Electrochemical Approach for Isolation of Chitin from the Skeleton of the Black Coral Cirrhipathes sp. (Antipatharia).

The development of novel and effective methods for the isolation of chitin, which remains one of the fundamental aminopolysaccharides within skeletal structures of diverse marine invertebrates, is still relevant. In contrast to numerous studies on chitin extraction from crustaceans, mollusks and sponges, there are only a few reports concerning its isolation from corals, and especially black corals (Antipatharia). In this work, we report the stepwise isolation and identification of chitin from Cirrhipathes sp. (Antipatharia, Antipathidae) for the first time. The proposed method, aiming at the extraction of the chitinous scaffold from the skeleton of black coral species, combined a well-known chemical treatment with in situ electrolysis, using a concentrated Na2SO4 aqueous solution as the electrolyte. This novel method allows the isolation of α-chitin in the form of a microporous membrane-like material. Moreover, the extracted chitinous scaffold, with a well-preserved, unique pore distribution, has been extracted in an astoundingly short time (12 h) compared to the earlier reported attempts at chitin isolation from Antipatharia corals.

Insights into the evolution of metazoan regenerative mechanisms: roles of TGF superfamily members in tissue regeneration of the marine sponge Chondrosia reniformis.

Tissue repair is an adaptive and widespread metazoan response. It is characterised by different cellular mechanisms and complex signalling networks that involve numerous growth factors and cytokines. In higher animals, transforming growth factor-ß (TGF-ß) signalling plays a fundamental role in wound healing. In order to evaluate the involvement of TGF superfamily members in lower invertebrate tissue regeneration, sequences for putative TGF ligands and receptors were isolated from the transcriptome of the marine sponge Chondrosia reniformis We identified seven transcripts that coded for TGF superfamily ligands and three for TGF superfamily receptors. Phylogenetically, C. reniformis TGF ligands were not grouped into any TGF superfamily clades and thus presumably evolved independently, whereas the TGF receptors clustered in the Type I receptor group. We performed gene expression profiling of these transcripts in sponge regenerating tissue explants. Data showed that three ligands (TGF1, TGF3 and TGF6) were mainly expressed during early regeneration and seemed to be involved in stem cell maintenance, whereas two others (TGF4 and TGF5) were strongly upregulated during late regeneration and thus were considered pro-differentiating factors. The presence of a strong TGF inhibitor, SB431542, blocked the restoration of the exopinacoderm layer in the sponge explants, confirming the functional involvement of the TGF pathway in tissue regeneration in these early evolved animals.

Mediterranean Bioconstructions Along the Italian Coast.

Marine bioconstructions are biodiversity-rich, three-dimensional biogenic structures, regulating key ecological functions of benthic ecosystems worldwide. Tropical coral reefs are outstanding for their beauty, diversity and complexity, but analogous types of bioconstructions are also present in temperate seas. The main bioconstructions in the Mediterranean Sea are represented by coralligenous formations, vermetid reefs, deep-sea cold-water corals, Lithophyllum byssoides trottoirs, coral banks formed by the shallow-water corals Cladocora caespitosa or Astroides calycularis, and sabellariid or serpulid worm reefs. Bioconstructions change the morphological and chemicophysical features of primary substrates and create new habitats for a large variety of organisms, playing pivotal roles in ecosystem functioning. In spite of their importance, Mediterranean bioconstructions have not received the same attention that tropical coral reefs have, and the knowledge of their biology, ecology and distribution is still fragmentary. All existing data about the spatial distribution of Italian bioconstructions have been collected, together with information about their growth patterns, dynamics and connectivity. The degradation of these habitats as a consequence of anthropogenic pressures (pollution, organic enrichment, fishery, coastal development, direct physical disturbance), climate change and the spread of invasive species was also investigated. The study of bioconstructions requires a holistic approach leading to a better understanding of their ecology and the application of more insightful management and conservation measures at basin scale, within ecologically coherent units based on connectivity: the cells of ecosystem functioning.

A large non-parasitic population of Savalia savaglia (Bertoloni, 1819) in the Boka Kotorska Bay (Montenegro).

The golden coral Savalia savaglia is a long-living ecosystem engineer of Mediterranean circalittoral assemblages, able to induce necrosis of gorgonians' and black corals' coenenchyme and grow on their cleaned organic skeleton. Despite its rarity, in Boka Kotorska Bay (Montenegro) a shallow population of more than 1000 colonies was recorded close to underwater freshwater springs, which create very peculiar environmental conditions. In this context, the species was extremely abundant at two sites, while gorgonians were rare. The abundance and size of S. savaglia colonies and the diversity of the entire benthic assemblage were investigated by photographic sampling in a depth range of 0-35 m. Several living fragments of S. savaglia spread on the sea floor and small settled colonies (< 5 cm high) suggested a high incidence of asexual reproduction and a non-parasitic behaviour of this population. This was confirmed by studying thin sections of the basal portion of the trunk where the central core, generally represented by the remains of the gorgonian host skeleton, was lacking. The S. savaglia population of Boka Kotorska Bay forms the unique Mediterranean assemblage of the species deserving the definition of animal forest. Recently, temporary mitigation measures for anthropogenic impact were issued by the Government of Montenegro. Nevertheless, due to the importance of the sites the establishment of a permanent Marine Protected Area is strongly recommended.

Sponges associated with octocorals in the Indo-Pacific, with the description of four new species.

Sponges are characterised by high levels of phenotypic plasticity, thus allowing the same species to live in different habitats by taking different shapes. Here we describe 28 sponge species associated with the octocorals Carijoa riisei, Paratelesto rosea and Alertigorgia hoeksemai in Indonesia, Hawai'i and Vietnam, including four species that are new to science (Chondropsis subtilis, Hymedesmia (Hymedesmia) spinata, Hymedesmia (Stylopus) perlucida, Mycale (Aegogropila) furcata). Moreover, a large proportion of the described sponge species (21.4%) represent new records for the studied areas (Indonesia and Hawai'i). In total, we have studied 47 colonies of C. riisei associated with 24 sponge species, 5 colonies of P. rosea associated with 4 species and one colony of A. hoeksemai associated with one sponge species. Collectively, these examples of associations highlight the importance of epibiosis as a biodiversity enhancing process.

Sinularioside, a triacetylated glycolipid from the Indonesian soft coral Sinularia sp., is an inhibitor of NO release.

Chemical analysis of the Indonesian soft coral Sinularia sp. (order Alcyonacea, family Alcyoniidae) afforded a known glucosylcerebroside of the sarcoehrenoside-type and sinularioside (2), a new naturally triacetylated glycolipid containing two α-D-arabinopyranosyl residues and a myristyl alcohol unit. Their complete stereostructures were solved by interpretation of MS and NMR data along with CD analysis of degradation products. Sinularioside proved to moderately inhibit LPS-induced NO release, providing interesting clues into the poorly understood structure-activity relationships for anti-inflammatory glycolipids.

Manadoperoxides, a new class of potent antitrypanosomal agents of marine origin.

Chemical investigation of the marine sponge Plakortis cfr. lita afforded a library of endoperoxyketal polyketides, manadoperoxides B-K (3-5 and 7-13) and peroxyplakoric esters B(3) (6) and C (14). Eight of these metabolites are new compounds and some contain an unprecedented chlorine-bearing THF-type ring in the side chain. The library of endoperoxide derivatives was evaluated for in vitro activity against Trypanosoma brucei rhodesiense and Leishmania donovani. Some compounds, such as manadoperoxide B, exhibited ultrapotent trypanocidal activity (IC(50) = 3 ng mL(-1)) without cytotoxicity. Detailed examination of the antitrypanosomal activity data and comparison with those available in the literature for related dioxane derivatives enabled us to draw a series of structure-activity relationships. Interestingly, it appears that minor structural changes, such as a shift of the methyl group around the dioxane ring, can dramatically affect the antitrypanosomal activity. This information can be valuable to guide the design of optimized antitrypanosomal agents based on the dioxane scaffold.

Leucettamols, bifunctionalized marine sphingoids, act as modulators of TRPA1 and TRPM8 channels.

Leucettamols, bifunctionalized sphingoid-like compounds obtained from a marine sponge Leucetta sp., act as non-electrophilic activators of the TRPA1 channel and potent inhibitors of the icilin-mediated activation of the TRPM8 channel, while they are inactive on CB1, CB2 and TRPV1 receptors. Leucettamols represent the first compounds of marine origin to target TRPA1 and the first class of natural products to inhibit TRPM8 channels. The preparation of a small series of semi-synthetic derivatives revealed interesting details on the structure-activity relationships within this new chemotype of simple acyclic TRP modulators.

Aurantoside J: a new tetramic acid glycoside from Theonella swinhoei. Insights into the antifungal potential of aurantosides.

The chemical investigation of an Indonesian specimen of Theonella swinhoei afforded four aurantosides, one of which, aurantoside J (5), is a new compound. The structure of this metabolite, exhibiting the unprecedented N-α-glycosidic linkage between the pentose and the tetramate units, has been determined through detailed spectroscopic analysis. The four obtained aurantosides have been tested against five fungal strains (four Candida and one Fusarium) responsible of invasive infections in immuno-compromised patients. The non-cytotoxic aurantoside I (4) was the single compound to show an excellent potency against all the tested strains, thus providing valuable insights about the antifungal potential of this class of compounds and the structure-activity relationships.

Fate of lost fishing gears: Experimental evidence of biofouling colonization patterns from the northwestern Mediterranean Sea.

ALDFGs (abandoned, lost or otherwise discharged fishing gears) represent a major pollutant in the world's oceans, including the Mediterranean Sea, but very little is known about their long-term fate in the underwater environment. Here we investigate the destiny of ALDFGs in a coralligenous benthic community by studying the biofouling growing on them at 30 m depth over a three-year period. Results indicate that ALDFGs are quickly covered by biofouling, reaching maximum values of cover in one year. At this depth, the biofouling community results highly influenced by light-dependent seasonal cycles and a general four-steps colonization scheme is tentatively depicted: i) biofouling community is initially dominated by fast-growing organisms, with algae and hydrozoans settling after two weeks; ii) small-sized carbonatic organisms (e.g. bryozoans) appear after two months; iii) the same organisms (including coralline algae) become abundant after five to eight months; iv) conspicuous carbonatic skeletons (serpulids, bivalves and bryozoans) occur after one year increasing in complexity in the following months. The biofouling settled on ALDFGs, as well as growth rates of large carbonatic species, could provide useful information to estimate the age of lost lines and nets in retrieved material and in situ photo footage complementing the overview of the fishing impact in this specific environment. Finally, no signs of nylon degradation are reported, suggesting that the prevailing physical conditions and the biofouling cover extent could alter microplastics release from nylon ALDFGs.

Immunomodulatory Activity of the Marine Sponge, Haliclona (Soestella) sp. (Haplosclerida: Chalinidae), from Sri Lanka in Wistar Albino Rats: Immunosuppression and Th1-Skewed Cytokine Response.

Natural secondary metabolites of sponges of the genus Haliclona are associated with an array of biological activity with therapeutic usage. We investigated the immunopharmacological properties of a presumably novel marine sponge species from Sri Lanka, Haliclona (Soestella) sp. Sponge material was collected from southern Sri Lanka by scuba diving. Sponge identification was based on spicule and skeleton morphology using light microscopy. Selected in vivo and ex vivo tests investigated nonfunctional and functional immunomodulatory activity of the Haliclona (Soestella) sp. crude extract (HSCE) in the Wistar rat model. Compared to the controls, rats orally gavaged daily for 14 consecutive days with 15 mg/kg dose of the HSCE manifested a significant reduction of immune cell counts of total WBCs (by 17%; p < 0.01), lymphocytes (38%), platelets (52%), splenocytes (20%), and bone marrow cells (BMC; 60%) (p < 0.001), with a concurrent increase in the neutrophil : lymphocyte ratio (p < 0.05); RBC counts abated by 53% (p < 0.001). A significant reduction of the splenosomatic index was evident with the 10 and 15 mg/kg doses (p < 0.001). Rat plasma TNF-α cytokine level was augmented by tenfold (p < 0.001), IL-6 level by twofold (p < 0.01) with the 15 mg/kg HSCE treatment, while IL-10 was detectable in rat plasma only with this treatment; the corresponding Th1 : Th2 cytokine ratio (TNF-α : IL-10) was indicative of an unequivocal Th1-skewed cytokine response (p < 0.01). Ex vivo bone marrow cell and splenocyte proliferation were significantly and dose dependently impaired by HSCE (IC50 0.719 and 0.931 µg/mL, respectively; p < 0.05). Subacute toxicity testing established that HSCE was devoid of general toxic, hepatotoxic, and nephrotoxic effects. In conclusion, HSCE was orally active, nontoxic, and effectively suppressed nonfunctional and functional immunological parameters of Wistar rats, suggestive of the potential use of the HSCE as an immunosuppressant drug lead.

Acanthella danerii sp. nov. (Demospongiae, Bubarida, Dictyonellidae) from Chilean fjords (South Pacific Ocean).

The use of SCUBA diving for marine research in the Chilean fjords has increased the possibility of obtaining information on the sponge assemblages living in this environment. However, much work is still needed to achieve a satisfactory knowledge of the benthos of this wide region. As to sponges, just consider that seventeen new species have been recently described by several authors (Hajdu et al., 2013; Fernandez et al., 2016; Bertolino et al., 2019) with detailed information of this benthic fauna for this area. Aim of this work is the description of a new species of Acanthella (Dictyonellidae van Soest, Diaz Pomponi, 1990), Acanthella danerii sp. nov.. It was collected at "Seno Magdalena" (44°61'48.63" S 72°95'83.12" W) (Chilean fjords) by SCUBA, at 20 m depth, on hard substrates of a vertical wall ending on a rocky bottom that slopes down to 32 m during a survey in August 2016.

Assessment and distribution of seafloor litter on the deep Ligurian continental shelf and shelf break (NW Mediterranean Sea).

Litter has become a common component of marine ecosystems, and its assessment and monitoring are now of primary interest. Here, we characterized seafloor litter in the mesophotic depth range of the Ligurian Sea, one of the most urbanized basins of the Mediterranean Sea. Eighty sites (30-220 m depth) were investigated by by means of ROV video transects. Data regarding environmental parameters and anthropic pressures were used to explain the distribution of human-derived objects. Litter resulted widely distributed in all the investigated areas, with maximum densities of 7900 items ha-1. It was mainly represented by abandoned fishing gears, with highest abundances found on the deep coastal rocky shoals exploited by artisanal and recreational fishermen. Urban litter, primarily represented by plastics, was generally higher close to shore, especially near river mouths and large harbors. Litter densities reported here are among the highest ever recorded in Europe, denoting the urgency for specific actions to prevent further spreading.

The sponge fauna of the Seno Magdalena and Puyuhuapi Fjord (Chile), with a description of two new species.

The presence of fjords, islands and channels originating from glacial erosion and ice cap retreat, makes the Chilean benthic biodiversity difficult to explore and study. Our survey of this region allowed the identification of 29 Demospongiae species in total. Two of them are new to science and here described: Biemna lutea sp. nov., and Hamigera cleistochela sp. nov.. Two species (Clathria (Clathria) microxa and Lissodendoryx (Ectyodoryx) patagonica,) are new for the region and the Chilean fjords. Lissodendoryx (Ectyodoryx) patagonica was found for the second time after the original description by Ridley Dendy, 132 years ago. These results - considering the small number of species identified on the whole - are promising and confirm that the marine biodiversity of Chilean fjords is remarkable but not well known yet.

Megabenthic communities of the Ligurian deep continental shelf and shelf break (NW Mediterranean Sea).

The Ligurian Sea is one of the most studied Mediterranean basins. Since the beginning of the last century, many research expeditions have characterized its benthic and pelagic fauna through scuba diving and trawl surveys. However, a large knowledge gap exists about the composition of benthic communities extending into the so-called mesophotic or twilight depth range, currently under intense pressure from commercial and recreational fishing. A series of visual surveys, carried out by means of remotely operated vehicles between 2012 and 2018, were conducted along the Ligurian deep continental shelf and shelf break, between 30 and 210 m depth, in order to characterize the main benthic biocoenoses dwelling at this depth range and to determine the most relevant environmental factors that explain their spatial distribution. Deep circalittoral communities of the Ligurian Sea were represented by a mixture of species belonging to the deepest extension of shallow-water habitats and deep circalittoral ones. Twelve major biocoenoses were identified, each one characterized by specific preferences in depth range, substrate type and seabed slope. Those biocoenoses included gorgonian and hydrozoan forests, dense keratose sponge grounds, Dendrophyllia cornigera gardens, bryozoan beds and soft-bottom meadows of sabellid polychaetes and soft-corals. Other less common aggregations included six forests of black corals and two populations of Paramuricea macrospina. A georeferenced database has been created in order to provide information to managers and stakeholders about the location of the identified communities and high-diversity areas, aiming to facilitate sustainable long-term conservation of the Ligurian benthic ecosystem.

Thirty year ecosystem trajectories in a submerged marine cave under changing pressure regime.

Marine caves are unique and vulnerable habitats exhibiting high biodiversity and heterogeneity, but threatened by multiple global and local disturbances. Marine caves, although widely distributed along the Mediterranean coast, suffer for the lack of quantitative data on their structure and function, which hinder their conservation status assessment. Thanks to the availability of a nearly 30-year-long series of data (1986-2013), we evaluated ecosystem change in the Bergeggi marine cave (Ligurian Sea, NW Mediterranean), a cave with a complex shape and high habitat heterogeneity. Non-taxonomic descriptors were adopted, namely growth forms (GF) and trophic guilds (TG), which are informative about ecosystem structure and functioning, respectively. The cave experienced a general trend of change during the last three decades, mainly due to the decline in the cover of sessile organisms (especially 3-dimensional forms) matched by an increase of turf and sediment, thus causing the structural and functional homogenization of the cave community. While change before 2004 had been attributed to climatic factors (especially to the summer heat waves of 1999 and 2003), the most important rate of change was observed between 2009 and 2013, coinciding with recent major beach nourishments and the extension of the neighbouring Vado Ligure harbour, thus providing evidences on the importance of local disturbances deriving from coastal interventions. Monitoring the status of cave ecosystems is urgently needed, and the use of effective indicators, such as the specific traits here adopted (morphology and feeding strategy), could provide effective tools to assist marine cave conservation.

A predictive approach to benthic marine habitat mapping: Efficacy and management implications.

The availability of marine habitats maps remains limited due to difficulty and cost of working at sea. Reduced light penetration in the water hampers the use of optical imagery, and acoustic methods require extensive sea-truth activities. Predictive spatial modelling may offer an alternative to produce benthic habitat maps based on complete acoustic coverage of the seafloor together with a comparatively low number of sea truths. This approach was applied to the coralligenous reefs of the Marine Protected Area of Tavolara - Punta Coda Cavallo (NE Sardinia, Italy). Fuzzy clustering, applied to a set of observations made by scuba diving and used as sea truth, allowed recognising five coralligenous habitats, all but one existing within EUNIS (European Nature Information System) types. Variable importance plots showed that the distribution of habitats was driven by distance from coast, depth, and lithotype, and allowed mapping their distribution over the MPA. Congruence between observed and predicted distributions and accuracy of the classification was high. Results allowed calculating the occurrence of the distinct coralligenous habitats in zones with different protection level. The five habitats are unequally protected since the protection regime was established when detailed marine habitat maps were not available. A SWOT (Strengths-Weaknesses-Opportunities-Threats) analysis was performed to identify critical points and potentialities of the method. The method developed proved to be reliable and the results obtained will be useful when modulating on-going and future management actions in the studied area and in other Mediterranean MPAs to develop conservation efforts at basin scale.

From depth to regional spatial genetic differentiation of Eunicella cavolini in the NW Mediterranean.

Connectivity studies in the marine realm are of great importance to understand the evolutionary potential of populations in a context of growing pressures on the marine environment. Here, we investigated the effect of the local, regional, and depth spatial scale on the population genetic structure of the yellow gorgonian Eunicella cavolini, one of the most common octocoral species of the Mediterranean hard-bottom communities. This species, along with other sessile metazoans typical of coralligenous ecosystems, plays an important role in supporting biodiversity, but is also impacted by direct and indirect consequences of human activities, such as physical destruction or mortality events due to thermal anomalies. Samples were taken from 15 sites located in the Ligurian Sea (NW Mediterranean) in two adjacent regions 100 kilometres apart, i.e. from the areas of Marseille (France) and Portofino (Genoa, Italy), and were analysed using six microsatellite loci. A pattern of isolation by distance was observed at the regional as well as the local scales. Although E. cavolini showed less genetic structure than other Mediterranean octocorallian species, we observed a significant genetic differentiation between populations a few kilometres apart. A low genetic differentiation was also observed between shallow and deep populations. The occurrence of genetically differentiated populations of E. cavolini at the scale of kilometres has important consequences for the management of this species and of the associated communities.

The two facets of species sensitivity: Stress and disturbance on coralligenous assemblages in space and time.

Marine coastal ecosystems are affected by a vast array of human-induced disturbances and stresses, which are often capable of overwhelming the effects of natural changes. Despite the conceptual and practical difficulty in differentiating between disturbance and stress, which are often used interchangeably, the two terms bear different ecological meanings. Both are external agents, but the former causes mortality or physical damage (subtraction of biomass), whereas the latter causes physiological alteration (reduction in productivity). Sensitivity of marine organisms may thus have a dual connotation, being influenced in different ways by disturbance and by stress following major environmental change. Coralligenous assemblages, which shape unique biogenic formations in the Mediterranean Sea, are considered highly sensitive to change. In this paper, we propose a method to differentiate between disturbance and stress to assess the ecological status of the coralligenous assemblages. Disturbance sensitivity level (DSL) and stress sensitivity level (SSL) of the sessile organisms thriving in the coralligenous assemblages were combined into the integrated sensitivity level of coralligenous assemblages (ISLA) index. Changes in the coralligenous status were assessed in space, along a gradient of stress (human-induced pressures) at several sites of the western Mediterranean, and in time, from a long-term series (1961-2008) at Mesco Reef (Ligurian Sea) that encompasses a mass mortality event in the 1990s. The quality of the coralligenous assemblages was lower in highly urbanised sites than that in sites in both marine protected areas and areas with low levels of urbanisation; moreover, the quality of the assemblages at Mesco Reef decreased during the last 50years. Reduction in quality was mainly due to the increase in stress-tolerant and/or opportunist species (e.g. algal turfs, hydroids and encrusting sponges), the disappearance of the most sensitive macroalgae (e.g. Udoteaceae and erect Rhodophyta) and macro-invertebrates (e.g. Savalia savaglia, Alcyonium coralloides and Smittina cervicornis), and the appearance of invasive alien algal species. Although the specific indices of SSL or DSL well illustrated the changes in the spatial or temporal datasets, respectively, their integration in the ISLA index was more effective in measuring the change experienced by the coralligenous assemblages in both space and time.

Demosponge diversity from North Sulawesi, with the description of six new species.

Sponges are key components of the benthic assemblages and play an important functional role in many ecosystems, especially in coral reefs. The Indonesian coral reefs, located within the so-called "coral triangle", are among the richest in the world. However, the knowledge of the diversity of sponges and several other marine taxa is far from being complete in the area. In spite of this great biodiversity, most of the information on Indonesian sponges is scattered in old and fragmented literature and comprehensive data about their diversity are still lacking. In this paper, we report the presence of 94 species recorded during different research campaigns mainly from the Marine Park of Bunaken, North Sulawesi. Six species are new for science and seven represent new records for the area. Several others are very poorly known species, sometimes recorded for the second time after their description. For most species, besides field data and detailed descriptions, pictures in vivo are included. Moreover, two new symbiotic sponge associations are described. This work aims to increase the basic knowledge of Indonesian sponge diversity as a prerequisite for monitoring and conservation of this valuable taxon.