Environmental Flow Regimes for Dysidea avara Sponges

The aim of our research is to design tank systems to culture Dysidea avara for the production of avarol. Flow information was needed to design culture tanks suitable for effective production. Water flow regimes were characterized over a 1-year period for a shallow rocky sublittoral environment in the Northwestern Mediterranean where D. avara sponges are particularly abundant. Three-dimensional Doppler current velocities at 8¿10-m depths ranged from 5 to 15 cm/s over most seasons, occasionally spiking to 30¿66 cm/s. A thermistor flow sensor was used to map flow fields in close proximity (¿2 cm) to individual sponges at 4.5-, 8.8-, and 14.3-m depths. These ¿proximal flows¿ averaged 1.6 cm/s in calm seas and 5.9 cm/s during a storm, when the highest proximal flow (32.9 cm/s) was recorded next to a sponge at the shallowest station. Proximal flows diminished exponentially with depth, averaging 2.6 cm/s¿±¿0.15 SE over the entire study. Flow visualization studies showed that oscillatory flow (0.20¿0.33 Hz) was the most common regime around individual sponges. Sponges at the 4.5-m site maintained a compact morphology with large oscula year-around despite only seasonally high flows. Sponges at 8.8 m were more erect with large oscula on tall protuberances. At the lowest-flow 14.3-m site, sponges were more branched and heavily conulated, with small oscula. The relationship between sponge morphology and ambient flow regime is discussed

Diversity, structure and spatial distribution of megabenthic communities in Cap de Creus continental shelf and submarine canyon (NW Mediterranean)

The continental shelf and submarine canyon off Cap de Creus (NW Mediterranean) were declared a Site of Community Importance (SCI) within the Natura 2000 Network in 2014. Implementing an effective management plan to preserve its biological diversity and monitor its evolution through time requires a detailed character ization of its benthic ecosystem. Based on 60 underwater video transects performed between 2007 and 2013 (before the declaration of the SCI), we thoroughly describe the composition and structure of the main mega benthic communities dwelling from the shelf down to 400 m depth inside the submarine canyon. We then mapped the spatial distribution of the benthic communities using the Random Forest algorithm, which incor porated geomorphological and oceanographic layers as predictors, as well as the intensity of the bottom-trawling fishing fleet. Although the study area has historically been exposed to commercial fishing practices, it still holds a rich benthic ecosystem with over 165 different invertebrate (morpho)species of the megafauna identified in the video footage, which form up to 9 distinct megabenthic communities. The continental shelf is home to coral gardens of the sea fan Eunicella cavolini, sea pen and soft coral assemblages, dense beds of the crinoid Leptometra phalangium, diverse sponge grounds and massive aggregations of the brittle star Ophiothrix fragilis. The submarine canyon off Cap de Creus is characterized by a cold-water coral community dominated by the scleractinian coral Madrepora oculata, found in association with several invertebrate species including oysters, brachiopods and a variety of sponge species, as well as by a community dominated by cerianthids and sea urchins, mostly in sedimentary areas. The benthic communities identified in the area were then compared with habitats/biocenoses described in reference habitat classification systems that consider circalittoral and bathyal environments of the Mediterranean. The complex environmental setting characteristic of the marine area off Cap de Creus likely produces the optimal conditions for communities dominated by suspension- and filter-feeding species to develop. The uniqueness of this ecosystem and the anthropogenic pressures that it faces should prompt the development of effective management actions to ensure the long-term conservation of the benthic fauna representative of this marine area3,26

Grazing, differential size-class dynamics and survival of the Mediterranean sponge Corticium candelabrum

The growth dynamics and survival of the sponge Corticium candelabrum (Demospongiae: Homosclerophorida) were surveyed in the northwestern Mediterranean for more than 3 yr. Growth and regeneration rates, fission and fusion events and survival were monitored monthly. Moreover, in situ punctual clearance experiments were conducted seasonally searching for possible relationships between food uptake and sponge dynamics. The monthly growth rates (GR) of C. candelabrum were low (0.19 ± 0.02 mean [±SE] for the 3 yr of study), variable and seasonal, with the highest values in summer. The cumulative survival function followed a stepped profile with several consecutive months without mortality separated by shorter mortality events, which mainly occurred in cold months (winter¿spring). However, an event of high mortality (76% of the monitored individuals died) took place in the particularly warm summer 2003. Fission events were frequent after previous damage (e.g. partial predation) and only one fusion event was recorded along the study period. The diet of C. candelabrum was highly heterogeneous. Differences in clearance rates (CR) among picoplankton types with season indicated that the sponge retained with different efficiency the several picoplankton types present in the water. Survival and GRs were significantly different for small, medium and large individuals (size-classes I, II and III), with the small sponges showing the lowest survival (56.6% cumulative mortality for the last 2 yr of study) and the highest GRs (0.18 ± 0.03 mo¿1, mean ± SE). On the whole, the results indicate that C. candelabrum is a slow-growing but dynamic sponge

Cell culture from sponges: pluripotency and immortality

Sponges are a source of compounds with potential pharmaceutical applications. In this article, methods of sponge cell culture for production of these bioactive compounds are reviewed, and new approaches for overcoming the problem of metabolite supply are examined. The use of embryos is proposed as a new source of sponge material for cell culture. Stem cells are present in high amounts in embryos and are more versatile and resistant to infections than adult cells. Additionally, genetic engineering and cellular research on apoptotic mechanisms are promising new fields that might help to improve cell survival in sponge-cell lines. We propose that one topic for future research should be how to reduce apoptosis, which appears to be very high in sponge cell cultures

Long-term culture of sponge explants: conditions enhancing survival and growth, and assessment of bioactivity

Sponges are an important source of secondary metabolites with pharmaceutical interest. This is the main reason for the increasing interest of sponge culture recent years. The optimal culture system depends on the species to be cultured: while some species easily produce sponge aggregates after dissociation (primmorphs), others show a great capacity to regenerate after fragmentation (explants). Corticium candelabrum is a Mediterranean bacteriosponge that can undergo asexual reproduction. We have taken advantage of this capability and cultured C. candelabrum explants under several experimental conditions. To find the best conditions for obtaining functional explants, we assayed a range of conditions, including seasons of collection, culture temperature, filtered versus filtered-sterile seawater, addition of antibiotics and proportion of ectosome. We monitored the changes in shape and ultrastructure during the formation of explants. After 24 h, TEM images showed the aquiferous system disarranged, in particular at the sponge periphery. From 2 to 4 weeks later, the aquiferous system regenerated, and fragments became functional sponges (explants). Explants were cultured under two regimes: in vitro and in a closed aquarium system. Antibiotics were only added to the in vitro culture to assess their effect on the symbiotic bacteria, which remained healthy despite the presence of antibiotics. Two food regimens (marine bacteria and green algae) were assayed for their ability to satisfy the metabolic requirements of explants. We monitored explant survival and growth. Explants showed a high long-term survival rate (close to 100%). Growth rates were higher in the closed aquarium system, without antibiotic addition, and fed with algae. Explants cultures were hardly contaminated because manipulation was reduced to a minimum and we used sterilized seawater. C. candelabrum produces bioactive molecules, which may play a defensive role in the sponge and may have pharmaceutical interest. The bioactivity of the explants was similar to that of wild sponges

Vertical transmission and successive location of symbiotic bacteria during embryo development and larva formation in Corticium candelabrum (Porifera: Demospongiae)

This study reports on the transfer of heterotrophic bacteria from parental tissue to oocytes in the Mediterranean bacteriosponge Corticium candelabrum (Homosclerophorida) and the description of the successive locations of the microsymbionts during embryo development through transmission and scanning electron microscopy. Eight different types of symbiotic bacteria are described morphologically. These eight bacteria morphotypes are found in both adult individuals and larvae. Symbiotic bacteria are transferred to oocytes, but not to spermatocytes. Bacteria are first located at the oocyte periphery below a thick collagen layer and then they migrate to the oocyte cytoplasm, forming spherical clusters. After cleavage, the bacteria can be found in the free space between blastomeres but mainly accumulate at the embryo periphery below the follicular cells that surround the embryo. Once the blastocoel is formed, the symbiotic bacteria move to this central cavity where they actively divide by bipartition, increasing their number considerably. Many examples of phagocytosed bacteria in the proximal zone of the larval cells are observed at this stage. Consequently, bacteria may represent a complementary source of energy for free larvae and settlers before they are able to capture food from the surrounding water

Structural and Stereochemical Studies of C-21 Terpenoids from the Mediterranean Spongiidae Sponges

The degraded C-21 sesterterpenoid (+)-3, enantiomeric with (-)-untenospongin B, has been isolated from the Mediterranean sponge Spongia virgultosa. The absolute stereochemistry of 3 was assigned by applying Mosher’s method. On the basis of this work, the absolute stereochemistry at C-11 of nitenin (1) and dihydronitenin (2) has been reanalyzed by applying Mosher’s method, whereas the R chirality at C-8 of 2 was determined by recording NOE spectra.The structures of two known C-21 furanoterpenes, tetradehydrofurospongin-1 (8) and 7, have been revised as (+)-3 (named “revised tetradehydrofurospongin-1”) and its acetyl derivative 5, respectively. Finally, a comparison between the Mosher and Horeau methods was carried out, paying attention to the reliability of the stereochemical predictions obtained by the two approaches applied to compounds (+)-3, nitenin (1), and dihydronitenin (2), containing aliphatic-type alcohols

Embryo development of Corticium candelabrum (Demospongiae: Homosclerophorida)

Corticium candelabrum is a homosclerophorid sponge widespread along the rocky Mediterranean sublittoral. Scanning and transmission electron microscopy were used to describe the gametes and larval development. The species is hermaphroditic. Oocytes and spermatocytes are clearly differentiated in April. Embryos develop from June to July when the larvae are released spontaneously. Spermatic cysts originate from choanocyte chambers and spermatogonia from choanocytes by choanocyte mitosis. Oocytes have a nucleolate nucleus and a cytoplasm filled with yolk granules and some lipids. Embryos are surrounded by firmly interlaced follicular cells from the parental tissue. A thin collagen layer lies below the follicular cells. The blastocoel is formed by migration of blastomeres to the morula periphery. Collagen is spread through the whole blastocoel in the embryo, but is organized in a dense layer (basal lamina) separating cells from the blastocoel in the larva. The larva is a typical cinctoblastula. The pseudostratified larval epithelium is formed by ciliated cells. The basal zone of the ciliated cells contains lipid inclusions and some yolk granules; the intermediate zone is occupied by the nucleus; and the apical zone contains abundant electron-lucent vesicles and gives rise to cilia with a single cross-striated rootlet. Numerous paracrystalline structures are contained in vacuoles within both apical and basal zones of the ciliated cells. Several slightly differentiated cell types are present in different parts of the larva. Most cells are ciliated, and show ultrastructural particularities depending on their location in the larvae (antero-lateral, intermediate, and posterior regions). A few smaller cells are non-ciliated. Several features of the C. candelabrum larva seem to support the previously proposed paraphyletic position of homoscleromorphs with respect to the other demosponges

Embryo development of Corticium candelabrum (Demospongiae: Homosclerophorida)

Corticium candelabrum is a homosclerophorid sponge widespread along the rocky Mediterranean sublittoral. Scanning and transmission electron microscopy were used to describe the gametes and larval development. The species is hermaphroditic. Oocytes and spermatocytes are clearly differentiated in April. Embryos develop from June to July when the larvae are released spontaneously. Spermatic cysts originate from choanocyte chambers and spermatogonia from choanocytes by choanocyte mitosis. Oocytes have a nucleolate nucleus and a cytoplasm filled with yolk granules and some lipids. Embryos are surrounded by firmly interlaced follicular cells from the parental tissue. A thin collagen layer lies below the follicular cells. The blastocoel is formed by migration of blastomeres to the morula periphery. Collagen is spread through the whole blastocoel in the embryo, but is organized in a dense layer (basal lamina) separating cells from the blastocoel in the larva. The larva is a typical cinctoblastula. The pseudostratified larval epithelium is formed by ciliated cells. The basal zone of the ciliated cells contains lipid inclusions and some yolk granules; the intermediate zone is occupied by the nucleus; and the apical zone contains abundant electron-lucent vesicles and gives rise to cilia with a single cross-striated rootlet. Numerous paracrystalline structures are contained in vacuoles within both apical and basal zones of the ciliated cells. Several slightly differentiated cell types are present in different parts of the larva. Most cells are ciliated, and show ultrastructural particularities depending on their location in the larvae (antero-lateral, intermediate, and posterior regions). A few smaller cells are non-ciliated. Several features of the C. candelabrum larva seem to support the previously proposed paraphyletic position of homoscleromorphs with respect to the other demosponges