The sterols of calcareous sponges (Calcarea, Porifera)

Sponges are sessile suspension-feeding organisms whose internal phylogenetic relationships are still the subject of intense debate. Sterols may have the potential to be used as independent markers to test phylogenetic hypotheses. Twenty representative specimens of calcareous sponges (class Calcarea, phylum Porifera) with a broad coverage within both subclasses Calcinea and Calcaronea were analysed for their sterol content. Two major pseudohomologous series were found, accompanied by some additional sterols. The first series encompassing conventional C(27) to C(29)Delta(5,7,22) sterols represented the major sterols, with ergosterol (ergosta-5,7,22-trien-3beta-ol, C(28)Delta(5,7,22)) being most prominent in many species. The second series consisted of unusual C(27) to C(29)Delta(5,7,9(11),22) sterols. Cholesterol occurred sporadically, mostly in trace amounts. The sterol patterns did not resolve intraclass phylogenetic relationships, namely the distinction between the subclasses, Calcinea and Calcaronea. This pointed towards major calcarean lipid traits being established prior to the separation of subclasses. Furthermore, calcarean sterol patterns clearly differ from those found in Hexactinellida, whereas partial overlap occurred with some Demospongiae. Hence, sterols only partly reflect the phylogenetic separation of Calcarea from both of the other poriferan classes that was proposed by recent molecular work and fatty acid analyses.     

The fatty acids of calcareous sponges (Calcarea, Porifera)

Twenty-nine specimens of calcareous sponges (Class Calcarea, Phylum Porifera), covering thirteen representative species of the families Soleneiscidae, Leucaltidae, Levinellidae, Leucettidae, Clathrinidae, Sycettidae, Grantiidae, Jenkinidae, and Heteropiidae were analysed for their fatty acids. The fatty acids of Calcarea generally comprise saturated and monounsaturated linear (n-), and terminally methylated (iso-, anteiso-) C(14)-C(20) homologues. Furthermore, polyunsaturated C(22) fatty acids and the isoprenoic 4,8,12-trimethyltridecanoic acid were found. The most prominent compounds are n-C(16), iso-C(17), iso-C(18), n-C(18), n-C(20). In addition, a high abundance of the exotic 16-methyloctadecanoic acid (anteiso-C(19)) appears to be a characteristic trait of Calcarea. Long-chain 'demospongic acids', typically found in Demospongiae and Hexactinellida, are absent in Calcarea. The completely different strategy of calcarean fatty acid synthesis supports their phylogenetic distinctiveness from a common Demospongiae/Hexactinellida taxon. Both intraspecific and intraclass patterns of Calcarea showed great similarity, suggesting a conserved fatty acid composition that already existed in the last common ancestor of Calcinea and Calcaronea, i.e. before subclasses diverged.     

Molecular phylogenetic evaluation of classification and scenarios of character evolution in calcareous sponges (Porifera, Class Calcarea)

Calcareous sponges (Phylum Porifera, Class Calcarea) are known to be taxonomically difficult. Previous molecular studies have revealed many discrepancies between classically recognized taxa and the observed relationships at the order, family and genus levels; these inconsistencies question underlying hypotheses regarding the evolution of certain morphological characters. Therefore, we extended the available taxa and character set by sequencing the complete small subunit (SSU) rDNA and the almost complete large subunit (LSU) rDNA of additional key species and complemented this dataset by substantially increasing the length of available LSU sequences. Phylogenetic analyses provided new hypotheses about the relationships of Calcarea and about the evolution of certain morphological characters. We tested our phylogeny against competing phylogenetic hypotheses presented by previous classification systems. Our data reject the current order-level classification by again finding non-monophyletic Leucosolenida, Clathrinida and Murrayonida. In the subclass Calcinea, we recovered a clade that includes all species with a cortex, which is largely consistent with the previously proposed order Leucettida. Other orders that had been rejected in the current system were not found, but could not be rejected in our tests either. We found several additional families and genera polyphyletic: the families Leucascidae and Leucaltidae and the genus Leucetta in Calcinea, and in Calcaronea the family Amphoriscidae and the genus Ute. Our phylogeny also provided support for the vaguely suspected close relationship of several members of Grantiidae with giantortical diactines to members of Heteropiidae. Similarly, our analyses revealed several unexpected affinities, such as a sister group relationship between Leucettusa (Leucaltidae) and Leucettidae and between Leucascandra (Jenkinidae) and Sycon carteri (Sycettidae). According to our results, the taxonomy of Calcarea is in desperate need of a thorough revision, which cannot be achieved by considering morphology alone or relying on a taxon sampling based on the current classification below the subclass level.     

Non-monophyly of most supraspecific taxa of calcareous sponges (Porifera, Calcarea) revealed by increased taxon sampling and partitioned Bayesian analysis of ribosomal DNA

Calcareous sponges (Porifera, Calcarea) play an important role for our understanding of early metazoan evolution, since several molecular studies suggested their closer relationship to Eumetazoa than to the other two sponge 'classes,' Demospongiae and Hexactinellida. The division of Calcarea into the subtaxa Calcinea and Calcaronea is well established by now, but their internal relationships remain largely unresolved. Here, we estimate phylogenetic relationships within Calcarea in a Bayesian framework, using full-length 18S and partial 28S ribosomal DNA sequences. Both genes were analyzed separately and in combination and were further partitioned by stem and loop regions, the former being modelled to take non-independence of paired sites into account. By substantially increasing taxon sampling, we show that most of the traditionally recognized supraspecific taxa within Calcinea and Calcaronea are not monophyletic, challenging the existing classification system, while monophyly of Calcinea and Calcaronea is again highly supported.     

Molecular systematics of sponges (Porifera)

The first application of molecular systematics to sponges was in the 1980s, using allozyme divergence to dis-criminate between conspecific and congeneric sponge populations. Since this time, a fairly large database has been accumulated and, although the first findings seemed to indicate that sponge species were genetically more divergent than those of other marine invertebrates, a recent review of the available dataset indicates that levels of interspecific gene identities in most sponges fall within the normal range found between species of other invertebrates. Nevertheless, some sponge genera have species that are extremely divergent from each other, suggesting a possible polyphyly of these genera. In the 1990s, molecular studies comparing sequences of ribosomal RNA have been used to reappraise the phylogenetic relationships among sponge genera, families, orders and classes. Both the 18S small subunit and the 28S large subunit rRNA genes have been sequenced (41 complete or partial and 75 partial sequences, respectively). Sequences of 18S rRNA show good support for Porifera being true Metazoa, but they are not informative for resolving relationships among genera, families or orders. 28S rRNA domains D1 and D2 appear to be more informative for the terminal nodes and provide resolution for internal topologies in sufficiently closely related species, but the deep nodes between orders or classes cannot be resolved using this molecule. Recently, a more conserved gene, Hsp70, has been used to try to resolve the relationships in the deep nodes. Metazoan monophyly is very well supported. Nevertheless, the divergence between the three classes of Porifera, as well as the divergence between Porifera, Cnidaria and Ctenophora, is not resolved. Research is in progress using other genes such as those of the homeodomain, the tyrosine kinase domain, and those coding for the aggregation factor. For the moment the dataset for these genes is too restricted to resolve the phylogenetic relationships of these phyla. However, whichever the genes, the phylogenies obtained suggest that Porifera could be paraphyletic and that the phylogenetic relationships of most of the families and orders of the Demospongiae have to be reassessed. The Calcarea and Hexactinellida are still to be studied at the molecular level.     

Revision of the genus Clathrina (Porifera, Calcarea)

The genus Clathrina has one of the most difficult systematic arrangements in the Porifera, Class Calcarea. Few morphological characters can be used to describe its species, and the systematics and the geographical distribution of its species have changed several times, according to the point of view of the systematists. 'Lumpers' consider that clathrinas are morphologically plastic, while 'splitters' believe that even slight morphological differences should be considered sufficient to distinguish between species. The morphology of several specimens/species of Clathrina , including the type species, when possible, was studied and used to produce the first revision of this genus. Using results obtained from previous morphological and molecular studies, the morphological characters were chosen and analysed in all studied specimens. In total, 43 species were found, nine of which are new to science. These results agree with the viewpoint that morphological characters such as type and distribution of spicules, size of actines, spines, anastomosis of the cormus, organization of the osculum and presence of granules in cells, are useful when establishing the taxonomy of the genus.  © 2003 The Linnean Society of London, Zoological Journal of the Linnean Society , 2003, 139 , 1−62.     

Oogenesis and spermatogenesis in Paraleucilla magna (Porifera, Calcarea)

Gametogenesis in Calcarea is still poorly known; therefore, in this work, we describe the gametogenesis of Paraleucilla magna (Porifera, Calcarea) using light and electron microscopy. Male and female gametes derived from choanocytes. Oocytes grew inside a follicle-like structure composed of pinacocyte-like cells and choanocytes. They were nourished through nutrient transfer from nurse cells and through bacteria phagocytosis, which was never documented for a calcarean species. Mature oocytes were typical, spherical, with nucleolated nuclei and a cytoplasm filled with different types of inclusions and digestive vesicles. Nuage was also found. Spermatogenesis occurred in the lumen of differentiated choanocyte chambers, and spermatic cysts were never formed. Spherical spermatogonia without flagellum or collar were derived from hourglass-shaped primordial germ cells. Spermatozoa were of the ‘aberrant type’, showed a compact electron-dense nucleus and a homogeneous granulated body in its cytoplasm. The gametogenesis of P. magna is similar to what has been observed for other Porifera and Eumetazoa.     

The stem cell concept in sponges (Porifera): Metazoan traits

Sponges are considered the oldest living animal group and provide important insights into the earliest evolutionary processes in the Metazoa. This paper reviews the evidence that sponge stem cells have essential roles in cellular specialization, embryogenesis and Bauplan formation. Data indicate that sponge archaeocytes not only represent germ cells but also totipotent stem cells. Marker genes have been identified which are expressed in totipotent stem cells and gemmule cells. Furthermore, genes are described for the three main cell lineages in sponge, which share a common origin from archaeocytes and result in the differentiation of skeletal, epithelial, and contractile cells.     

Oogenesis and larval development of Scypha ciliata (Porifera,Calcarea)

Summary Scypha ciliata is a syconoid sponge. Its oocytes differentiate from choanocytes located near the apopyle of a flagellated chamber, and initially they remain in that location, in a trophic complex with neighbouring choanocytes. When this first growth phase is completed, the oocyte migrates to the periphery of the sponge. There it undergoes a second growth phase, in which it phagocytizes choanocytes and mesenchyme cells.Fertilization of the mature egg is assisted by a converted choanocyte, the sperm carrier cell. This cell penetrates the oocyte and transfers to it the sperm contained in a carriercell vacuole. No meiotic events have yet been observed.Cleavage is asynchronous, with holoblastic, approximately equal divisions. After the first cleavage steps the blastomeres often contain multiple nuclei. The single-layered blastoderm of the stomoblastula consists of many micromeres with flagella that project into the blastocoel, a few macromeres and four cruciform cells. There is no development of a follicle epithelium.The stomoblastula develops into the amphiblastula by inversion; with the assistance of the maternal choanocyte epithelium, the hollow sphere turns inside out, simultaneously moving out of the mesoderm and into the lumen of the adjacent flagellated chamber. In this process, the blastocoel of the stomoblastula is lost. The flagellated cells that form the wall of the amphiblastula now have their flagella extending outward; the amphiblastula also comprises four cruciform cells, macrogranular and agranular cells. The larval cavity of the amphiblastula is a newly formed structure.Abbreviations AB amphiblastula - AP apopyle - BC blastocoel - aC agranular cell - maC macrogranular cell - miC microgranular cell - CB crystalline body - CC central cavity - Ch choanocyte - fCh flat choanocyte - gCh granulate choanocyte - CM cell membrane - Co collar of choanocyte - CrC cruciform cell - DM dense material - EM electron micrograph - F flagellum - FC flagellated cell - FCm flagellated chamber - FL free larva - FV food vacuole - IR interior region - LC larval cavity - M mesenchyme - Ma macromere - MC mesenchyme cell - Mi micromere - N nucleus - Nu nucleolus - O opening - OC oocyte - P psudopodium - PC pinacocyte - PhM phase-contrast micrograph - Po pore - PP prosopyle - S sperm - SB stomoblastula - SC segmentation cavity - SCC sperm-carrier cell - SV sperm vacuole - lT large trophocyte - sT small trophocyte - V vacuole - VC vesicular cytoplasm - VM vacuole membrane     

The choanoderm of Sycettusa hastifera (Calcarea,Porifera) is able to generate new individuals

One of the main characteristics of sponges is their capacity for cell dedifferentiation. This capability can allow an impressive amount of asexual reproduction in these animals, because they are able to develop new individuals from just a few somatic cells. Studies of dedifferentiation, however, have focused mainly on sponges of the class Demospongiae. Therefore, we investigated here whether individuals of three different species of Calcarea are able to reconstitute new individuals following artificial fragmentation. We observed that fragmentation releases clumps of choanoderm able to initiate somatic embryogenesis. In Borojevia brasiliensis (asconoid aquiferous system, subclass Calcinea) and Paraleucilla magna (leuconoid aquiferous system, subclass Calcaronea), these clumps started to develop, but they did not pass through the first developmental phases. In Sycettusa hastifera (syconoid aquiferous system, subclass Calcaronea), the choanoderm was reorganized into primmorphs that fused to each other and formed an exopinacoderm. The first primmorphs’ spicules were triactines. Despite a large mortality rate, the primmorphs developed into olynthus stages. The somatic embryogenesis and the metamorphosis of the olynthus were similar to those observed during the sexual development of this and other calcareous sponge species. Our results show that in S. hastifera, and perhaps in other syconoid calcareous sponges, somatic embryogenesis occurs mainly from choanocytes, at least in vitro. However, primmorph development does not follow the same pattern observed in post‐metamorphic sexual development, as in that case diactines are always the first spicules to be synthesized in calcaronean species.     

First report on chitinous holdfast in sponges (Porifera)

A holdfast is a root- or basal plate-like structure of principal importance that anchors aquatic sessile organisms, including sponges, to hard substrates. There is to date little information about the nature and origin of sponges’ holdfasts in both marine and freshwater environments. This work, to our knowledge, demonstrates for the first time that chitin is an important structural component within holdfasts of the endemic freshwater demosponge Lubomirskia baicalensis. Using a variety of techniques (near-edge X-ray absorption fine structure, Raman, electrospray ionization mas spectrometry, Morgan–Elson assay and Calcofluor White staining), we show that chitin from the sponge holdfast is much closer to α-chitin than to β-chitin. Most of the three-dimensional fibrous skeleton of this sponge consists of spicule-containing proteinaceous spongin. Intriguingly, the chitinous holdfast is not spongin-based, and is ontogenetically the oldest part of the sponge body. Sequencing revealed the presence of four previously undescribed genes encoding chitin synthases in the L. baicalensis sponge. This discovery of chitin within freshwater sponge holdfasts highlights the novel and specific functions of this biopolymer within these ancient sessile invertebrates.     

First evidence of miniature transposable elements in sponges (Porifera)

Transposable elements play a vital role in genome evolution and may have been important for the formation of the early metazoan genome, but only little is known about transposons at this interface between unicellular opisthokonts and Metazoa. Here, we describe the first miniature transposable elements (MITEs, Queen1 and Queen2) in sponges. Queen1 and Queen2 are probably derived from Tc1/mariner-like MITE families and are represented in more than 3,800 and 1,700 copies, respectively, in the Amphimedon queenslandica genome. Queen elements are located in intergenic regions as well as in introns, providing the potential to induce new splicing sites and termination signals in the genes. Further possible impacts of MITEs on the evolution of the metazoan genome are discussed.     

Condensation rhythm of fresh-water sponges (Spongillidae, Porifera)

The mesenchyme continuum of spongillids exhibits rhythmic changes that at first glance appear to be contractions. Actually, however, the process is a condensation initiated by the formation of punctate cell contacts and a rapid swelling of all mesenchymal cells. As the cells come into closer contact and the spaces between them are constricted, the volume of the mesenchyme shrinks, giving the impression of a contraction. It seems likely that rhythmic mesenchyme condensation assists the choanocyte chambers in pumping water through the sponge.     

Cloning of Hsp70 genes from the marine sponges Sycon raphanus (Calcarea) and Rhabdocalyptus dawsoni (Hexactinellida). An approach to solve the phylogeny of sponges

The phylogenetic relationships among the three classes of the Porifera-Demospongiae, Calcarea and Hexactinellida-are still unresolved, despite the use of molecular analyses of rRNA. To determine whether phylogenetic resolution of these classes is possible based on genes coding for specific proteins, in the present study the genes for the 70 kDa heat shock protein [Hsp70] were isolated from Rhabdocalyptus dawsoni [Hexactinellida] and from Sycon raphanus [Calcarea], and compared to that previously isolated from the demosponge Geodia cydonium. The gene from R. dawsoni is 2021 bp long and encodes a predicted Hsp70 of Mr 77, 697; the protein comprises the characteristic sites of eukaryotic, cytoplasmic Hsp70 polypeptides. The Hsp70 isolated from cDNA from S. raphanus is 2326 bp long. It encodes a potential polypeptide of Mr 85, 927 and belongs to the same class of Hsp70s. All three sponge sequences for Hsp70 were found to be highly identical to both human and plant Hsp70s. The degree of identity at the amino acid (aa) level between the sponge sequences and the human sequence for Hsp70 is 77%-84% and at the nucleotide (nt) level, between 69% and 75%. Resolution of the phylogenetic relationship between the three classes of sponges based on the Hsp70 was not possible due to the high degree of identity [similarity] of their respective aa sequences, which ranged from 80% [90%] to 82% [91%]. The evolutionary rates-K_aa-values-calculated for the sponge Hsp70 molecules, are low, reflecting the strong functional contraints placed upon these polypeptides. These values range from 0.125 times 10^-9 for G. cydonium and R. dawsoni to 0.087 times 10^-9 for S. raphanus. Higher values have previously been reported for the G. cydonium galectin molecule [K_aa-value of 1.7 times 10^-9] and the receptor tyrosine kinase [1.24 times 10^-9] from the same animal. The occurrence of at least one double mutation, in the codon for the aa Ser in the conserved regions of the Hsp70 sequences, also suggests that these molecules are subjected to strong functional constraints.     

Ultrastructural study of embryonic development in Grantia compressa F. (Porifera, Calcarea)

The embryonic development of Grantia compressa is studied by means of the electron microscope from the blastula inside the mesenchyme to the mature amphi-blastula released in the excurrent canals. The study of the different cellular categories of the embryon shows the distribution of the vitellin inclusions and their evolution. The ultrastructure of the "cellules en croix" is not in favour of a photoreceptor part.     

Calcareous sponges of the genera Clathrina and Guancha (Calcinea, Calcarea, Porifera) of Norway (north-east Atlantic) with the description of five new species

The taxonomy and distribution of 11 species of calcareous sponges of the subclass Calcinea from the Norwegian coast are reviewed. The Norwegian Calcinea represents a mixture of southern boreal/boreal and boreoarctic species, and the calcinean sponge fauna of northern Norway has strong similarities to the Greenlandic and the White Sea/Barents Sea sponge faunas. Most Norwegian Calcinea have their main distribution between 20 and 100 m depth, although some species are found only in the shallow sublittoral or from sublittoral to abyssal depths. Six species were previously reported in the area: Clathrina coriacea (Montagu, 1818), Clathrina cribrata Rapp et al ., 2001, Clathrina nanseni (Breitfuss, 1896), Clathrina septentrionalis Rapp et al ., 2001, Guancha blanca Miklucho-Maclay, 1868 and Guancha lacunosa (Johnston, 1842). Five species are new to science: Clathrina corallicola , Clathrina jorunnae , Guancha arnesenae , Guancha camura , and Guancha pellucida spp. nov. A key to the known Norwegian Calcinea is provided.  © 2006 The Linnean Society of London, Zoological Journal of the Linnean Society , 2006, 147 , 331–365.     

The filtration apparatus for food collection in freshwater sponges (Porifera,Spongillidae)

Summary The freshwater sponges (Spongillidae) feed by filtering out small particles from the water passing through them by means of strainer devices in the flagellated chambers. These are filamentous, fine-meshed structures at the distal ends of the choanocyte collars formed of a mucous material similar to that in the glycocalyx. Each strainer separates its flagellated chamber into an outer and an inner zone. The strainers are an extremely efficient filtering mechanism.