 to 26% of the stone inspected. Ticks are probably absent in the south.     

Functional morphology of the branchial basket of ascidia paratropa (Tunicata,Ascidiacea)

Summary The filtering surface of the branchial basket of Ascidia paratropa (Huntsman, 1912) consists of two adjacent and connected sheets. The stigmatal surface is loosely folded. The secondary gill screen, composed of ciliated longitudinal vessels with primary and secondary papillae, is connected to the stigmatal surface on the inside of the branchial basket. Continuous mucus secretion results in a net with elastic and adhesive properties, and a pore size of less than 0.5 m in its longest dimension. Net production cannot be interrupted by external mechanical stimuli. The net is usually supported by and transported across the secondary screen, forming a filtering surface that parallels the undulations of the stigmatal surface; however, localized muscular activity of the dorsal laminar fold and longitudinal vessels are also involved in mucous net transport. Squirts occur more frequently than in many species. At these times the net is transported as a flat sheet, independent of the secondary screen. Squiring does not interrupt continuous feeding activity.     

Morphological aspects of fertilization in Phallusia (Ascidia) nigra (Ascidiacea,Tunicata)

The spermatozoa of Phallusia (Ascidia) nigra have an elongated head (approximately 5 m in length) in which a nucleus and a single mitochondrion are located side by side. There is no midpiece. The apex of the head is wedge-shaped. Acrosomal vesicles (approximately 55–65 nm in diameter) and moderately electron-dense material (MEDM) are present between the plasmalemma and the nuclear membranes in the anterior tip of the head. The MEDM occupies a central position and three or four acrosomal vesicles are seen in a line alongside it. The acrosomal vesicles disappear as the sperm makes contact with the surface of the chorion. Gamete fusion most likely occurs between a small process extending from the peripheral margin of the sperm apex and the egg surface, resulting in incorporation of the sperm into the egg from the anterior region of its head.     

Ovulation in Diplosoma (Tunicata, Ascidiacea, Didemnidae): a light microscopical study

Serial sections of non-ovulated and ovulated eggs of Diplosoma migrans (Tunicata, Ascidiacea, Didemnidae) were compared in order to study the process of ovulation on the level of light microscopy. Fully grown eggs are surrounded by an outer follicular epithelium consisting of cubic cells, and an inner follicular epithelium consisting of flat cells. Egg release is initiated by a fusion of the epidermis with the outer follicular epithelium at the distal pole of the egg. A contraction of the outer follicular epithelium leads to an opening of both epidermis and outer follicular epithelium, and their removal from the egg. The envelope of ovulated eggs corresponds to the former inner follicular epithelium. Mainly due to a contraction of the epidermis, the outer follicular epithelium is shifted into the abdomen and separates from the epidermis. It transforms into a vestigial body (corpus luteum) and finally atrophies. The epidermis flattens and resumes its typical appearance. On the basis of these observations, the results of studies on the closely related Diplosoma listerianum are discussed and questioned. Electronic Publication     

Glycogen and degeneration in the pyloric gland of Dendrodoa grossularia (Ascidiacea,Tunicata)

Summary Degeneration is observed in cells of the pyloric gland of Dendrodoa grossularia in which glycogen storage occurs. The ultrastructure of four phases of the degeneration cycle is described. Natural senescence seems to be the cause of the degeneration. Glycogen storage might be the result of metabolic disturbance, but its presence reveals the importance of glycogen in the function of the organ. The role of the pyloric gland is discussed.     

The alien ascidian Styela clava now invading the Sea of Marmara (Tunicata: Ascidiacea)

During the implementation of a large project aimed to investigate the benthic community structures of the Sea of Marmara, specimens of the invasive ascidian species Styela clava were collected on natural substrata (rocks) at 10 m depth at one locality (Karamürsel) in İzmit Bay. The specimens were mature, containing gametes, indicating that the species had become established in the area. The Sea of Marmara seems to provide suitable conditions for this species to survive and form proliferating populations.     

Foraminiferal distribution on the continental margin off Nova Scotia

Q-mode factor analysis of total foraminiferal abundance data (living plus dead) from 250 grab samples taken from the continental margin off Nova Scotia allows the determination of eight factor assemblages. On the northeastern shelf, an exclusively agglutinated assemblage dominated byAdercotryma glomerata occupies both banks and basins. Central shelf basins contain a predominantly calcareous assemblage dominated byGlobobulimina auriculata andNonionellina labradorica. Transitional between these two factor assemblages is an agglutinated assemblage dominated bySaccammina atlantica. Consistently present along the shelf edge is aTrifarina angulosa assemblage. In northeastern bays and a few samples near Sable Island, an agglutinatedEggerella advena assemblage is found. A relict and transport affected assemblage dominated byElphidium excavatum occurs in the southwestern approaches to Emerald Basin. Sandy/gravelly areas of the inner shelf and outer bank regions are characterized by aCibicides lobatulus assemblage.The statistical relationships of these defined assemblages to various aspects of the marine environment (depth, temperature, salinity, percent gravel, sand and mud) were investigated through multiple regression techniques. Results indicate that the present foraminiferal distribution patterns off Nova Scotia are influenced by the prevailing watermass characters and substrate. TheAdercotryma glomerata assemblage is influenced by the presence of cold, (0–4°C) low salinity waters (32–33‰) of arctic, Labrador Current origin. The central basin assemblage (G. auriculata) is related to warmer (8–12°C) more saline waters (35‰) of slope origin. The transition between these two bottom waters is marked by the opportunisticSaccammina atlantica assemblage. Preferred substrate character possibly determines the occurrence of theCibicides lobatulus, Islandiella islandica andEggerella advena assemblages.Trifarina angulosa shows a significant relationship to salinity and depth.Although the surficial sediments on the Nova Scotian Shelf are largely the product of reworking of glacial deposits during late glacial and early Holocene times, all but theE. excavatum factor assemblage appear to be in equilibrium with the modern oceanographic regime.     

陕北烟田多异瓢虫空间分布型与抽样技术研究

根据1995年烟田多异瓢虫Hippodamiavariegats（Goeze）调查,计算了6种分布型指数,以及频次拟合优度检验,结果表明：该虫幼虫、蛹在陕北烟田里聚集分布,其聚集强度随种群密度增加而降低,聚集原因是环境差异;卵块分布同时具有聚集和随机的趋势,成虫为随机分布。同时建立了各虫态的理论抽样量公式,估测种群密度公式。     

Cytotoxic activity of a dichloromethane extract and fractions obtained from Eudistoma vannamei (Tunicata: Ascidiacea)

This study consists of the bioassay-guided fractionation of the dichloromethane extract from Eudistoma vannamei and the pharmacological characterization of the active fractions. The dried hydromethanolic extract dissolved in aqueous methanol was partitioned with dichloromethane and chromatographed on a silica gel flash column. The anti-proliferative effect was monitored by the MTT assay. Four of the latest fractions, numbered 14 to 17, which held many chemical similarities amongst each other, were found to be the most active. The selected fractions were tested for viability, proliferation and death induction on cultures of HL-60 promyeloblastic leukemia cells. The results suggested that the observed cytotoxicity is related to apoptosis induction.     

Cytochemical investigations on tunic morphogenesis in the sea peach Halocynthia papillosa (Tunicata, Ascidiacea) 2: demonstration of proteins

In a previous paper, cellulose fibres were demonstrated in the larval, the metamorphosing, and the juvenile tunics. In this paper we used cytochemical methods and X-ray microanalysis to obtain additional information on tunic morphogenesis in Halocynthia papillosa. The chemical composition of the tunic evolves with its structural complexity. The larval and juvenile fibres are shown to be structurally and chemically different. While neither proteins nor glycosaminoglycans seem to be associated with the larval fibres, the juvenile fibres consist of a cellulose core wrapped in a sheath of tannophilic proteins. Patches of glycosaminoglycans line their longitudinal axes. In the course of metamorphosis, the cuticle undergoes profound modifications in regions of spine morphogenesis. Granular material that was previously called fibro-granular material (Lübbering et al., 1993) is essential to the formation of cuticular plates and spines. During metamorphosis, this material accumulates in epidermal granules and is discharged into the tunic. It crosses the fundamental layer of the tunic and reaches the cuticle. Our results strongly suggest that this material consists of proteins rich in cysteine and hydrophobic amino acids.     

An experimental test of colonization ability in the potentially invasive Didemnum perlucidum (Tunicata, Ascidiacea)

Exotic species invasions are one of the greatest threats to marine systems and ascidians have many characteristics that favor transport, colonization and establishment into new regions. Didemnum perlucidum is a widespread species that has been introduced into tropical ports around the world. Here we examine the colonizing ability of D. perlucidum by experimental use of artificial plates in a shellfish culture. The experiment comprised paired plates for colonization (bare and occupied) in 16 monthly replicates. Recruitment and space occupation were compared between bare and occupied plates and an estimation of reproductive effort was based on the number of larvae produced in each of ten colonies collected on the culture structures. D. perlucidum reproduced continuously but greatest reproduction occurred between December 2006 and May 2007. While recruitment was somewhat greater (number of new colonies) on bare plates, this species can colonize already occupied substrates and, surprisingly, colony area was always similar between treatments. Thus, while fewer colonies formed on occupied plates, once formed, colonies grew at rates similar to those on clean plates. Thus, D. perlucidum colonizes substrates very efficiently, especially when unoccupied space is available.     

Environmental determinants of top predator distribution within the dynamic winter pack ice zone of the northern Antarctic Peninsula

Global warming is predicted to reduce the amount of sea ice concentration in polar environments, thus presenting profound changes for populations of seabirds and marine mammals dependent on sea ice. Using data from a shipboard survey during August 2012, I test the hypothesis that relative abundance of seabird and marine mammals reflects environmental variability associated with the dynamic pack ice zone. Using environmental data and observations of sea ice concentration, I quantified an environmental gradient that describes the spatial organization of the dynamic pack ice zone. The relationship of top predators to this environmental gradient revealed three important aspects: (1) an open water and pack ice community is present with some top predator species exhibiting higher abundance associated with moderate sea ice concentration (40–60 %) as opposed to the pack ice edge (10 %), (2) Antarctic fur seals (Arctocephalus gazella) were the most abundant pinniped and they were observed resting on ice floes and foraging within leads and polynyas, and (3) for the most abundant species, spatial regression models indicate that latitude and sea ice concentration (a principal north/south gradient) are the most important environmental determinants. Winter ocean conditions may strongly influence population dynamics of top predators; therefore, information regarding their habitat use during winter is needed for understanding ecosystem dynamics.