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Confined marine environments are particularly susceptible to climate change and anthropic pressures. Indeed, the long-term monitoring of benthic assemblages in these environments allows us to understand the direction of changes over time. The demosponge Geodia cydonium is a suitable study case, since it is widely represented in many Mediterranean environments, while being a long-living and important habitat-forming species. Here, we report the results of a descriptive study on temporal and spatial variations of this demosponge in three semi-enclosed environments along the Italian coast: Marsala Lagoon, Porto Cesareo Bay, and Mar Piccolo of Taranto. At Marsala and Porto Cesareo, the study compares the present data with those reported by the literature at the end of the 1990s. Caused by the modification of its preferential habitats, the data indicated the loss and a remarkable regression of this species at Porto Cesareo and Marsala, respectively. In addition, we hypothesized that the increase in severe weather phenomena in the small Bay of Porto Cesareo recorded during the last 20 years may have had a marked impact on water mass, thus affecting the sponge assemblages. At Taranto, despite a remarkable environmental degradation, G. cydonium has appeared stable and persistent in the last 45 years, thus representing one of the richest and most well-preserved populations in the Mediterranean Sea.

Climate change is increasing the frequency, magnitude, and socio-economic costs of unprecedented extreme weather events and disasters. An exceptional stranding event of benthic species, mainly represented by the ecosystem engineer Porifera (Demospongiae) Geodia cydonium (Linnaeus, 1767) was observed in the Gulf of Oristano (Sardinia, Italy, Western Mediterranean) in 2018 after an extreme meteorological event. The stranding phenomenon was assessed and quantified by means of quadrats sampled along the beach highlighting all the conspicuous species; the benthic sponge community in the area was evaluated through underwater SCUBA transects. Stranding reached density peaks of up to 0.88 ind m−2, and the most abundant species reported was the sponge G. cydonium, followed by molluscs, bryozoans, a rare teleost fish, and many other species. The putative source populations were identified in quite shallow (less than 2.5-m depth) Posidonia oceanica (Linnaeus) Delile, 1813 meadows, where the sponge shows a mean density of 2.0 ind m−2. The present massive stranding event is the first in Sardinia and in the Mediterranean for this species, whose distribution is still poorly known. The living population of the gulf still seems in good status, and probably only a little percentage of specimens has been struck by the exceptionally strong hurricane. This work highlights the relevance of stranding events as opportunities for providing information about local biodiversity; biological traits of G. cydonium and the need of effective monitoring plans to fill the lack of knowledge about the distribution of this protected species are discussed.

Marine sponges (Porifera) live in a symbiotic relationship with microorganisms, primarily bacteria. Recently, several studies indicated that sponges are the most prolific source of biologically-active compounds produced by symbiotic microorganisms rather than by the sponges themselves. In the present study we characterized the bacterial symbionts from two Demospongiae, Ircinia muscarum and Geodia cydonium. We amplified 16S rRNA by PCR, using specific bacterial-primers. The phylogenetic analysis revealed the presence of nine bacterial clones from I. muscarum and ten from G. cydonium. In particular, I. muscarum resulted enriched in Bacillus species and G. cydonium in Proteobacterium species. Since these bacteria were able to produce secondary metabolites with potential biotechnological and biopharmaceutical applications, we hypothesized that I. muscarum and G. cydonium could be a considered as a “gold mine” of natural products.

The [beta][gamma]-crystallin superfamily includes highly diverse proteins belonging to all of the kingdoms of life. Based on structural topology, these proteins are considered to be evolutionarily related to the long-lived [beta][gamma]-crystallins that constitute the vertebrate eye lens. This study reports the crystallographic structure at 0.99 Å resolution of the two-domain [beta][gamma]-crystallin (geodin) from the sponge Geodia cydonium. This is the most ancient member of the [beta][gamma]-crystallin superfamily in metazoans. The X-ray structure shows that the geodin domains adopt the typical [beta][gamma]-crystallin fold with a paired Greek-key motif, thus confirming the hypothesis that the crystallin-type scaffold used in the evolution of bacteria and moulds was recruited very early in metazoans. As a significant new structural feature, the sponge protein possesses a unique interdomain interface made up by pairing between the second motif of the first domain and the first motif of the second domain. The atomic resolution also allowed a detailed analysis of the calcium-binding site of the protein.

The reproductive cycle of Geodia cydonium in a semi-enclosed Mediterranean bay (Porto Cesareo, SW Apulia) was studied with monthly frequency over a 3-year period. The investigation was carried out by uti- lizing a technique consisting of tagging ten individuals with a PVC stick and cutting oV, by means of a metal cork borer, small samples (cylinders about 5 cm 3 in vol- ume) from each of them for histological analysis. Sex- ual reproductive elements were detected in all individuals, but in the third year some specimens showed reduced reproductive activity or complete infertility. Spermatogenesis occurred in a short period (from June to August) whereas oogenesis lasted longer (from spring to late summer). Spermatic cysts occurred when the frequency of specimens with oocytes reached its maximum values. A relationship between water temperature and the onset of gamete diVerentiation was observed. G. cydonium is here conWrmed ovipa- rous and gonochoric with a sex ratio in favour of the females. However, in contrast with current literature on Porifera—which suggests that only a limited num- ber of sponges, all belonging to the same species are sexually active—the sexual reproduction of this species involves all the examined individuals. This Wnding can be explained by the methodological approach used in this research, which diVers from the traditional way of assessing the sponge reproductive cycle by analysing randomly collected specimens within a population. The study of a series of individuals over time represents a better "model technique" for investigating sponge sex- ual reproduction and the eVect of environmental parameters on gamete diVerentiation.

The sterol and fatty acid composition of three Adriatic Sea sponges (Geodia cydonium and two unidentified Tedania sp.), collected at the same time and same place, was established. Twenty-four sterols and forty fatty acids were identified. The identical ecological conditions, including the diet, allowed us to apply the results obtained for taxonomical conclusions, based on the biodiversity of the investigated sponges. On the basis of the sterol composition they can be separated into two groups: Tedania and Geodia sponges. The sterol and fatty acid composition indicates that the two investigated Tedania samples might be different species or subspecies

Sponges (Porifera) represent the lowest metazoan phylum, characterized by a pronounced plasticity in the determination of cell lineages, and they are the closest related taxon to the hypothetical ancestral animal, the Urmetazoa, from which the metazoan lineages diverged. In a first approach to elucidate the molecular mechanisms controlling the switch from the cell lineage with a putative indefinite growth capacity to senescent, somatic cells, the activity of the telomerase as an indicator for immortality has been determined. The studies were performed with the marine demosponges Suberites domuncula and Geodia cydonium, in vivo with tissue but also in vitro using the primmorph system. Primmorphs are formed from dissociated cells which have retained their proliferation potency. It was found that the activity of telomerase in tissue of both sponges is high. Based on this and additional findings it is assumed that the separation of the senescent sponge cell lineage from the immortal germ-/somatic cell lineage is triggered by the loss of contact to cell adhesion factors. First evidence is included which suggests that the final progress of the senescent, telomerase-negative cells to cell death is caused by apoptosis.

A high (2',5')oligoadenylate (2-5A) synthetase activity was found in the marine sponge Geodia cydonium. Here we demonstrate that the 2-5A synthetase activity is present also in other sponge species although the level of the 2-5A synthetase activity varies in several magnitudes in different sponges. The 2-5A synthesizing activity was maintained in the primary culture produced from a sponge.

Sponges (Porifera) represent the lowest metazoan phylum, characterized by a pronounced plasticity in the determination of cell lineages, and they are the closest related taxon to the hypothetical ancestral animal, the Urmetazoa, from which the metazoan lineages diverged. In a first approach to elucidate the molecular mechanisms controlling the switch from the cell lineage with a putative indefinite growth capacity to senescent, somatic cells, the activity of the telomerase as an indicator for immortality has been determined. The studies were performed with the marine demosponges Suberites domuncula and Geodia cydonium, in vivo with tissue but also in vitro using the primmorph system. Primmorphs are formed from dissociated cells which have retained their proliferation potency. It was found that the activity of telomerase in tissue of both sponges is high. Based on this and additional findings it is assumed that the separation of the senescent sponge cell lineage from the immortal germ-/somatic cell lineage is triggered by the loss of contact to cell adhesion factors. First evidence is included which suggests that the final progress of the senescent telomerase- negative cells to cell death is caused by apoptosis.

Among a sample of 39 Geodia cydonium (Demospongiae, Porifera) genes, with an average Gh+hC content of 51.2%, extensive structural heterogeneity and considerable variations in synonymous codon usage were found. The G + C content of coding sequences and G + C content at silent codon positions (GC3S) varied from 42.4 to 59.2% and from 35.6 to 76.5%, respectively. Correspondence analysis of 39 genes revealed that putative highly expressed genes preferentially use a limited subset of codons, which were therefore defined as preferred codons in G. cydonium. A total of 22 preferred codons for 18 amino acids with synonyms in codons were identified and they all (with one exception) end with C or G. Among these codons there are also C- and G-ending codons which were previously identified as codons optimal for translation in a variety of eukaryotes, including metazoans and plants. The bias in synonymous codon usage in putative highly expressed G. cydonium genes is moderate, indicating that these genes are not shaped under strong natural selection. We postulate that the preference for C- and G-ending codons was already established in the ancestor of all Metazoa, including also sponges. This ancestor most probably also had a G + C rich genome. The selection toward C- and G-ending codons has been largely conserved throughout eukaryote evolution; exceptions are, for example, mammals for which strong mutational biases caused switches from that rule. Der Codongebrauch im Kieselschwamm Geodia cydonium: Hochexprimierte Gene in den einfachsten multizellularen Tieren benutzen bevorzugt C- und G-endende Codons In einer Auswahl von 39 Genen des Kieselschwammes Geodia cydonium (Demospongiae, Porifera), die einen durchschnittlichen G + C Gehalt von 51.2% haben, wurde eine betrachtliche strukturelle Heterogenitat und eine hohe Variabilitat an synonymem Codongebrauch bestimmt. Der G + C Gehalt der codierenden Sequenzen sowie der G + C Gehalt an den neutralen Codon-Positionen (GC3S) variierte von 42.4–59.2%, beziehungsweise von 35.6–76.5%. Eine Vergleichsanalyse der 39 Gene ergab, das die wahrscheinlich hochexprimierten Gene bevorzugt nur eine begrenzte Auswahl an Codons benutzen, welche als ‘bevorzugte’ Codons von G. cydonium definiert werden. 22 ‘bevorzugte’ Codons fur die 18 Aminosauren, die synonyme Codons aufweisen, wurden untersucht; alle diese (mit einer Ausnahme) enden mit C oder G. Unter diesen Codons befinden sich auch diejenigen auf C und G-endenden Codons, die bereits in fruheren Untersuchungen als die bei der Translation von einer Vielzahl von Eukaryota, einschlieslich der vielzelligen Tiere und Pflanzen, bevorzugten Codons identifiziert wurden. Die Abweichung (Bias) des synonymen Codongebrauchs in den hochexprimierten G. cydonium Genen ist gering, was darauf hindeutet, das diese Gene nur unter einem geringen naturlichen Selektionsdruck stehen. Wir nehmen an, das der bevorzugte Gebrauch fur C-und G-endende Codons bereits in dem gemeinsamen Vorfahren der Metazoa, einschlieslich der Schwamme, etabliert war; es ist wahrscheinlich, das dieser gemeinsamen Vorfahre ein G + C-reiches Genom aufwies. Die bevorzugte Selektion C-und G-endender Codons blieb wahrend der Evolution der Eukaryota weitgehend konserviert; Ausnahmen sind die Saugetiere, die einen starken Bias aufweisen und deshalb von diesem Trend abweichen.

One autapomorphic character restricted to all Metazoa including Porifera [sponges] is the existence of transmembrane receptor tyrosine kinases (RTKs). In this study we screened for molecules from one subfamily within the superfamily of the insulin receptors. The subfamily includes the insulin receptors (InsR), the insulin-like growth factor I receptors, and the InsR-related receptors--all found in vertebrates--as well as the InsR-homolog from Drosophila melanogaster. cDNAs encoding putative InsRs were isolated from the hexactinellid sponge Aphrocallistes vastus, the demosponge Suberites domuncula, and the calcareous sponge Sycon raphanus. Phylogenetic analyses of the catalytic domains of the putative RTKs showed that the sponge polypeptides must be grouped with the InsRs. The relationships revealed that all sponge sequences fall into one branch of this group, whereas related sequences from mammals (human, mouse, and rat), insects and molluscs, and polypeptides from one cephalochordate, fall together into a second branch. We have concluded that (i) the InsR-like molecules evolved in sponges prior to the "Cambrian Explosion" and contributed to the rapid appearance of the higher metazoan phyla; (ii) the sponges constitute a monophyletic taxon, and (iii) epidermal growth factor (EGF)-like domains are present in sponges, which allows the insertion of this domain into potential receptor and matrix molecules.

Sponges (Porifera) are among the major phyla inhabiting the marine hard-substrate benthos, both in respect to the number of species and their biomass. Hence reliable biomarkers need to be developed to monitor the environmental load in those animals. Recently, the cDNA for the heat shock protein HSP70 has been isolated from the sponge Geodia cydonium and found to be a reliable indicator for temperature stress. In the present study, we have isolated the Rab GDP-dissociation inhibitor (GDI), which has previously been shown to be a key element in the intracellular traffic system. The 1521 bp long cDNA, encoding sponge GDI, has been isolated and analyzed. The deduced aa sequence ( M r = 50215) displays high homology to vertebrate, invertebrate and also yeast GDIs. Northern blot analysis revealed that the 1.6 kb sponge transcript that hybridized with the sponge GDI is down-regulated under temperature stress; in contrast, the transcription of the sponge HSP70 is strongly enhanced. Due to the high sequence conservation of the GDIs, one can assume that sponge GDI probes can be used as functional biomarkers for temperature stress, especially when applied simultaneously with the highly conserved HSP70 probes.

Geodia cidnium は古くから細胞凝集のモデルとして知られる海綿の一種である。この生物における細胞間相互作用には、凝集因子 (aggregation factor) と呼ばれる複合体の関与が知られていたが、最近、この複合体はある種のレクチンを介して細胞膜に存在する凝集受容体 (aggregation receptor) と結合していることがわかった。このレクチンはβガラクトシド結合性であり、そのcDNAを単離したところ推定アミノ酸配列からガレクチン家系と高い類似性を持つことが示された。ここに、系統発生上、最古の多細胞動物とされる海綿にすでにガレクチンが存在することが明らかになった。

GEODIA CYDONIUM (JAMESON, 1811) Records in the boreo-arctic region North-eastern Kara Sea: Off Sewernaja Semlja (Gorbunov, 1946: p. 37). South-western Barents Sea: Kola Fjord (Breitfuss, 1912: p. 62, as Cydonium mülleri; also referred to by Hentschel, 1929: p. 920, as Geodia mülleri). Norway: Off Vadsø, Varanger Fjord (Burton, 1930: p. 490, G. mülleri); Røberg, Trondheim Fjord (Arndt, 1913: p. 112, G. mülleri); Korsfjord near Bergen (Norman, 1879: p. 13, Geodia sp.; Brunchorst, 1891: p. 31, Geodia sp., according to Arndt, 1935: p. 30, both G. cydonium); off Haugesund (Schmidt, 1875: p. 120, Geodia gigas, according to Arndt, 1935: p. 30, G. cydonium); off Stavanger (Burton, 1930: p. 490, G. mülleri). Iceland: 64°56′N, 11°48′W, 216 m, 25.08.1902 (Burton, 1959: p. 9); Faxa Bay (Einarsson, 1941: p. 23, as G. mülleri). Discussion: Koltun (1966) reinvestigated the specimens of Gorbunov (1946) and Breitfuss (1912) and found that they are G. phlegraei. The specimen of Arndt (1913) could not be traced. A.B.K. and H.T.R. have sampled intensively on the same locality, Røberg in the Trondheimsfjord, and found many specimens of G. barretti and G. phlegraei, but not a single specimen referable to G. cydonium; note that the specimen of Arndt was probably about 15 cm in diameter. We conclude that Arndt’s specimen must have been misidentified. Arndt (1935) referred to some Geodia ‘sp.’s in the literature as G. cydonium. Nothing indicates that Arndt ever saw any of these specimens, rather he just felt certain that G. cydonium was an inhabitant of Norwegian waters. We have worked along most of the Norwegian coast and we have not found specimens that could be referred to G. cydonium. Probably all those referred to above represent G. barretti, which is very common along the entire Norwegian coastline. The Icelandic records are doubtful, too. We reinvestigated the specimen of Burton (1959) stored at ZMUC; in our opinion it is a fragment of G. barretti. Einarsson (1941) wrote ‘... enormous masses of sponges (G. mülleri ?) are encountered...’ Unfortunately we have no other samples from the same area, but everything considered, if it is a Geodia at all, it is presumably G. barretti. Our conclusion is that there are no certain records of G. cydonium north of the line Shetland Islands– Lousy Bank, west of the Faroe Islands. The last mentioned locality is listed by Burton (1959), and no description or further reference is given; accordingly, it may also be considered doubtful until a control is possible., Published as part of Cárdenas, Paco, Rapp, Hans Tore, Klitgaard, Anne Birgitte, Best, Megan, Thollesson, Mikael & Tendal, Ole Secher, 2013, Taxonomy, biogeography and DNA barcodes of Geodia species (Porifera, Demospongiae, Tetractinellida) in the Atlantic boreo-arctic region, pp. 251-311 in Zoological Journal of the Linnean Society 169 (2) on page 298, DOI: 10.1111/zoj.12056, http://zenodo.org/record/5287119, {"references":["Jameson R. 1811. Catalogue of animals of the class Vermes found in the Firth of Forth, and other parts of Scotland. Memoirs of the Wernerian Natural-History Society 1: 556 - 565.","Gorbunov GP. 1946. Benthonic populations of the novosibirsk shallow and central part of the Arctic Ocean. Tr. dreif. eksped. Glavsevmorputi na 1 / p ' G. Sedov' 1937 - 1940 III: 30 - 138 (in Russian).","Breitfuss JS. 1912. Zur Kenntnis der Spongio-Fauna des Kola Fjords. Travaux de la Societe Imperiale des Naturalistes de St. Petersbourg, Section Zoologie 41: 61 - 80, pls I - II.","Hentschel E. 1929. Die Kiesel- und Hornschwamme des Nordlichen Eismeers. In: Romer F, Schaudinn F, Brauer A, Arndt W, eds. Fauna Arctica. Eine Zusammenstellung der arktischen Tierformen mit besonderer Berucksichtigung des Spitzbergen-Gebietes auf Grund der Ergebnisse der Deutschen Expedition in das Nordliche Eismeer im Jahre 1898. Jena: G. Fischer, 857 - 1042, pls XII - XIV.","Burton M. 1930. Norwegian sponges from the Norman collection. Proceedings of the Zoological Society of London 1930: 487 - 546, pls I - II.","Arndt W. 1913. Zoologische Ergebnisse der ersten Lehr- Expedition der P. Scottlanderschen Jubilaums-Stifting. Jahresberichte der Schlesischen Gesellschaft fur vaterlandische Cultur 90: 110 - 119.","Norman AM. 1879. The Mollusca of the fjords near Bergen, Norway. Journal of Conchology 2: 8 - 77.","Brunchorst J. 1891. Die biologische Meeresstation in Bergen. Bergens Museums Aarsberetning for 1890 5: 1 - 31.","Arndt W. 1935. Porifera. III. a Die Tierwelt der Nord- und Ostsee. Leipzig. 1 - 140.","Schmidt O. 1875. Spongien Die Expedition zur physikalischchemischen und biologischen Untersuchung der Nordsee im Sommer 1872. Jahresbericht der Commission zur Wissenschaftlichen Untersuchung der Deutschen Meere in Kiel. 115 - 120, pl. I.","Burton M. 1959. Spongia. In: Fridriksson A, Tuxen SL, eds. The zoology of Iceland. Copenhagen: Ejnar Munksgaard, 1 - 71.","Einarsson H. 1941. Survey of the benthonic animal communities of Faxa Bay (Iceland). Meddelelser fra Kommissionen for Danmarks fiskeri- og HavundersOgelser. Serie: Fiskeri 11: 1 - 46.","Koltun VM. 1966. Four-rayed sponges of Northern and Far Eastern seas of the USSR (order Tetraxonida). Opredeliti Faunei SSSR 90. (Zoological Institute of the Academy of Sciences of the USSR: Moscow, Leningrad): 1 - 112, pls I - XXXVIII (in Russian)."]}

The DNA of the marine sponge Geodia cydonium (G.c.), a member of the phylogenetically old phylum Porifera, was characterized by density gradient centrifugation and by determining its genetic complexity by reassociation kinetics. At least five subcomponents were identified by curve-fit analyses of analytical density gradient centrifugation profiles of total G.c.-DNA. Four of these subcomponents were isolated from total G.c.-DNA by preparative density gradient centrifugation. The GC-contents of the subcomponents were determined to be 36.4%, 44.0%, 58.7%, and 66.1%, respectively. To our knowledge, such an extreme heterogeneity of DNA composition has never before been observed for any organism. The genetic complexities within the subcomponents were determined by reassociation kinetics to 2.1×108, 2.8×108, 9.2×108, and 1.4×109 bp, respectively. The orders of magnitude of the genetic complexities clearly indicate that the DNA subcomponents mainly contain eukaryotic single copy DNA, since DNA of symbiotic prokaryotes should show significantly lower complexities.

Four 4-ene-3,6-diketo steroids were isolated from the sponge G. cydonium along with 4-ene-3-keto steroids. The structure of these compds. was deduced by anal. of spectral data. Partial synthesis of compd. I confirmed the structure assignment of 4-ene-3,6-diketones

The genome of sponges has only been investigated so far by Bartmann-Lindholm et al. [Progr. Colloid. Polym. Sci. 107 (1997) 122-126] who reported a multimodal CsCl profile which could be resolved into five peaks for Geodia cydonium. This problem was reinvestigated here on both G. cydonium and Suberites domuncula. It was shown that DNAs from both sponges are characterized by unimodal CsCl profiles, additional peaks being due to contaminating prokaryotic and eukaryotic microorganisms.

In 1876, Campbell (Campbell, 1876 [p. 446]) wrote “those beautiful ‘glass-rope sponges’, Hyalonema etc., have been found by our researchers to be ‘the most characteristic inhabitants of the great depths all over the world, and with them ordinary siliceous sponges, some of which rival Hyalospongiae in beauty’ “. The admiration for the beauty of sponges is documented since Aristotle (cited in Camus 1783), however the nature of these organisms and their phylogenetic position remained enigmatic until less than 10 years ago. E.g., in 1988 Loomis (Loomis, 1988 [p. 186]) wrote “the sponge cells are unspecialized flagellates held together by a glycoprotein extracellular matrix... they are multicellular, but just barely so”. This view changed drastically since the introduction of modern molecular biological techniques; the informational genes investigated in detail now group the sponges to the Metazoa leaving any doubt on their evolutionary origin behind. The first breakthrough came with the study of the galectin molecule from Geodia cydonium, when it was discovered that the deduced polypeptide shared high sequence similarity only with metazoan proteins and comprised all characteristic amino acid moieties required for the binding to the sugar (Pfeifer et al., 1993).

Sponges (Porifera) are the phylogenetically oldest still extant metazoan phylum. Recently elements of their immune system have been cloned and analyzed, primarily from the demosponges Suberites domuncula and Geodia cydonium. By differential display, two genes were identified in S. domuncula, whose translation products are involved in graft rejection/fusion: the allograft inflammatory factor (AIF-1) and the Tcf-like transcription factor (TCF). Since the AIF-1 and TCF genes are upregulated in vivo after tissue transplantation, especially in allografts, we investigated whether this reaction can be monitored in vitro. Therefore, the autogeneic and the allogeneic mixed sponge cell reaction (MSCR) system was applied for the first time to identify distinct factors in sponges in vitro. The results confirm that the two AIF-1 and TCF genes are induced during allogeneic MSCR. Furthermore, the recombinant sponge AIF-1 causes an upregulation of the expression of the TCF. We conclude that the AIF-1 and TCF genes are upregulated in sponges during histoincompatibility reactions ; the data support the view that sponges have immune systems composed of highly complex elements related to those found in mammalian systems.

Polychaete assemblages associated to the sponge Geodia cydonium were investigated at two sampling sites in the Mediterranean Sea: Porto Cesareo Basin (Apulia) and Marsala Lagoon (Sicily), both characterized by sheltered hydrodynamic conditions. Samples were seasonally performed during 1997, in order to compare the assemblages coming from the two localities studied, considering separately the internal and external tissues of the sponge, and with the aim of evaluating the influence of sponge size on polychaete colonization. The examined sponge is characterized by a peculiar stratification of its tissues: an external thick and hard layer, the cortex, and an internal softer one, the choanosome. Statistical analysis showed that this was the main factor controlling polychaete assemblage, with the internal tissue, less rich and diversified, appearing impoverished with respect to the external layer. A similarity in species composition was observed between sites, even though some differences were evidenced in the abundance of some species, mainly reflecting differences in local environmental conditions. Species richness and density increased with the increasing sponge size. Such a situation is particularly evident at Porto Cesareo, where sponges are covered by an algal layer which is particularly rich on the largest specimens, thus suggesting that most of the species of polychaetes were linked more to the neighbouring environment than to the sponge itself.

Among Metazoa, sponges (phylum Porifera) are the richest source for different bioactive compounds. The availability of the raw material is, however, restricted. To obtain enough of the bioactive compounds for application in human therapy, sponges have to be cultured in in vitro systems. One technique for the establishment of a long-term cell culture from sponges has recently been elaborated. Here, we present a procedure to cultivate tissue samples from sponges in an open system. The species Geodia cydonium, which produces bioactive compounds, has been selected. Tissue samples of approximately 10 g were attached to the bottoms of cultivation trays. After 2 to 3 days, the tissue samples formed a robust contact with the metal support. Subsequently, sets of trays, called tray batteries, either remained in huge aquaria at the Center for Marine Research or were transferred to the vicinity of a fish and mussel farm. The growth rates of the samples remained unchanged within the first month; however, after 3 and 6 months, they increased to 147% and 189%, respectively. In parallel, extracts were prepared from the tissue samples and tested for cytotoxicity in a mouse lymphoma cell assay system. Extracts from cultured tissue initially had a low inhibitory potency; however, after cultivation for 3 or 6 months, values comparable to those of extracts from sponges taken from the biotope were found. In addition, a molecular marker was applied to document. the response (health state) of the tissue and the identity of the material in culture. The CD63 molecule was chosen because the expression of this molecule in mammalian systems changes with the age of the animals. The corresponding complementary DNA was isolated from Geodia cydonium. With this probe, the level of expression in cultured tissue samples decreased immediately after starting cultivation; after a cultivation period of 6 months, however, values were similar to those found in controls. These data show that sponge species that produce bioactive compounds can be cultivated in open systems, in which they retain their potency to produce bioactive compounds as well as their health state.

Recently cDNAs coding for cell surface molecules have been isolated from sponges. The molecules for alpha-integrin, galectin, and receptor tyrosine kinase (RTK), obtained from the marine sponge, Geodia cydonium?, have been described earlier, In the present study also the cDNA for one putative beta-integrin has been identified from G, cydonium. The deduced aa sequence comprises the characteristic signatures, found in other metazoan beta-integrin molecules; the estimated size is 95,215 Da, To obtain first insights into the molecular events which proceed during autograft fusion, the expressions of these genes were determined on transcriptional and translational level. The cDNAs as well as antibodies raised against the recombinant sponge proteins alpha-integrin, RTK and galectin were used and Northern blot experiments and immunocytochemical analyses have been performed. The results show that transcription of the two subunits of an integrin receptor as well as of the RTK are strongly upregulated after grafting; levels of > 10-fold have been determined in the fusion zone of the grafts after a 10 days incubation. Immunofluorescence studies of sections through the fusion zone support these data, In contrast the transcription of the gene encoding galectin is drastically downregulated after grafting, In a parallel series of experiments the level of the heat-shock protein-70 was determined and it was found that it remained unchanged after grafting, We conclude that integrin subunits and the RTK molecule are involved in self-self recognition of sponge.

Polychlorinated biphenyls (PCBs) are ubiquitous industrial compounds found in almost every component of the terrestrial and marine ecosystem. Most of the PCB congeners bind to the aryl hydrocarbon receptor and in turn cause expression of stress response genes. here we report for the first time that PCB 118 acts in marine sponge Geodia cydonium as an inducer of 2 chaperons, the 14-3-3 protein(s)(a protein targeting molecule) and the heat shock protein HSP70 (a chaperone, primarily involved in folding of proteins). While the cDNA encoding the latter protein has been cloned previously, the 14-3-3 cDNA from sponges is reported in this study. The full-length cDNA clone of G.cydonium, GC14-3-3, has a size of 912 nucleotides (nt) and contains a 744 nt long potential open reading frame ; the relative molecular weight (Mr) of the deduced aa sequence is 28 378 Da. The sponge polypeptide is closely related to the deduced polypeptides of the 14-3-3 sequences belongig to isoforms ni and gama. Using the cDNAs, coding for the 14-3-3 and the HSP70 proteins as well as antibodies raised against these 2 proteins, it was demonstrated that neither chaperon can be detected in the absence of PCB. However, after incubation of sponge tissue with PCB 118 the transcripts of the 2 chaperones are detectable after 12h, while the corresponding proteins appear after 1d. Subsequently, the levels of the transcripts and of the proteins increase steadily. From these data we conclude that the 2 chaperones, 14-3-3 and HSP70, are useful biomarkers in sponges. Due to the broad cross-reactivity of their antibodies throughout the Metazoa, these chaperones may be useful biomarkers for monitoring environmental contaminants, as shown here for PCB 118, in all organisms.

The marine demosponge Geodia cydonium encodes a gene, termed GCXPB, which displays 62% identity to the human XPB/ERCC-3 gene. The degree of DNA strand breaks induced by UVB light (300 or 1000 Jm-2)in the marine sponge paralleled the increase of expression of the GCXPB gene.

Because sponges (Porifera) are generally the dominant group of multicellular animals found in the marine benthos they are key animals for the testing of biomarkers in this biotope. Previously, it was shown that the heat shock protein hsp70 from the marine sponge Geodia cydonium is a useful biomarker for following biological responses to selected physical and chemical stressors. One protein which is critical for hsp70 function in other organisms is the DnaJ-like protein. In this study we therefore isolated and characterized a cDNA encoding the DnaJ-like protein from G. cydonium designated GCDNAJ. GCDNAJ is 1476 base pairs (bp) long with a 1236 bp open reading frame encoding a protein of 45 542 daltons. Northern blot analysis revealed a transcript size of 1.5 kilobase (kb). The deduce amino acid (aa) sequence of GCDNAJ contains motifs characteristic of most prokaryotic and eukaryotic DnaJ family members. In response to a 7oC temperature increase, expression of the transcripts encoding hsp70 and DnaJ-like protein increased as much as 18-fold and 3-fold, respectively, within a 8 h period. From these data we concluded that the levels of both hsp70 and DnaJ-like protein are likely to be important to the adaptation to temperature increases in G. cydonium.

Sponges [Porifera] are the phylogenetically oldest phylum of the Metazoa. They are provided with both cellular and humoral allorecognition systems. The underlying molecules are not yet known. To study allorecognition in sponges we first determined the frequency of graft rejection in a natural population of the marine sponge Geodia cydonium. We then determined, for the first time at the molecular level, the degree of sequence polymorphism in segments of one molecule which may be related to sponge allorecognition and host defense: the Ig-like domains from the receptor tyrosine kinase [RTK]. Thirty six pairs of auto- and allografts were assayed, either by parabiotic attachment or insertion of grafts. All of the autografts fused, while only two allografts fused and 34 pairs were incompatible. Rejection among the parabiotic allografts was characterized by the formation of a collagenous barrier, while the allografts that were inserted into the host underwent destruction. At the molecular level we first cloned to completion the 5'-end of sponge RTK, which displays a Pro-Ser-Thr-rich sequence; this is thought to act as a module of cell adhesion proteins. Then we analyzed RT-PCR products of amplification across the two Ig-like domains of RTK (about 500 bp), from two pairs of fusing sponges and one pair of rejecting sponges. High levels of polymorphism were recorded, including 18 nucleotide-substitution positions and a tri-nucleotide deletion, which translate into 13 polymorphic amino acid positions. Two of the six sponges were scored as heterozygotes. Among 9 informative polymorphic sites that were tested for linkage disequilibrium, 11 pairwise comparisons were found to be significant, implying the possibility of distinguishable alleles in this locus. To the best of our knowledge this is the first report of polymorphism in Ig-like domains of a receptor from invertebrates that may be associated with allorecognition. This data attests also that fusion in sponges is not confined to genetically identical individuals.

The stress proteins, the "classic" heat shock proteins, have a high potential as biomarkers for general sterss. In the present study it is shown that this heat shock response is a fundamental aspect of cellular physiology also in freshwater- and marine sponges, in which exposure to temperature stress or xenobiotics result in a dramatic reaction, induction of hsps (especially hsp70) and a suppression of other cellular proteins. With the isolation of the cDNA, coding for hsp70 in the marine sponge Geodia cydonium, a tool is now available to analyze for the first time the reaction of a sponge to stressors on molecular level.

Protein kinases C (PKCs) comprise closely related Ser/Thr kinases, ubiquitously present in animal tissues ; they respond to second messengers, e.g., Ca2+ and/or diacylglycerol, to express their activities. Two PKCs have been sequenced from Geodia cydonium, a member of the lowest multicellular animals, the sponges (Porifera). One sponge G. cydonium PKC, GCPKC1, belongs to the ''novel'' (Ca2+-independent) PKC (nPKC) subfamily while the second one, GCPKC2, has the hall-marks of the ''conventional'' (Ca2+-dependent) PKC (cPKC) subfamily. The alignment of the Ser/Thr catalytic kinase domains, of the predicted aa sequences for these cDNAs with respective segments from previously reported sequences, revealed highest homology to PKCs from animals but also distant relationships to Ser/Thr kinases from protozoa, plants, and bacteria. However, a comparison of the complete structures of the sponge PKCs, which are-already-identical to those of nPKCs and cPKCs from higher metazoa, with the structures of protozoan, plant, and bacterial Ser/Thr kinases indicates that the metazoan PKCs have to be distinguished from the nonmetazoan enzymes. These data indicate that metazoan PKCs have a universal common ancestor which they share with the nonmetazoan Ser/Thr kinases with respect to the kinase domain, but they differ from them in overall structural composition.

The biomarker concept involves the use of biochemical, cellular and physiological parameters as screening tools in environmental surveillance. Stress proteins, such as heat shock proteins (HSPs), fulfill many of the requirements for being ideal candidates in a biomarker strategy for environmental monitoring. Sponges (Porifera) are one of the major phyla found in the marine hard-substrate benthos, both with respect to the number of species and biomass. However, only recently have genes from sponges been cloned. Here we describe the isolation of the cDNA encoding a heat shock protein of M(r) (relative molecular weight) 70 kDa (HSP70). The cDNA HSP70 from the marine sponge Geodia cydonium has a length of 2.3 kb and encodes an AA sequence of M(r) 72 579. The sponge HSP70 displays characteristic features of the HSP70 family. The HSP70 protein is induced by natural stressors including changes in temperature as well as pH, as demonstrated by Western blot analysis. No response was observed after treating the samples with hypotonic or hypertonic conditions. Our results provide the first molecular evidence that HSP70 of sponges is a useful biomarker.

During the development of higher animals, morphogenetic programs are switched on which are frequently controlled by homeotic genes. Until now these genes have not been identified in the lowest animals, the marine sponges. Since sponges show (i) an antero-posterior and/or dorso-ventral axis during embryogenesis and (ii) a complex differentiation pattern during spicula formation, we hypothesized that in sponges homeotic genes — if present — are also involved in the control of these processes. Therefore, we searched for homeobox or homeobox-like sequences in the marine sponge Geodia cydonium . Here we describe a homeobox-like sequence from these animals; it was isolated from a cDNA library of an adult specimen. The deduced amino acid sequence of the complete homeodomain shares over 70% similarity with other homeodomain sequences, including those from hydra, insects and vertebrates. These data indicate that the sponge homeodomain-like sequence is similar with respect to structure to those of other animals and may suggest that the sponge homeodomain-like sequence(s) might function during developmental processes and/or during spiculogenesis in a similar manner to that known for higher animals.

We have isolated circular proteid particles (CPP) from the sponge Geodia cydonium , which carry the aggregation factor, and have freed them from concomitant impurities. The CPP have been visualized by electron microscopy as annular particles with a circular contour length of 3530 A. The circles are surrounded by 25 filaments 610 A in length. The CPP are characterized by a sedimentation coefficient ( S ° 20, w ) of 3000, by a buoyant density of 1.32 g/cm 3 and by a particle weight of about 2–5 · 10 9 daltons. They consist primarily of protein. It is suggested that the fibrous complex found electron microscopically is only the core structure of a more complex particle.

In the extracellular space of the tissue of the sponge Geodia cydonium, circular proteid particles are found which carry as subunits the aggregation factor and a series of glycosyltransferases. Using the technique of velocity sucrose gradient centrifugation, the sedimentation coefficient ( s 20, w 0 ) of the particle-monosomes was determined to be 90. By means of the Svedberg equation a molecular weight of 1.3 · 108 daltons could be estimated. The monosomes aggregate in the presence of Ca2+ to higher complexes via disomes, trisomes, and pentasomes. The complexes can be redissociated by dodecyl sulfate but not by EDTA. During the Ca2+-mediated self-aggregation, the particles lose their biological activity with respect to their aggregation promoting function.

The siliceous sponge Geodia cydonium was induced to regenerate. In this condition the nucleic acid fractions were used as indicators to study the influence of pollution in marine environments. Besides conventional analytical methods a newly developed method is introduced. The cetyltrimethylammonium salts of the polyanions are formed which are soluble at high ionic strength and are precipitated sequentionally as measurable turbidities by dilution. The dilution turbidity curves of regenerating sponge in different environments aided with polyribosomal profiles can be interpreted. It is shown that under the conditions of intermittent pollution the amount of some high molecular RNA fractions decrease. The turbidity-dilution technique may become a useful tool in environmental investigations.

An aggregation receptor (AR) from the siliceous sponge Suberites domuncula has been isolated and purified by chromatography to about 55% purity. The AR consists primarily of neutral carbohydrate and is characterized by a buoyant density of 1.59 g/ml and by an apparent molecular weight of 42,500. The average density of the AR on Suberite cells is about 3.8 × 10 5 per μm 2 . The AR contains considerable amounts of hexuronic acid. The isolated AR can bind not only to receptor-depleted Suberites cells but also to receptor depleted cells from another siliceous species (Geodia cydontum) . After being charged with Suberites ARs, Geodia cells form aggregates in the presence of the species-specific Suberites aggregation factor.

Chemisch dissoziierte Zellen des KieselschwammesGeodia cydonium reaggregieren aufgrund zweier verschiedenr Reaggregationsprinzipien. Der Aggnegationsfaktor, auf den die Primaraggregation zuruckgeht, ist membrangebunden und wird durch Proteasen nicht inaktiviert. Der sekundare Aggregationsfaktor wurde 500fach angereichert. Das Molekulargewicht dieses Aggregationsfaktors betragt etwa 20000 Daltons; er ist mit einem ringformigen Makromolekul (2×109 Daltons) assoziiert.

In vertebrates cytokines mediate innate (natural) immunity and protect them against viral infections. The cytokine interferon causes the induction of the (2′-5′)oligoadenylate synthetase [(2-5)A synthetase], whose product, (2′-5′)oligoadenylate, activates the endoribonuclease L which in turn degrades (viral) RNA. Three isoforms of (2-5)A synthetases exist, form I (40–46 kDa), form II (69 kDa), and form III (100 kDa). Until now (2-5)A synthetases have only been cloned from birds and mammals. Here we describe the cloning of the first putative invertebrate (2-5)A synthetase from the marine sponge Geodia cydonium. The deduced amino acid sequence shows signatures characteristic for (2-5)A synthetases of form I. Phylogenetic analysis of the putative sponge (2-5)A synthetase indicates that it diverged first from a common ancestor of the hitherto known members of (vertebrate) (2-5)A synthetases I, (2-5)A synthetases II and III. Moreover, it is suggested that the (2-5)A synthetases II and III evolved from this common ancestor (very likely) by gene duplication. Together with earlier results on the existence of the (2′-5′)oligoadenylates in G. cydonium, the data presented here demonstrate that also invertebrates, here sponges, are provided with the (2-5)A system. At present, it is assumed that this system might be involved in growth control, including control of apoptosis, and acquired its additional function in innate immune response in evolutionarily younger animals, in vertebrates.

Recently, the presence of 2′,5′-linked oligoadenylates and a high 2′,5′-oligoadenylate synthetase activity were discovered in a lower invertebrate, the marine sponge Geodia cydonium. It has been demonstrated that mammalian 2–5A synthetase isozymes require a dsRNA cofactor for their enzymatic activity. Our results show that, unlike mammalian 2–5A synthetases, the 2–5A synthetase from the sponge acts in a dsRNA-independent manner in vitro. A prolonged incubation of the G. cydonium extract with a high concentration of a micrococcal nuclease had no effect on the activity of the 2–5A synthetase. At the same time, the micrococcal nuclease was effective within 30 min in degrading dsRNA needed for the enzymatic activity in IFN-induced PC12 cells. These results indicate that the 2–5A synthetase from G. cydonium may be active per se or is activated by some other mechanism. The sponge enzyme is capable of synthesizing a series of 2–5A oligomers ranging from dimers to octamers. The accumulation of a dimer in the predominant proportion during the first stage of the reaction was observed, followed by a gradual increase in longer oligoadenylates. By its product profile and kinetics of formation, the sponge 2–5A synthetase behaves like a specific isoform of enzymes of the 2–5A synthetase family.

1. 1. Ergosta-4,24(28)-dien-3-one (1) is found in Geodia cydonium and other Astrophorida demosponges, as well as in subantarctic shallow-water hexactinellides. 2. 2. In accordance with previous views, derived from the similar composition of fatty acids for both Demospongia and Hexactinellida, this provides no biochemical ground in support of the separation of Hexactinellida from the class Demospongia.