Minimalist barcodes for sponges: a case study classifying African freshwater Spongillida.

African sponges, particularly freshwater sponges, are understudied relative to demosponges in most other geographical regions. Freshwater sponges (Spongillida) likely share a common ancestor; however, their evolutionary history, particularly during their radiation into endemic and allegedly cosmopolitan groups, is unclear. Freshwater sponges of at least 58 species of 17 genera and four families are described from Central and Eastern Africa, but the diversity is underestimated due to limited distinguishable morphological features. The discovery of additional cryptic species is very likely with the use of molecular techniques such as DNA barcoding. The Royal Museum of Central Africa (MRAC, Tervuren, Belgium) hosts one of the largest collections of (Central) African freshwater sponge type material. Type specimens in theory constitute ideal targets for molecular taxonomy; however, the success is frequently hampered by DNA degradation and deamination, which are a consequence of suboptimal preservation techniques. Therefore, we genotyped African demosponge holotype material of the MRAC with specific short primers suitable for degenerated tissue and compare the results with the current, morphology-based classification. Our results demonstrate the utility of minimalistic barcodes for identification of sponges, potentially enabling efficient identification of individuals in taxonomic or metabarcoding studies, and highlight inconsistencies in the current freshwater sponge classification.

A Collection of Bioactive Nitrogen-Containing Molecules from the Marine Sponge Acanthostrongylophora ingens.

Thirteen nitrogen-containing molecules (1a/1b and 2-12) were isolated from the Indonesian sponge Acanthostrongylophora ingens, highlighting the richness of this organism as a source of alkaloids. Their structures were elucidated using one- and two-dimensional NMR spectroscopy and HR-ESI-MS, while the stereochemistry of the diketopiperazines was established using Marfey's method. All compounds were screened in our standard bioactivity assays, including antibacterial, antikinases, and amyloid ß-42 assays. The most interesting bioactivity result was obtained with the known acanthocyclamine A (3), which revealed for the first time a specific Escherichia coli antimicrobial activity and an inhibitory effect on amyloid ß-42 production induced by aftin-5 and no cytotoxicity at the dose of 26 µM. These results highlight the potentiality of a bipiperidine scaffold as a promising skeleton for preventing or reducing the production of amyloid ß-42, a key player in the initiation of Alzheimer's disease.

Chloromethylhalicyclamine B, a Marine-Derived Protein Kinase CK1δ/ε Inhibitor.

The halogenated alkaloid chloromethylhalicyclamine B (1), together with the known natural compound halicyclamine B (2), was isolated from the extract of the sponge Acanthostrongylophora ingens. The structure of compound 1 was determined by spectroscopic means, and it was shown that 1 is produced by reaction of 2 with CH2Cl2 used for extraction. Compound 1 was a selective CK1δ/ε inhibitor with an IC50 of 6 µM, while the natural compound 2 was inactive. The absolute configuration of 1 was determined by quantum mechanical calculation of its ECD spectrum, and this also determined the previously unknown absolute configuration of the parent halicyclamine B (2). Computational studies, validated by NOESY data, showed that compound 1 can efficiently interact with the ATP-binding site of CK1δ in spite of its globular structure, very different from the planar structure of known inhibitors of CK1δ. This opens the way to the design of a new structural type of CK1δ/ε inhibitors.

Reef sponges of the genus Agelas (Porifera: Demospongiae) from the Greater Caribbean.

The genus Agelas comprises a group of tropical and subtropical reef sponges that contains large, long-lived, often brightly colored and conspicuous species, distributed throughout the tropica l western Atlantic, temperate northern Atlantic (Mediterranean Sea), and western and central Indo-Pacific Realms. Among tropical sponge genera, Agelas is one with similar species richness in the Greater Caribbean in comparison to the Indo Pacific. The presence of verticillated acanthostyle spicules and a fibroreticulate skeleton of spongin fibres cored and/or echinated by spicules characterize this group. Taxonomic identification relies on a combination of characters, where external morphology and color play a key role, owing to the paucity of microscopical characters. Thus, there is still a great deal of taxonomic confusion, even for the more common species. We carried out a detailed revision of Agelas species throughout the Greater Caribbean area using classic taxonomic tools. Samples and observations covered Colombia, Belize, Jamaica, the Bahamas, Barbados, Curaçao and Venezuela, and included type material from major museum collections. According to our results, the genus Agelas in the Caribbean has at least thirteen valid species, viz. Agelas sceptrum (Lamarck, 1815); A. dispar Duchassaing & Michelotti, 1864;  A. dilatata Duchassaing & Michelotti, 1864; A. clathrodes (Schmidt, 1870);  A. cervicornis (Schmidt, 1870); A. conifera (Schmidt, 1870); A. schmidti Wilson, 1902;   A. tubulata Lehnert & van Soest, 1996; A. wiedenmayeri Alcolado, 1984;  A. citrina Gotera & Alcolado, 1987; A. sventres Lehnert & van Soest, 1996; A. repens Lehnert & van Soest, 1998; and A. cerebrum Assmann et al., 2001. We found that variation of microscopic characteristics like skeleton arrangement, number of verticills and their spines, and spicule length and width, can be used as taxonomic tools, but only in a thorough comparison with other species in the same sub-regional context. Thus, a certain degree of familiarity with the genus' regional variation is often required. The richness and distribution of these species in the Caribbean area show north/south differences and other ecological patterns are evident. 

Diversity of sponges (Porifera) from cryptic habitats on the Belize barrier reef near Carrie Bow Cay.

The Caribbean barrier reef near Carrie Bow Cay, Belize, has been a focus of Smithsonian Institution (Washington) reef and mangrove investigations since the early 1970s. Systematics and biology of sponges (Porifera) were addressed by several researchers but none of the studies dealt with cryptic habitats, such as the shaded undersides of coral rubble, reef crevices, and caves, although a high species diversity was recognized and samples were taken for future reference and study. This paper is the result of processing samples taken between 1972 and 2012. In all, 122 species were identified, 14 of them new (including one new genus). The new species are Tetralophophora (new genus) mesoamericana, Geodia cribrata, Placospongia caribica, Prosuberites carriebowensis, Timea diplasterina, Timea oxyasterina, Rhaphidhistia belizensis, Wigginsia curlewensis, Phorbas aurantiacus, Myrmekioderma laminatum, Niphates arenata, Siphonodictyon occultum, Xestospongia purpurea, and Aplysina sciophila. We determined that about 75 of the 122 cryptic sponge species studied (61%) are exclusive members of the sciophilic community, 47 (39 %) occur in both, light-exposed and shaded or dark habitats. Since we estimate the previously known sponge population of Carrie Bow reefs and mangroves at about 200 species, the cryptic fauna makes up 38 % of total diversity.

Structure, synthesis, and biological activity of a C-20 bisacetylenic alcohol from a marine sponge Callyspongia sp.

An optically inactive C-20 bisacetylenic alcohol, (4E,16E)-icosa-4,16-diene-1,19-diyne-3,18-diol, was isolated from a marine sponge Callyspongia sp. as a result of screening of antilymphangiogenic agents from marine invertebrates. An optical resolution using chiral-phase HPLC gave each enantiomer, (-)-1 and (+)-2. Because the natural and synthetic enantiomers 1 and 2 showed different biological properties, we investigated the structure-activity relationships of bisacetylenic alcohols using 11 synthetic derivatives, and it is clarified that the essential structural unit for antiproliferative activity is the "1-yn-3-ol" on both termini and that there is a minimum chain length that connects the "1-yn-3-ol" moieties.

In vitro pharmacological and toxicological effects of norterpene peroxides isolated from the Red Sea sponge Diacarnus erythraeanus on normal and cancer cells.

Eight cyclic peroxide norterpenoids, compounds 1-8, have been isolated and characterized from the Red Sea sponge Diacarnus erythraeanus, including two new norsesterterpene derivatives (3, 4). Among these metabolites, (-)-muqubilin A (5) (nine cell lines analyzed) and the new compounds 3 and 4 (seven cell lines analyzed) displayed mean IC50 growth inhibitory concentrations in vitro of <10 µM, while the remaining compounds (1, 6-8) were inactive in these cancer cell lines. Compound 5 displayed no selectivity between normal and cancer cells in terms of in vitro growth inhibition. Quantitative video microscopy analysis carried out on (-)-muqubilin A-treated cells validated the data obtained by means of the MTT colorimetric assay, while flow cytometry analysis revealed ROS production but no induction of apoptosis in cancer cells.

Acetylcholinesterase-inhibitory activities of the extracts from sponges collected in mauritius waters.

Patients diagnosed with Alzheimer's disease (AD) show a characteristic neurochemical deficit of acetylcholine, especially in the basal forebrains. The use of acetylcholinesterase (AChE) inhibitors to retard the hydrolysis of acetylcholine has been suggested as a promising strategy for AD treatment. In this study, we evaluated the acetylcholinesterase inhibitory (AChEI) activities of 134 extracts obtained from 45 species of marine sponges. Thin-layer chromatography (TLC) and microplate assays reveal potent acetylcholinsterase inhibitory activities of two AcOEt extracts from the sponges Pericharax heteroraphis and Amphimedon navalis PULITZER-FINALI. We further investigated the inhibitory kinetics of the extracts and found them to display mixed competitive/noncompetitive inhibition and associated their inhibitory activity partly to terpenoids. Acetylcholinesterase inhibitors from marine organisms have been rarely studied, and this study demonstrated the potential of marine sponges as a source of pharmaceutical leads against neurodegenerative diseases.

Genetic data confirms the enigmatic demosponge Janulum as haplosclerid.

Historically, sponge classification is based on the interpretation of morphological characters, whose phylogenetic information content is frequently limited, subject to homoplasies, or prone to environmental plasticity (e.g., Chombard et al. 1998). Therefore, the currently accepted order-level classification of its largest class, Demospongiae, has been largely revised with molecular phylogenetic data (Morrow & Cárdenas 2015). Nevertheless, numerous sponge genera with ambiguous or provisoric phylogenetic placement still await definite classification.

Halenaquinone, a chemical compound that specifically inhibits the secondary DNA binding of RAD51.

Mutations and single-nucleotide polymorphisms affecting RAD51 gene function have been identified in several tumors, suggesting that the inappropriate expression of RAD51 activity may cause tumorigenesis. RAD51 is an essential enzyme for the homologous recombinational repair (HRR) of DNA double-strand breaks. In the HRR pathway, RAD51 catalyzes the homologous pairing between single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA), which is the central step of the HRR pathway. To identify a chemical compound that regulates the homologous-pairing activity of RAD51, in the present study, we screened crude extract fractions from marine sponges by the RAD51-mediated homologous-pairing assay. Halenaquinone was identified as an inhibitor of the RAD51 homologous-pairing activity. A surface plasmon resonance analysis indicated that halenaquinone directly bound to RAD51. Intriguingly, halenaquinone specifically inhibited dsDNA binding by RAD51 alone or the RAD51-ssDNA complex, but only weakly affected the RAD51-ssDNA binding. In vivo, halenaquinone significantly inhibited the retention of RAD51 at double-strand break sites. Therefore, halenaquinone is a novel type of RAD51 inhibitor that specifically inhibits the RAD51-dsDNA binding.

Horny sponges and their affairs: on the phylogenetic relationships of keratose sponges.

The demosponge orders Dictyoceratida and Dendroceratida are historically assigned to the keratose (or "horny") sponges, which are mostly devoid of primary skeletal elements, but possess an elaborate skeleton of organic fibres instead. This paucity of complex mineral skeletal elements makes their unambiguous classification and phylogenetic reconstruction based on morphological features difficult. Here we present the most comprehensive molecular phylogeny to date for the Dendroceratida, Dictyoceratida, and also other sponge orders that largely lack a mineral skeleton or skeletal elements at all (i.e. Verongida, Halisarcida, Chondrosida), based on independent mitochondrial and nuclear markers. We used molecular data to validate the coherence of all recognised orders, families and subfamilies that are currently defined using morphological characteristics. We discussed the significance of morphological and chemotaxonomic characters for keratose sponges, and suggested adapted definitions for the classification of dendroceratid, dictyoceratid, and verongid higher taxa. Also, we found that chondrosid sponges are non-monophyletic with respect to Halisarcida. Verongida and Dendroceratida were monophyletic, however most of their classically recognised families were not recovered. This indicated that the current distinction between dendritic and mesh-like fibre skeletons is not significant at this level of classification. Dysideidae were found to be the sister-group to the remaining Dictyoceratida. Irciniidae formed a distinct clade, however Thorectidae and Spongiidae could not be separated with the molecular markers used. Finally, we are establishing the name Verongimorpha for the clade combining verongid, chondrosid and halisarcid taxa and readjust the content of its sister-clade Keratosa.

The terminology of sponge spicules.

Sponges (Porifera) are a diverse and globally distributed clade of benthic organisms, with an evolutionary history reaching at least the Ediacaran-Cambrian (541 Ma) boundary interval. Throughout their research history, sponges have been subjects of intense studies in many fields, including paleontology, evolutionary biology, and even bioengineering and pharmacology. The skeletons of sponges are mostly characterized by the presence of mineral elements termed spicules, which structurally support the sponge bodies, though they also minimize the metabolic cost of water exchange and deter predators. The description of the spicules' shape and the skeleton organization represents the fundamental basis of sponge taxonomy and systematics. Here, we provide an illustrated catalogue of sponge spicules, which is based on previous works on sponge spicules, for example, and gathers and updates all terms that are currently used in sponge descriptions. Each spicule type is further illustrated through high quality scanning electron microscope micrographs. It is expected to be a valuable source that will facilitate spicule identification and, in certain cases, also enable sponge classification.

Shishicrellastatins, inhibitors of cathepsin B, from the marine sponge Crella (Yvesia) spinulata.

Two dimeric steroid derivatives, shishicrellastatin A (1) and B (2), have been isolated as cathepsin B inhibitors from the marine sponge Crella (Yvesia) spinulata. Their structures were determined by interpretation of spectroscopic data. Shishicrellastatins inhibit cathepsin B with an IC(50) value of 8 µg/mL each.

Bioactive terpenes from Spongia officinalis.

The terpene metabolite pattern of Mediterranean Spongia officinalis was chemically investigated. This study resulted in the isolation of a series of sesterterpenes and C21 furanoterpenes, according to the literature data on this sponge. Four new oxidized minor metabolites (compounds 1, 2, 3, and 4) were isolated along with six known compounds of the furospongin series (compounds 5-8, 9, and 10) and three scalarane sesterterpenes (compounds 11-13). Interestingly, tetrahydrofurospongin-2 (6) and dihydrofurospongin-2 (7), which were among the main metabolites, induced biofilm formation by Escherichia coli. All compounds isolated were also assayed for antibacterial and antifungal properties.

9'-Epi-3ß,3'ß-dimethylxestospongin C, a new macrocyclic diamine alkaloid from the Hainan sponge Neopetrosia exigua.

A new BIS-1-oxaquinolizidine alkaloid, 9'- EPI-3 ß,3' ß-dimethylxestospongin C (2), and three known related analogues have been isolated from the Hainan sponge Neopetrosia exigua.

Trans-dimer D, a novel dimeric sesquiterpene with a bis-bisabolene skeleton from a Hainan sponge Axinyssa variabilis.

A novel bisabolene sesquiterpene dimer named trans-dimer D (1) and its diastereoisomer trans-dimer C (2) reported in our previous work have been isolated as an inseparable mixture in a ratio of 1.5:1 from the South China Sea sponge Axinyssa variabilis. The structure of 1 was determined on the basis of extensive spectroscopic analysis and by comparison of its NMR spectral data with those of structurally related compounds. Compound 1 represents the fourth example of a sesquiterpene dimer with a bis-bisabolene skeleton.

emAlcyonium/em embursa/em Linnaeus, 1758 is not a sponge.

In the 10th edition of the Systema Naturae (Linnaeus, 1758), which is the starting point of the Code for Zoological Nomenclature (ICZN Art. 3), Linnaeus named three species of the genus Alcyonium, A. arboreum, A. digitatum, and A. bursa. The genus name Alcyonium was based on the 16th and 17th century pre-Linnaean use for a diversity of marine organisms, including cnidarians, sponges, bryozoans, and algae. In the first valid presentation of the genus name, Linnaeus narrowed this down to comprise two clear cnidarians (A. arboreum, currently Paragorgia arborea, and A. digitatum, still accepted under this name and subsequently assigned as type species), but the pre-Linnaean diversity perhaps explains why the third species, A. bursa, was not recognized as a cnidarian. Linnaeus defined it as 'Alcyonium acaule pulposum subglobosum. Habitat in O. Europaea.' (translated as: Alcyonium without stalk, fleshy, semiglobular. From the European Ocean).' Attempts to fix its identity among contemporary authors at the end of the 18th and beginning of the 19th century followed a checkered course, with opinions varying from algae to tunicates and sponges.

Mycale species of the tropical Indo-West Pacific (Porifera, Demospongiae, Poecilosclerida).

The species of the cosmopolitan sponge genus Mycale occurring in the tropical Indo-West Pacific region and adjacent subtropical waters are reviewed taxonomically. Specimens incorporated in the collections of the Naturalis Biodiversity Center form the basis of this comprehensive study, supplemented by (type) specimens borrowed from or examined in other institutions. Specimens available numbered 351, belonging to 44 species, including 14 species new to science, Mycale (Aegogropila) prognatha sp.nov., Mycale (Carmia) amiri sp.nov., Mycale (Carmia) fungiaphila sp.nov., Mycale (Carmia) monomicrosclera sp.nov., Mycale (Carmia) tenuichela sp.nov., Mycale (Carmia) tubiporicola sp.nov., Mycale (Carmia) tydemani sp.nov., Mycale (Mycale) asigmata sp.nov., Mycale (Mycale) grandoides sp.nov., Mycale (Mycale) sundaminorensis sp.nov., Mycale (Naviculina) mascarenensis sp.nov., Mycale (Paresperella) sceptroides sp.nov., Mycale (Paresperella) seychellensis sp.nov., Mycale (Zygomycale) sibogae sp.nov. Three species, indicated by the designation 'aff.', were not definitely assigned to known or new species due to uncertainty of their identity. The genus Kerasemna, previously considered a junior synonym of Mycale, was revived as an additional subgenus Mycale (Kerasemna). One species, previously assigned to the genus Desmacella as D. lampra De Laubenfels is here reassigned to Mycale, subgenus at present undecided. Additionally, species previously reported from the region but not represented in our collections are briefly characterized and discussed. We propose new names Mycale (Mycale) mauricei nom.nov. for Mycale macrochela Burton (junior primary homonym of Mycale fistulata var. macrochela Hentschel) and Mycale (Mycale) bouryesnaultae nom.nov. for Mycale (Mycale) fibrosa Boury-Esnault Van Beveren (junior primary homonym of Mycale (Aegogropila) adhaerens subsp. fibrosa Koltun). Keys to the species of each subgenus occurring in the region are provided. The opportunity of having studied this comprehensive set of species and specimens from the tropical Indo-West Pacific is taken to review and discuss the morphological and biogeographical data gathered so far on the genus Mycale. The genus currently comprises approximately 255 accepted species, with highest diversity focused in tropical Atlantic and Indo-West Pacific regions as well as in warm-temperate Mediterranean-Atlantic regions.

Species boundaries and phylogenetic relationships between Atlanto-Mediterranean shallow-water and deep-sea coral associated Hexadella species (Porifera, Ianthellidae).

Coral reefs constitute the most diverse ecosystem of the marine realm and an increasing number of studies are focusing on coral species boundaries, distribution, and on processes that control species ranges. However, less attention has been paid to coral associated species. Deep-sea sponges dominate cold-water coral ecosystems, but virtually nothing is known about their molecular diversity. Moreover, species boundaries based on morphology may sometimes be inadequate, since sponges have few diagnostic characters. In this study, we investigated the molecular diversity within the genus Hexadella (Porifera, Demospongiae, Verongida, Ianthellidae) from the European shallow-water environment to the deep-sea coral ecosystems. Three molecular markers were used: one mitochondrial (COI) and two nuclear gene fragments (28S rDNA and the ATPS intron). Phylogenetic analyses revealed deeply divergent deep-sea clades congruent across the mitochondrial and nuclear markers. One clade contained specimens from the Irish, the Scottish, and the Norwegian margins and the Greenland Sea (Hexadella dedritifera) while another clade contained specimens from the Ionian Sea, the Bay of Biscay, and the Irish margin (H. cf. dedritifera). Moreover, these deeply divergent deep-sea clades showed a wide distribution suggesting a connection between the reefs. The results also point to the existence of a new deep-sea species (Hexadella sp.) in the Mediterranean Sea and of a cryptic shallow-water species (Hexadella cf. pruvoti) in the Gorringe Bank. In contrast, low genetic differentiation between H. cf. dedritifera and H. pruvoti from the Mediterranean Sea was observed. All Hexadella racovitzai specimens from the Mediterranean Sea (shallow and deep) to the Atlantic formed a monophyletic group.

Aaptamine, an alkaloid from the sponge Aaptos suberitoides, functions as a proteasome inhibitor.

Aaptamine (1), isoaaptamine (2), and demethylaaptamine (3) were isolated from the marine sponge Aaptossuberitoides collected in Indonesia as inhibitors of the proteasome. They inhibited the chymotrypsin-like and caspase-like activities of the proteasome with IC(50) values of 1.6-4.6 microg/mL, while they showed less inhibition of the trypsin-like activity of the proteasome. The three compounds showed cytotoxic activities against HeLa cells, but their cytotoxicity did not correlate with their potency as proteasome inhibitors, strongly suggesting that their proteasomal inhibitory activity is dispensable to their cytotoxicity.