Vicingus serpentipes gen. nov., sp. nov., a new member of the Flavobacteriales from the North Sea.

A new member of the Flavobacteriales was isolated from the surface of a stone collected on the German North Sea shore. The bacterium, strain ANORD5T, is a mesophilic, chemoheterotrophic aerobic, typical marine bacterium. Optimal growth was observed at 20-30 °C, pH 7.0-8.5 and 1-2 % sea salt. The 16S rRNA gene sequence revealed a distant relationship with the representatives of the Cryomorphaceae, with less than 90 % sequence similarity. Strain ANORD5T forms a cluster together with Owenweeksia hongkongensis UST20020801T (89.9 %), Cryomorpha ignava 1-22T (87.9 %), Luteibaculum oceani CC-AMWY-103BT (88.1 %) and Phaeocystidibacter luteus PG2S01T (87.3 %). Strain ANORD5T has a low DNA G+C content (31 mol%). Based on morphological, physiological and phylogenetic data, strain ANORD5T is considered a type strain of a new species and a new genus of the family Cryomorphaceae for which the name Vicingus serpentipes is proposed. The type strain is ANORD5T (=NCIMB 15042T=DSM 103558T=MTCC 12686T).

Asperentin B, a New Inhibitor of the Protein Tyrosine Phosphatase 1B.

In the frame of studies on secondary metabolites produced by fungi from deep-sea environments we have investigated inhibitors of enzymes playing key roles in signaling cascades of biochemical pathways relevant for the treatment of diseases. Here we report on a new inhibitor of the human protein tyrosine phosphatase 1B (PTP1B), a target in the signaling pathway of insulin. A new asperentin analog is produced by an Aspergillussydowii strain isolated from the sediment of the deep Mediterranean Sea. Asperentin B (1) contains an additional phenolic hydroxy function at C-6 and exhibits an IC50 value against PTP1B of 2 µM in vitro, which is six times stronger than the positive control, suramin. Interestingly, asperentin (2) did not show any inhibition of this enzymatic activity. Asperentin B (1) is discussed as possible therapeutic agents for type 2 diabetes and sleeping sickness.

Biscogniauxone, a New Isopyrrolonaphthoquinone Compound from the Fungus Biscogniauxia mediterranea Isolated from Deep-Sea Sediments.

The properties and the production of new metabolites from the fungal strain LF657 isolated from the Herodotes Deep (2800 m depth) in the Mediterranean Sea are reported in this study. The new isolate was identified as Biscogniauxia mediterranea based on ITS1-5.8S-ITS2 and 28S rRNA gene sequences. A new isopyrrolonaphthoquinone with inhibitory activity against glycogen synthase kinase (GSK-3ß) was isolated from this fungus. This is the first report of this class of compounds from a fungus isolated from a deep-sea sediment, as well as from a Biscogniauxia species.

Marine Fungi as Producers of Benzocoumarins, a New Class of Inhibitors of Glycogen-Synthase-Kinase 3ß.

The glycogen-synthase-kinase 3 (GSK-3) is an important target in drug discovery. This enzyme is involved in the signaling pathways of type 2 diabetes, neurological disorders, cancer, and other diseases. Therefore, inhibitors of GSK-3 are promising drug candidates for the treatment of a broad range of diseases. Here we report pannorin (1), alternariol (2), and alternariol-9-methylether (3) to be promising inhibitors of the isoform GSK-3ß showing sub-µM IC50 values. The in vitro inhibition is in the range of the known highly active GSK-3ß inhibitor TDZD-8. Compounds 1-3 have a highly oxygenated benzocoumarin core structure in common, which suggests that this may be a new structural feature for efficient GSK-3ß inhibition.

Engyodontochones, Antibiotic Polyketides from the Marine Fungus Engyodontium album Strain LF069.

Six new (2, 4-8) and two known polyketides with a basic structure of an anthraquinone-xanthone were isolated from mycelia and culture broth of the fungus Engyodontium album strain LF069. The structures and relative configurations of these compounds were established by spectroscopic means, and their absolute configurations were defined mainly by comparison of quantum chemical TDDFT calculated and experimental ECD spectra. Compounds 2 and 4-8 were given the trivial names engyodontochone A (2) and B-F (4-8). Compounds 5-8 represent the first example of a 23,28 seco-beticolin carbon skeleton. The relative and absolute configurations of two known substances JBIR-97/98 (1) and JBIR-99 (3) were determined for the first time. All isolated compounds were subjected to bioactivity assays. Compounds 1-4 exhibited inhibitory activity against methicillin-resistant Staphylococcus aureus (MRSA) that was 10-fold stronger than chloramphenicol.

Lindgomycin, an Unusual Antibiotic Polyketide from a Marine Fungus of the Lindgomycetaceae.

An unusual polyketide with a new carbon skeleton, lindgomycin (1), and the recently described ascosetin (2) were extracted from mycelia and culture broth of different Lindgomycetaceae strains, which were isolated from a sponge of the Kiel Fjord in the Baltic Sea (Germany) and from the Antarctic. Their structures were established by spectroscopic means. In the new polyketide, two distinct domains, a bicyclic hydrocarbon and a tetramic acid, are connected by a bridging carbonyl. The tetramic acid substructure of compound 1 was proved to possess a unique 5-benzylpyrrolidine-2,4-dione unit. The combination of 5-benzylpyrrolidine-2,4-dione of compound 1 in its tetramic acid half and 3-methylbut-3-enoic acid pendant in its decalin half allow the assignment of a new carbon skeleton. The new compound 1 and ascosetin showed antibiotic activities with IC50 value of 5.1 (±0.2) µM and 3.2 (±0.4) µM, respectively, against methicillin-resistant Staphylococcus aureus.

Identification of Habitat-Specific Biomes of Aquatic Fungal Communities Using a Comprehensive Nearly Full-Length 18S rRNA Dataset Enriched with Contextual Data.

Molecular diversity surveys have demonstrated that aquatic fungi are highly diverse, and that they play fundamental ecological roles in aquatic systems. Unfortunately, comparative studies of aquatic fungal communities are few and far between, due to the scarcity of adequate datasets. We combined all publicly available fungal 18S ribosomal RNA (rRNA) gene sequences with new sequence data from a marine fungi culture collection. We further enriched this dataset by adding validated contextual data. Specifically, we included data on the habitat type of the samples assigning fungal taxa to ten different habitat categories. This dataset has been created with the intention to serve as a valuable reference dataset for aquatic fungi including a phylogenetic reference tree. The combined data enabled us to infer fungal community patterns in aquatic systems. Pairwise habitat comparisons showed significant phylogenetic differences, indicating that habitat strongly affects fungal community structure. Fungal taxonomic composition differed considerably even on phylum and class level. Freshwater fungal assemblage was most different from all other habitat types and was dominated by basal fungal lineages. For most communities, phylogenetic signals indicated clustering of sequences suggesting that environmental factors were the main drivers of fungal community structure, rather than species competition. Thus, the diversification process of aquatic fungi must be highly clade specific in some cases.The combined data enabled us to infer fungal community patterns in aquatic systems. Pairwise habitat comparisons showed significant phylogenetic differences, indicating that habitat strongly affects fungal community structure. Fungal taxonomic composition differed considerably even on phylum and class level. Freshwater fungal assemblage was most different from all other habitat types and was dominated by basal fungal lineages. For most communities, phylogenetic signals indicated clustering of sequences suggesting that environmental factors were the main drivers of fungal community structure, rather than species competition. Thus, the diversification process of aquatic fungi must be highly clade specific in some cases.

Acetylcholinesterase inhibitors from a marine fungus Talaromyces sp. strain LF458.

Two new oxaphenalenone dimers, talaromycesone A (1) and talaromycesone B (2), and a new isopentenyl xanthenone, talaroxanthenone (3), together with six known diphenyl ether derivatives, e.g., Δ(1',3'),-1'-dehydroxypenicillide (4), 1',2'-dehydropenicillide (5), vermixocin A (6), vermixocin B (7), 3'-methoxy-1'2'-dehydropenicillide (8), and AS-186c (9), were isolated from the culture broth and mycelia of a marine fungus Talaromyces sp. strain LF458. Compound 2 represents the first example of 1-nor oxaphenalenone dimer carbon skeleton. All isolated compounds were subjected to bioactivity assays. Compounds 1, 2, and 9 exhibited potent antibacterial activities with IC50 3.70, 17.36, and 1.34 µM, respectively, against human pathogenic Staphylococcus strains. Compounds 1, 3, and 9 displayed potent acetylcholinesterase inhibitory activities with IC50 7.49, 1.61, and 2.60 µM, respectively. Interestingly, phosphodiesterase PDE-4B2 was inhibited by compounds 3 (IC50 7.25 µM) and 9 (IC50 2.63 µM).

Spirocyclic drimanes from the marine fungus Stachybotrys sp. strain MF347.

A novel spirocyclic drimane coupled by two drimane fragment building blocks 2 and a new drimane 1 were identified in mycelia and culture broth of Stachybotrys sp. MF347. Their structures were established by spectroscopic means. This is the first example of spirocyclic drimane coupled by a spirodihydrobenzofuranlactam unit and a spirodihydroisobenzofuran unit; and the connecting position being N-C instead of an N and N connecting unit. Strain MF347 produced also the known spirocyclic drimanes stachybocin A (12) and stachybocin B (11) featured by two sesquiterpene-spirobenzofuran structural units connected by a lysine residue; the known spirocyclic drimanes chartarlactam O (5); chartarlactam K (6); F1839A (7); stachybotrylactam (8); stachybotramide (9); and 2α-acetoxystachybotrylactam acetate (10); as well as ilicicolin B (13), a known sesquiterpene. The relative configuration of two known spirobenzofuranlactams (3 and 4) was determined. All compounds were subjected to biological activity tests. The spirocyclic drimane 2, 11, and 12, as well as the sesquiterpene 13, exhibited antibacterial activity against the clinically relevant methicillin-resistant Staphylococcus aureus (MRSA).

Two new antibiotic pyridones produced by a marine fungus, Trichoderma sp. strain MF106.

Two unusual pyridones, trichodin A (1) and trichodin B (2), together with the known compound, pyridoxatin (3), were extracted from mycelia and culture broth of the marine fungus, Trichoderma sp. strain MF106 isolated from the Greenland Seas. The structures of the new compounds were characterized as an intramolecular cyclization of a pyridine basic backbone with a phenyl group. The structure and relative configuration of the new compounds were established by spectroscopic means. The new compound 1 and the known compound 3 showed antibiotic activities against the clinically relevant microorganism, Staphylococcus epidermidis, with IC50 values of 24 µM and 4 µM, respectively.

Widespread occurrence of two carbon fixation pathways in tubeworm endosymbionts: lessons from hydrothermal vent associated tubeworms from the mediterranean sea.

Vestimentiferan tubeworms (siboglinid polychetes) of the genus Lamellibrachia are common members of cold seep faunal communities and have also been found at sedimented hydrothermal vent sites in the Pacific. As they lack a digestive system, they are nourished by chemoautotrophic bacterial endosymbionts growing in a specialized tissue called the trophosome. Here we present the results of investigations of tubeworms and endosymbionts from a shallow hydrothermal vent field in the Western Mediterranean Sea. The tubeworms, which are the first reported vent-associated tubeworms outside the Pacific, are identified as Lamellibrachia anaximandri using mitochondrial ribosomal and cytochrome oxidase I (COI) gene sequences. They harbor a single gammaproteobacterial endosymbiont. Carbon isotopic data, as well as the analysis of genes involved in carbon and sulfur metabolism indicate a sulfide-oxidizing chemoautotrophic endosymbiont. The detection of a hydrogenase gene fragment suggests the potential for hydrogen oxidation as alternative energy source. Surprisingly, the endosymbiont harbors genes for two different carbon fixation pathways, the Calvin-Benson-Bassham (CBB) cycle as well as the reductive tricarboxylic acid (rTCA) cycle, as has been reported for the endosymbiont of the vent tubeworm Riftia pachyptila. In addition to RubisCO genes we detected ATP citrate lyase (ACL - the key enzyme of the rTCA cycle) type II gene sequences using newly designed primer sets. Comparative investigations with additional tubeworm species (Lamellibrachia luymesi, Lamellibrachia sp. 1, Lamellibrachia sp. 2, Escarpia laminata, Seepiophila jonesi) from multiple cold seep sites in the Gulf of Mexico revealed the presence of acl genes in these species as well. Thus, our study suggests that the presence of two different carbon fixation pathways, the CBB cycle and the rTCA cycle, is not restricted to the Riftia endosymbiont, but rather might be common in vestimentiferan tubeworm endosymbionts, regardless of the habitat.

Diacidene, a polyene dicarboxylic acid from a Micromonospora isolate from the German Wadden Sea.

Micromonospora sp. strain DB620 was isolated from a Wadden Sea sediment sample collected near Büsum (Germany) and is closely related (99% 16S-rRNA gene sequence similarity) to Micromonospora coxensis strain MTCC8093. It produced a new polyene dicarboxylic acid named diacidene (1) and in addition a derivative of chorismic acid, the known 3-[(1-carboxyvinyl)oxy]benzoic acid. The structure elucidation of 1 was achieved by applying different 1D and 2D NMR techniques as well as mass spectrometry and UV spectroscopy.

Phylogenetic identification of fungi isolated from the marine sponge Tethya aurantium and identification of their secondary metabolites.

Fungi associated with the marine sponge Tethya aurantium were isolated and identified by morphological criteria and phylogenetic analyses based on internal transcribed spacer (ITS) regions. They were evaluated with regard to their secondary metabolite profiles. Among the 81 isolates which were characterized, members of 21 genera were identified. Some genera like Acremonium, Aspergillus, Fusarium, Penicillium, Phoma, and Trichoderma are quite common, but we also isolated strains belonging to genera like Botryosphaeria, Epicoccum, Parasphaeosphaeria, and Tritirachium which have rarely been reported from sponges. Members affiliated to the genera Bartalinia and Volutella as well as to a presumably new Phoma species were first isolated from a sponge in this study. On the basis of their classification, strains were selected for analysis of their ability to produce natural products. In addition to a number of known compounds, several new natural products were identified. The scopularides and sorbifuranones have been described elsewhere. We have isolated four additional substances which have not been described so far. The new metabolite cillifuranone (1) was isolated from Penicillium chrysogenum strain LF066. The structure of cillifuranone (1) was elucidated based on 1D and 2D NMR analysis and turned out to be a previously postulated intermediate in sorbifuranone biosynthesis. Only minor antibiotic bioactivities of this compound were found so far.

Eutypoids B-E produced by a Penicillium sp. strain from the North Sea.

Crude extracts of the Penicillium sp. strain KF620 isolated from the North Sea showed antimicrobial activities against Xanthomonas campestris and Candida glabrata. Purification of the extracts led to the isolation of the new aromatic butenolides eutypoids B (1), C (2), D (3), and E (4). Their structures were elucidated by NMR spectroscopy and supported by HRESIMS and UV data. The antibacterial activity of the crude extracts was due to the presence of the known diketopiperazine fellutanine (cyclo(Trp-Trp)). The eutypoids were neither cytotoxic nor antibacterial, but inhibited the activity of glycogen synthase kinase-3ß.

New phylogenetic lineages of the Spirochaetes phylum associated with Clathrina species (Porifera).

Though spirochetes have been repeatedly found in marine sponges and other invertebrates, little attention has been paid to the specificity of this association. This study demonstrates that different geno-and morphotypes of spirochetes can reside within the same sponge individual and develop in considerable numbers. Specimens of the calcareous sponge Clathrina clathrus collected from the Adriatic Sea off Rovinj (Croatia) were found to harbor spirochete-like bacteria, which were characterized by scanning electron microscopy (SEM), 16S rRNA gene analysis, and catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH). Two novel spirochete sequence types related to the Brachyspiraceae could be retrieved. By use of specifically designed CARD-FISH probes, the C. clathrus-associated sequences could be assigned to a linear and a helical spirochete morphotype. Both were located within the sponge mesohyl and resembled the spirochete-like cells identified by SEM. In addition, from a Clathrina sp., most likely C. coriacea, that originated from Indonesian coastal waters, four different spirochete type sequences were recovered. Two of these also affiliated with the Brachyspiraceae, the other two were found associated with the Spirochaetaceae, one with the genera Borrelia and Cristispira.

Kiloniella laminariae gen. nov., sp. nov., an alphaproteobacterium from the marine macroalga Laminaria saccharina.

A novel alphaproteobacterium, strain LD81(T), was isolated from the marine macroalga Laminaria saccharina. The bacterium is mesophilic and shows a typical marine growth response. It is a chemoheterotrophic aerobe with the potential for denitrification. Growth optima are 25 degrees C, pH 5.5 and 3 % NaCl. Strain LD81(T) has a unique phylogenetic position, not fitting any of the known families of the Alphaproteobacteria. The 16S rRNA gene sequence revealed a distant relationship to species of several orders of the Alphaproteobacteria, with less than 90 % sequence similarity. Phylogenetically, strain LD81(T) is related to the type strains of Terasakiella pusilla (88.4 % 16S rRNA gene sequence similarity) and the three Thalassospira species (88.9-89.2 %). It forms a cluster with these bacteria and a novel as-yet undescribed isolate (KOPRI 13522; 96.6 % sequence similarity). Strain LD81(T) has a relatively low DNA G+C content (51.1 mol%) and, due to its distant phylogenetic position from all other alphaproteobacteria, strain LD81(T) (=NCIMB 14374(T) =JCM 14845(T)) is considered as the type strain of a novel species within a new genus, for which the name Kiloniella laminariae gen. nov., sp. nov. is proposed. The genus Kiloniella represents the type of the new family Kiloniellaceae fam. nov. and order Kiloniellales ord. nov.

A new member of the family Micromonosporaceae, Planosporangium flavigriseum gen. nov., sp. nov.

A novel actinomycete, designated strain YIM 46034(T), was isolated from an evergreen broadleaved forest at Menghai, in southern Yunnan Province, China. Phenotypic characterization and 16S rRNA gene sequence analysis indicated that the strain belonged to the family Micromonosporaceae. Strain YIM 46,034(T) showed more than 3 % 16S rRNA gene sequence divergence from recognized species of genera in the family Micromonosporaceae. Characteristic features of strain YIM 46,034(T) were the production of two types of spores, namely motile spores, which were formed in sporangia produced on substrate mycelia, and single globose spores, which were observed on short sporophores of the substrate mycelia. The cell wall contained meso-diaminopimelic acid, glycine, arabinose and xylose, which are characteristic components of cell-wall chemotype II of actinomycetes. Phosphatidylethanolamine was the major phospholipid (phospholipid type II). Based on morphological, chemotaxonomic, phenotypic and genetic characteristics, strain YIM 46,034(T) is considered to represent a novel species of a new genus in the family Micromonosporaceae, for which the name Planosporangium flavigriseum gen. nov., sp. nov. is proposed. The type strain of Planosporangium flavigriseum is YIM 46,034(T) (=CCTCC AA 205,013(T) =DSM 44,991(T)).

Scopularides A and B, cyclodepsipeptides from a marine sponge-derived fungus, Scopulariopsis brevicaulis.

Two novel cyclodepsipeptides, scopularides A and B, were found in the fungus Scopulariopsis brevicaulis, which was isolated from the marine sponge Tethya aurantium. In addition, the known fungal metabolite paxilline was identified. The structures of the scopularides were elucidated by NMR, MS, and chemical derivatization methods as cyclo-(3-hydroxy-4-methyldecanoyl-Gly-L-Val-D-Leu-L-Ala-L-Phe) and cyclo-(3-hydroxy-4-methyloctanoyl-Gly-L-Val-D-Leu-L-Ala-L-Phe) for scopularide A and B, respectively. Antibiotic activity against Gram-negative bacteria was absent and against Gram-positive bacteria was weak, but activity against several tumor cell lines was significant at 10 microg/mL.

Sponge-specific bacterial associations of the Mediterranean sponge Chondrilla nucula (Demospongiae, Tetractinomorpha).

A stable and specific bacterial community was shown to be associated with the Mediterranean sponge Chondrilla nucula. The associated bacterial communities were demonstrated to be highly similar for all studied specimens regardless of sampling time and geographical region. In addition, analysis of 16S rDNA clone libraries revealed three constantly C. nucula-associated bacterial phylotypes belonging to the Acidobacteria, the Gamma- and Deltaproteobacteria present in sponge specimens from two Mediterranean regions with distinct water masses (Ligurian Sea and Adriatic Sea). For the first time, candidate division TM7 bacteria were found in marine sponges. A major part (79%) of the C. nucula-derived 16S rDNA sequences were closely related to other sponge-associated bacteria. Phylogenetic analysis identified 14 16S rRNA gene sequence clusters, seven of which consisted of exclusively sponge-derived sequences, whereas the other seven clusters contained additional environmental sequences. This study adds to a growing database on the stability and variability of microbial consortia associated with marine sponges.

Spatial distribution of sponge-associated bacteria in the Mediterranean sponge Tethya aurantium.

The local distribution of the bacterial community associated with the marine sponge Tethya aurantium Pallas 1766 was studied. Distinct bacterial communities were found to inhabit the endosome and cortex. Clear differences in the associated bacterial populations were demonstrated by denaturing gradient gel electrophoresis (DGGE) and analysis of 16S rRNA gene clone libraries. Specifically associated phylotypes were identified for both regions: a new phylotype of Flexibacteria was recovered only from the sponge cortex, while Synechococcus species were present mainly in the sponge endosome. Light conduction via radiate spicule bundles conceivably facilitates the unusual association of Cyanobacteria with the sponge endosome. Furthermore, a new monophyletic cluster of sponge-derived 16S rRNA gene sequences related to the Betaproteobacteria was identified using analysis of 16S rRNA gene clone libraries. Members of this cluster were specifically associated with both cortex and endosome of T. aurantium.