The cladorhizid fauna (Porifera, Poecilosclerida) of the Caribbean and adjacent waters

The carnivorous sponge family Cladorhizidae has been subject to several recent studies, yet the cladorhizid fauna of the Caribbean and adjacent areas remain comparatively poorly known. In this article we provide a description of the novel species Abyssocladia polycephalus sp. nov. from the Muir Seamount NE of Bermuda, belonging to the mainly Pacific genus Abyssocladia, and Asbestopluma (Asbestopluma) caribica sp. nov. from the Beata Ridge. Additionally, we provide a re-description of the poorly known species Chondrocladia (Chondrocladia) verticillata Topsent, 1920, and compare this species with the closely related species C. (C.) concrescens Schmidt, 1880. Finally, we provide a brief overview of the carnivorous sponges known from the Gulf of Mexico, Caribbean Sea and adjacent Atlantic Ocean.publishedVersio

Molecular Techniques Reveal Wide Phyletic Diversity of Heterotrophic Microbes Associated with Discodermia spp. (Porifera: Demospongiae)

Sponges are well known to harbor large numbers of heterotrophic microbes within their mesohyl. Studies to determine the diversity of these associated microbes have been attempted for only a few shallow water species. We cultured various microorganisms from several species of Discodermia collected from deep water using the \u27Johnson-Sea-Link\u27 manned submersibles, and characterised them by standard microbiological identification methods. Characterisation of a small proportion (ca. 10%) of the total and potential eubacterial isolate collection with molecular systematics techniques revealed a wide diversity of microbes. Phylogenetic analyses of 32 small subunit (SSU) 16S-like rRNA gene sequences from different micorbes indicated high levels of taxonomic diversity assoiated with this genus of sponge. For example, bacteria from at least five cubacterial subdivisions - gamma, alpha, beta, Cytophaga and Gram positive - were isolated from the mesohyl of Discodermia. Several strains were unidentifiable from current sequence databases. No overlap was found between sequences of 24 isolates and 8 sequences obtained by PCR and cloning directly from sponge samples. The abundance and diversity of microbes associated with sponges such as Discodermia suggest that they may play important roles in marine microbial ecology, dispersal and evolution

Differential modulation of microglia superoxide anion and thromboxane B(2 )generation by the marine manzamines

BACKGROUND: Thromboxane B(2 )(TXB(2)) and superoxide anion (O(2)(-)) are neuroinflammatory mediators that appear to be involved in the pathogenesis of several neurodegenerative diseases. Because activated-microglia are the main source of TXB(2 )and O(2)(- )in these disorders, modulation of their synthesis has been hypothesized as a potential therapeutic approach for neuroinflammatory disorders. Marine natural products have become a source of novel agents that modulate eicosanoids and O(2)(- )generation from activated murine and human leukocytes. With the exception of manzamine C, all other manzamines tested are characterized by a complex pentacyclic diamine linked to C-1 of the β-carboline moiety. These marine-derived alkaloids have been reported to possess a diverse range of bioactivities including anticancer, immunostimulatory, insecticidal, antibacterial, antimalarial and antituberculosis activities. The purpose of this investigation was to conduct a structure-activity relationship study with manzamines (MZ) A, B, C, D, E and F on agonist-stimulated release of TXB(2 )and O(2)(- )from E. coli LPS-activated rat neonatal microglia in vitro. RESULTS: The manzamines differentially attenuated PMA (phorbol 12-myristate 13-acetate)-stimulated TXB(2 )generation in the following order of decreasing potency: MZA (IC(50 )<0.016 μM) >MZD (IC(50 )= 0.23 μM) >MZB (IC(50 )= 1.6 μM) >MZC (IC(50 )= 2.98 μM) >MZE and F (IC(50 )>10 μM). In contrast, there was less effect on OPZ (opsonized zymosan)-stimulated TXB(2 )generation: MZB (IC(50 )= 1.44 μM) >MZA (IC(50 )= 3.16 μM) >MZC (IC(50 )= 3.34 μM) >MZD, MZE and MZF (IC(50 )>10 μM). Similarly, PMA-stimulated O(2)(- )generation was affected differentially as follows: MZD (apparent IC(50)<0.1 μM) >MZA (IC(50 )= 0.1 μM) >MZB (IC(50 )= 3.16 μM) >MZC (IC(50 )= 3.43 μM) >MZE and MZF (IC(50 )>10 μM). In contrast, OPZ-stimulated O(2)(- )generation was minimally affected: MZB (IC(50 )= 4.17 μM) >MZC (IC(50 )= 9.3 μM) >MZA, MZD, MZE and MZF (IC(50 )> 10 μM). From the structure-activity relationship perspective, contributing factors to the observed differential bioactivity on TXB(2 )and O(2)(- )generation are the solubility or ionic forms of MZA and D as well as changes such as saturation or oxidation of the β carboline or 8-membered amine ring. In contrast, the fused 13-membered macrocyclic and isoquinoline ring system, and any substitutions in these rings would not appear to be factors contributing to bioactivity. CONCLUSION: To our knowledge, this is the first experimental study that demonstrates that MZA, at in vitro concentrations that are non toxic to E. coli LPS-activated rat neonatal microglia, potently modulates PMA-stimulated TXB(2 )and O(2)(- )generation. MZA may thus be a lead candidate for the development of novel therapeutic agents for the modulation of TXB(2 )and O(2)(- )release in neuroinflammatory diseases. Marine natural products provide a novel and rich source of chemical diversity that can contribute to the design and development of new and potentially useful anti-inflammatory agents to treat neurodegenerative diseases

Breakthrough in marine invertebrate cell culture : Sponge cells divide rapidly in improved nutrient medium

Sponges (Phylum Porifera) are among the oldest Metazoa and considered critical to understanding animal evolution and development. They are also the most prolific source of marine-derived chemicals with pharmaceutical relevance. Cell lines are important tools for research in many disciplines, and have been established for many organisms, including freshwater and terrestrial invertebrates. Despite many efforts over multiple decades, there are still no cell lines for marine invertebrates. In this study, we report a breakthrough: we demonstrate that an amino acid-optimized nutrient medium stimulates rapid cell division in 9 sponge species. The fastest dividing cells doubled in less than 1 hour. Cultures of 3 species were subcultured from 3 to 5 times, with an average of 5.99 population doublings after subculturing, and a lifespan from 21 to 35 days. Our results form the basis for developing marine invertebrate cell models to better understand early animal evolution, determine the role of secondary metabolites, and predict the impact of climate change to coral reef community ecology. Furthermore, sponge cell lines can be used to scale-up production of sponge-derived chemicals for clinical trials and develop new drugs to combat cancer and other diseases.publishedVersio

Evolution of Group I Introns in Porifera: New Evidence for Intron Mobility and Implications for DNA Barcoding

BackgroundMitochondrial introns intermit coding regions of genes and feature characteristic secondary structures and splicing mechanisms. In metazoans, mitochondrial introns have only been detected in sponges, cnidarians, placozoans and one annelid species. Within demosponges, group I and group II introns are present in six families. Based on different insertion sites within the cox1 gene and secondary structures, four types of group I and two types of group II introns are known, which can harbor up to three encoding homing endonuclease genes (HEG) of the LAGLIDADG family (group I) and/or reverse transcriptase (group II). However, only little is known about sponge intron mobility, transmission, and origin due to the lack of a comprehensive dataset. We analyzed the largest dataset on sponge mitochondrial group I introns to date: 95 specimens, from 11 different sponge genera which provided novel insights into the evolution of group I introns. ResultsFor the first time group I introns were detected in four genera of the sponge family Scleritodermidae (Scleritoderma, Microscleroderma, Aciculites, Setidium). We demonstrated that group I introns in sponges aggregate in the most conserved regions of cox1. We showed that co-occurrence of two introns in cox1 is unique among metazoans, but not uncommon in sponges. However, this combination always associates an active intron with a degenerating one. Earlier hypotheses of HGT were confirmed and for the first time VGT and secondary losses of introns conclusively demonstrated. ConclusionThis study validates the subclass Spirophorina (Tetractinellida) as an intron hotspot in sponges. Our analyses confirm that most sponge group I introns probably originated from fungi. DNA barcoding is discussed and the application of alternative primers suggested

CRISPR/Cas12a-mediated gene editing in Geodia barretti sponge cell culture

Sponges and their associated microorganisms are the most prolific source of marine natural products, and many attempts have been made at creating a marine sponge cell line to produce these products efficiently. However, limited knowledge on the nutrients sponge cells require to grow and poor genetic accessibility have hampered progress toward this goal. Recently, a new sponge-specific nutrient medium M1 has been shown to stimulate sponge cells in vitro to divide rapidly. In this study, we demonstrate for the first time that sponge cells growing in M1 can be genetically modified using a CRISPR/Cas12a gene editing system. A short sequence of scrambled DNA was inserted using a single-stranded oligodeoxynucleotide donor template to disrupt the 2′,5′-oligoadenylate synthetase gene of cells from the boreal deep-sea sponge Geodia barretti. A blue fluorescent marker gene appeared to be inserted in an intron of the same gene and expressed by a small number of G. barretti cells. Our results represent an important step toward developing an optimized continuous sponge cell line to produce bioactive compounds.publishedVersio

Divergence times in demosponges (Porifera): first insights from new mitogenomes and the inclusion of fossils in a birth-death clock model

Background: Approximately 80% of all described extant sponge species belong to the class Demospongiae. Yet, despite their diversity and importance, accurate divergence times are still unknown for most demosponge clades. The estimation of demosponge divergence time is key to answering fundamental questions on the origin of Demospongiae, their diversification and historical biogeography. Molecular sequence data alone is not informative on an absolute time scale, and therefore needs to be "calibrated" with additional data such as fossils. Here, we calibrate the molecular data with the fossilized birth-death model, which compared to strict node dating, allows for the inclusion of young and old fossils in the analysis of divergence time. We use desma-bearing sponges, a diverse group of demosponges that form rigid skeletons and have a rich and continuous fossil record dating back to the Cambrian (similar to 500 Ma), to date the demosponge radiation and constrain the timing of key evolutionary events, like the transition from marine to freshwater habitats. To infer a dated phylogeny of Demospongiae we assembled the mitochondrial genomes of six desma-bearing demosponges from reduced-representation genomic libraries. The total dataset included 33 complete demosponge mitochondrial genomes and 30 fossils. Results: Our study supports a Neoproterozoic origin of Demospongiae. Novel age estimates for the split of freshwater and marine sponges dating back to the Carboniferous and the previously assumed recent (similar to 18 Ma) diversification of freshwater sponges is supported. Moreover, we provide detailed age estimates for a possible diversification of Tetractinellidae (similar to 315 Ma), the Astrophorina (similar to 240 Ma), the Spirophorina (similar to 120 Ma) and the family Corallistidae (similar to 88 Ma) all of which are considered as key groups for dating the Demospongiae due to their extraordinary rich and continuous fossil history. Conclusion: This study provides novel insights into the evolution of Demospongiae. Observed discrepancies of our dated phylogeny with their putative first fossil appearance dates are discussed for selected sponge groups. For instance, a Carboniferous origin of the order Tetractinellida seems to be too late, compared to their first appearance in the fossil record in the Middle Cambrian. This would imply that Paleozoic spicule forms are not homologous to post-Paleozoic forms

Reconstruction of Family-Level Phylogenetic Relationships within Demospongiae (Porifera) Using Nuclear Encoded Housekeeping Genes

Background: Demosponges are challenging for phylogenetic systematics because of their plastic and relatively simple morphologies and many deep divergences between major clades. To improve understanding of the phylogenetic relationships within Demospongiae, we sequenced and analyzed seven nuclear housekeeping genes involved in a variety of cellular functions from a diverse group of sponges. Methodology/Principal Findings: We generated data from each of the four sponge classes (i.e., Calcarea, Demospongiae, Hexactinellida, and Homoscleromorpha), but focused on family-level relationships within demosponges. With data for 21 newly sampled families, our Maximum Likelihood and Bayesian-based approaches recovered previously phylogenetically defined taxa: Keratosap, Myxospongiaep, Spongillidap, Haploscleromorphap (the marine haplosclerids) and Democlaviap. We found conflicting results concerning the relationships of Keratosap and Myxospongiaep to the remaining demosponges, but our results strongly supported a clade of Haploscleromorphap+Spongillidap+Democlaviap. In contrast to hypotheses based on mitochondrial genome and ribosomal data, nuclear housekeeping gene data suggested that freshwater sponges (Spongillidap) are sister to Haploscleromorphap rather than part of Democlaviap. Within Keratosap, we found equivocal results as to the monophyly of Dictyoceratida. Within Myxospongiaep, Chondrosida and Verongida were monophyletic. A well supported clade within Democlaviap, Tetractinellidap, composed of all sampled members of Astrophorina and Spirophorina (including the only lithistid in our analysis), was consistently revealed as the sister group to all other members of Democlaviap. Within Tetractinellidap, we did not recover monophyletic Astrophorina or Spirophorina. Our results also reaffirmed the monophyly of order Poecilosclerida (excluding Desmacellidae and Raspailiidae), and polyphyly of Hadromerida and Halichondrida. Conclusions/Significance: These results, using an independent nuclear gene set, confirmed many hypotheses based on ribosomal and/or mitochondrial genes, and they also identified clades with low statistical support or clades that conflicted with traditional morphological classification. Our results will serve as a basis for future exploration of these outstanding questions using more taxon- and gene-rich datasets