New 3,4-seco-3,19-Dinor- and Spongian-Based Diterpenoid Lactones from the Marine Sponge Spongia sp.

Continuing chemical investigation of the Red Sea sponge Spongia sp. led to the isolation of four new 3,4-seco-3,19-dinorspongian diterpenoid lactones, secodinorspongins A-D (1-4), along with a classical spongian diterpenoid lactone, sponginolide (5). The chemical structures, including the absolute configurations of these compounds, were elucidated using the extensive spectroscopic study composed of 1D and 2D NMR data analyses, and a comparison between calculated-electronic-circular-dichroism (ECD) and experimental-circular-dichroism (CD) spectra. A plausible biosynthetic pathway of 1-4 was also proposed. Furthermore, the cytotoxicity, antibacterial and anti-inflammatory activities of 1-5 were evaluated. Compound 1 was found to exhibit inhibitory activity against the growth of Staphylococcus aureus (S. aureus), and 4 and 5 exhibited suppression of superoxide-anion generation and elastase release in fMLF/CB-induced human neutrophils.

Assessing the genomic composition, putative ecological relevance and biotechnological potential of plasmids from sponge bacterial symbionts.

Plasmid-mediated transfer of genes can have direct consequences in several biological processes within sponge microbial communities. However, very few studies have attempted genomic and functional characterization of plasmids from marine host-associated microbial communities in general and those of sponges in particular. In the present study, we used an endogenous plasmid isolation method to obtain plasmids from bacterial symbionts of the marine sponges Stylissa carteri and Paratetilla sp. and investigated the genomic composition, putative ecological relevance and biotechnological potential of these plasmids. In total, we isolated and characterized three complete plasmids, three plasmid prophages and one incomplete plasmid. Our results highlight the importance of plasmids to transfer relevant genetic traits putatively involved in microbial symbiont adaptation and host-microbe and microbe-microbe interactions. For example, putative genes involved in bacterial response to chemical stress, competition, metabolic versatility and mediation of bacterial colonization and pathogenicity were detected. Genes coding for enzymes and toxins of biotechnological potential were also detected. Most plasmid prophage coding sequences were, however, hypothetical proteins with unknown functions. Overall, this study highlights the ecological relevance of plasmids in the marine sponge microbiome and provides evidence that plasmids of sponge bacterial symbionts may represent an untapped resource of genes of biotechnological interest.

Spongenolactones A-C, Bioactive 5,5,6,6,5-Pentacyclic Spongian Diterpenes from the Red Sea Sponge Spongia sp.

Three new 5,5,6,6,5-pentacyclic spongian diterpenes, spongenolactones A-C (1-3), were isolated from a Red Sea sponge Spongia sp. The structures of the new metabolites were elucidated by extensive spectroscopic analysis and the absolute configurations of 1-3 were determined on the basis of comparison of the experimental circular dichroism (CD) and calculated electronic circular dichroism (ECD) spectra. Compounds 1-3 are the first 5,5,6,6,5-pentacyclic spongian diterpenes bearing an ß-hydroxy group at C-1. These metabolites were assayed for their cytotoxic, antibacterial, and anti-inflammatory activities. All three compounds were found to exert inhibitory activity against superoxide anion generation in fMLF/CB-stimulated human neutrophils. Furthermore, 1 showed a higher activity against the growth of Staphylococcus aureus in comparison to 2.

The Chemically Highly Diversified Metabolites from the Red Sea Marine Sponge Spongia sp.

A polyoxygenated and halogenated labdane, spongianol (1); a polyoxygenated steroid, 3ß,5α,9α-trihydroxy-24S-ethylcholest-7-en-6-one (2); a rare seven-membered lactone B ring, (22E,24S)-ergosta-7,22-dien-3ß,5α-diol-6,5-olide (3); and an α,ß-unsaturated fatty acid, (Z)-3-methyl-9-oxodec-2-enoic acid (4) as well as five known compounds, 10-hydroxykahukuene B (5), pacifenol (6), dysidamide (7), 7,7,7-trichloro-3-hydroxy-2,2,6-trimethyl-4-(4,4,4-trichloro-3-methyl-1-oxobu-tylamino)-heptanoic acid methyl ester (8), and the primary metabolite 2'-deoxynucleoside thymidine (9), have been isolated from the Red Sea sponge Spongia sp. The stereoisomer of 3 was discovered in Ganoderma resinaceum, and metabolites 5 and 6, isolated previously from red algae, were characterized unprecedentedly in the sponge. Compounds 7 and 8 have not been found before in the genus Spongia. Compounds 1-9 were also assayed for cytotoxicity as well as antibacterial and anti-inflammatory activities.

Marine Sponge and Octocoral-Associated Bacteria Show Versatile Secondary Metabolite Biosynthesis Potential and Antimicrobial Activities against Human Pathogens.

Marine microbiomes are prolific sources of bioactive natural products of potential pharmaceutical value. This study inspected two culture collections comprising 919 host-associated marine bacteria belonging to 55 genera and several thus-far unclassified lineages to identify isolates with potentially rich secondary metabolism and antimicrobial activities. Seventy representative isolates had their genomes mined for secondary metabolite biosynthetic gene clusters (SM-BGCs) and were screened for antimicrobial activities against four pathogenic bacteria and five pathogenic Candida strains. In total, 466 SM-BGCs were identified, with antimicrobial peptide- and polyketide synthase-related SM-BGCs being frequently detected. Only 38 SM-BGCs had similarities greater than 70% to SM-BGCs encoding known compounds, highlighting the potential biosynthetic novelty encoded by these genomes. Cross-streak assays showed that 33 of the 70 genome-sequenced isolates were active against at least one Candida species, while 44 isolates showed activity against at least one bacterial pathogen. Taxon-specific differences in antimicrobial activity among isolates suggested distinct molecules involved in antagonism against bacterial versus Candida pathogens. The here reported culture collections and genome-sequenced isolates constitute a valuable resource of understudied marine bacteria displaying antimicrobial activities and potential for the biosynthesis of novel secondary metabolites, holding promise for a future sustainable production of marine drug leads.

An Anti-Inflammatory 2,4-Cyclized-3,4-Secospongian Diterpenoid and Furanoterpene-Related Metabolites of a Marine Sponge Spongia sp. from the Red Sea.

Chemical investigation of a Red Sea Spongia sp. led to the isolation of four new compounds, i.e., 17-dehydroxysponalactone (1), a carboxylic acid, spongiafuranic acid A (2), one hydroxamic acid, spongiafuranohydroxamic acid A (3), and a furanyl trinorsesterpenoid 16-epi-irciformonin G (4), along with three known metabolites (-)-sponalisolide B (5), 18-nor- 3,17-dihydroxy-spongia-3,13(16),14-trien-2-one (6), and cholesta-7-ene-3ß,5α-diol-6-one (7). The biosynthetic pathway for the molecular skeleton of 1 and related compounds was postulated for the first time. Anti-inflammatory activity of these metabolites to inhibit superoxide anion generation and elastase release in N-formyl-methionyl-leucyl phenylalanine/cytochalasin B (fMLF/CB)-induced human neutrophil cells and cytotoxicity of these compounds toward three cancer cell lines and one human dermal fibroblast cell line were assayed. Compound 1 was found to significantly reduce the superoxide anion generation and elastase release at a concentration of 10 µM, and compound 5 was also found to display strong inhibitory activity against superoxide anion generation at the same concentration. Due to the noncytotoxic activity and the potent inhibitory effect toward the superoxide anion generation and elastase release, 1 and 5 can be considered to be promising anti-inflammatory agents.

Characterization of putative circular plasmids in sponge-associated bacterial communities using a selective multiply-primed rolling circle amplification.

Plasmid transfers among bacterial populations can directly influence the ecological adaptation of these populations and their interactions with host species and environment. In this study, we developed a selective multiply-primed rolling circle amplification (smRCA) approach to enrich and characterize circular plasmid DNA from sponge microbial symbionts via high-throughput sequencing (HTS). DNA (plasmid and total community DNA) obtained from sponge (Cinachyrella sp.) samples and a bacterial symbiont (Vibrio sp. CyArs1) isolated from the same sponge species (carrying unknown plasmids) were used to develop and validate our methodology. The smRCA was performed during 16 hr with 141 plasmid-specific primers covering all known circular plasmid groups. The amplified products were purified and subjected to a reamplification with random hexamer primers (2 hr) and then sequenced using Illumina MiSeq. The developed method resulted in the successful amplification and characterization of the sponge plasmidome and allowed us to detect plasmids associated with the bacterial symbiont Vibrio sp. CyArs1 in the sponge host. In addition to this, a large number of small (<2 kbp) and cryptic plasmids were also amplified in sponge samples. Functional analysis identified proteins involved in the control of plasmid partitioning, maintenance and replication. However, most plasmids contained unknown genes, which could potentially serve as a resource of unknown genetic information and novel replication systems. Overall, our results indicate that the smRCA-HTS approach developed here was able to selectively enrich and characterize plasmids from bacterial isolates and sponge host microbial communities, including plasmids larger than 20 kbp.

Sponge-Derived 24-Homoscalaranes as Potent Anti-Inflammatory Agents.

Scalarane-type sesterterpenoids are known for their therapeutic potential in cancer treatments. However, the anti-inflammatory properties of this class of metabolites remain elusive. Our current work aimed to investigate the anti-inflammatory scalaranes from marine sponge Lendenfeldia sp., resulting in the isolation of six new 24-homoscalaranes, lendenfeldaranes E-J (1-6). The structures of the new metabolites were determined by extensive spectroscopic analyses, and the absolute configuration of 1 was established by electronic circular dichroism (ECD) calculations. Compounds 2 and 3 were discovered to individually reduce the generation of superoxide anions, and compound 1 displayed an inhibitor effect on the release of elastase. These three compounds were proven to be the first anti-neutrophilic scalaranes.

Probing Anti-Proliferative 24-Homoscalaranes from a Sponge Lendenfeldiasp.

In the current study, an NMR spectroscopic pattern-based procedure for probing scalarane derivatives was performed and four new 24-homoscalaranes, lendenfeldaranes A-D (1- 4), along with three known compounds, 12α-acetoxy-22-hydroxy-24-methyl-24-oxoscalar-16-en- 25-al (5), felixin F (6), and 24-methyl-12,24,25-trioxoscalar-16-en-22-oic acid (7) were isolated from the sponge Lendenfeldia sp. The structures of scalaranes 1-7 were elucidated on the basis of spectroscopic analysis. Scalaranes 1-7 were further evaluated for their cytotoxicity toward a series of human cancer cell lines and the results suggested that 5 and 7 dominated in the anti- proliferative activity of the extract. The 18-aldehyde functionality was found to play a key role in their activity.

Compositional variation between high and low prokaryotic diversity coral reef biotopes translates to different predicted metagenomic gene content.

In a previous study, we identified host species that housed high and low diversity prokaryotic communities. In the present study, we expand on this and assessed the prokaryotic communities associated with seawater, sediment and 11 host species from 7 different phyla in a Taiwanese coral reef setting. The host taxa sampled included hard, octo- and black corals, molluscs, bryozoans, flatworms, fish and sea urchins. There were highly significant differences in composition among host species and all host species housed distinct communities from those found in seawater and sediment. In a hierarchical clustering analysis, samples from all host species, with the exception of the coral Galaxea astreata, formed significantly supported clusters. In addition to this, the coral G. astreata and the bryozoan Triphyllozoon inornatum on the one hand and the coral Tubastraea coccinea, the hermit crab Calcinus laevimanus and the flatworm Thysanozoon nigropapillosum on the other formed significantly supported clusters. In addition to composition, there were highly pronounced differences in richness and evenness among host species from the most diverse species, the bryozoan T. inornatum at 2518 ± 240 OTUs per 10,000 sequences to the least diverse species, the octocoral Cladiella sp. at 142 ± 14 OTUs per 10,000 sequences. In line with the differences in composition, there were significant differences in predicted metagenomic gene counts among host species. Furthermore, there were pronounced compositional and predicted functional differences between high diversity hosts (Liolophura japonica, G. astreata, T. coccinea, C. laevimanus, T. inornatum) and low diversity hosts (Antipathes sp., Pomacentrus coelestis, Modiolus auriculatus, T. nigropapillosum, Cladiella sp. and Diadema savigny). In particular, we found that all tested low diversity hosts were predicted to be enriched for the phosphotransferase system compared to high diversity hosts.

The sponge microbiome within the greater coral reef microbial metacommunity.

Much recent marine microbial research has focused on sponges, but very little is known about how the sponge microbiome fits in the greater coral reef microbial metacommunity. Here, we present an extensive survey of the prokaryote communities of a wide range of biotopes from Indo-Pacific coral reef environments. We find a large variation in operational taxonomic unit (OTU) richness, with algae, chitons, stony corals and sea cucumbers housing the most diverse prokaryote communities. These biotopes share a higher percentage and number of OTUs with sediment and are particularly enriched in members of the phylum Planctomycetes. Despite having lower OTU richness, sponges share the greatest percentage (>90%) of OTUs with >100 sequences with the environment (sediment and/or seawater) although there is considerable variation among sponge species. Our results, furthermore, highlight that prokaryote microorganisms are shared among multiple coral reef biotopes, and that, although compositionally distinct, the sponge prokaryote community does not appear to be as sponge-specific as previously thought.