Composition and temporal variation of the algal assemblage associated with the haplosclerid sponge Haliclona indistincta (Bowerbank)

Although interactions between seaweeds and sponges have been studied in detail, general information concerning the whole epibiontic algal assemblage associated with a sponge species is virtually non-existent. We present here the first study in which the macroalgal community associated with a sponge, Haliclona indistincta (Bowerbank), was examined in detail. In the period October 2009-September 2010, the seaweed assemblage epibiontic on H. indistincta at a site of the Irish West coast was composed of 66 algal taxa (48 red algae, 7 green algae, 11 brown algae). The red algae Gelidium spinosum and Rhodothamniella floridula were the only epibionts associated with H. indistincta for the whole annual cycle. Most of the algal epibionts were filamentous species, which colonized the surface of the sponge and did not penetrate deeply into it. The algal assemblage was most abundant and species-diverse in the period late winter-spring; multivariate analyses revealed a significant variation of the community on the temporal scale of season and sampling date (weeks to months). The results indicate that the algal communities associated with sponges may be very diverse, showing that this type of assemblage deserves further detailed studies. © 2012 Elsevier B.V.

Comparison of two marine sponge-associated Penicillium strains DQ25 and SC10: differences in secondary metabolites and their bioactivities

Two strains of Penicillium, DQ25 and SC10, isolated from marine sponge Haliclona angulata (Bowerbank) and Hymeniacidon sp. respectively, were subjected to stationary cultivation under GYP medium for 30 days. The fermentation extracts were undergone bioactivities assays against human pathogens, phytopathogenic fungi and brine shrimp (Artemia salina). Bioassays-guided compounds isolation was performed by Silica gel columns and Sephadex LH-20 chromatography. Spectroscopic methods were used to structures elucidation of the compounds. Results showed the activities of secondary metabolites of strain DQ25 were generally stronger than that of strain SC10. Major bioactive molecules isolated from strain DQ25 were a 1,4-naphthoquinone derivative and an unidentified alkaloid. The two components were not isolated from the extract of strain SC10. ITS sequences revealed that these two species have the greatest similarity with Penicillium vinaceum and Penicillium granulatum respectively.

Biodiscovery of natural products from microbes associated with Irish coastal sponges

Marine sponges have been an abundant source of new metabolites in recent years. The symbiotic association between the bacteria and the sponge has enabled scientists to access the bacterial diversity present within the bacterial/sponge ecosystem. This study has focussed on accessing the bacterial diversity in two Irish coastal marine sponges, namely Amphilectus fucorum and Eurypon major. A novel species from the genus Aquimarina has been isolated from the sponge Amphilectus fucorum. The study has also resulted in the identification of an α–Proteobacteria, Pseudovibrio sp. as a potential producer of antibiotics. Thus a targeted based approach to specifically cultivate Pseudovibrio sp. may prove useful for the development of new metabolites from this particular genus. Bacterial isolates from the marine sponge Haliclona simulans were screened for anti–fungal activity and one isolate namely Streptomyces sp. SM8 displayed activity against all five fungal strains tested. The strain was also tested for anti–bacterial activity and it showed activity against both against B. subtilis and P. aeruginosa. Hence a combinatorial approach involving both biochemical and genomic approaches were employed in an attempt to identify the bioactive compounds with these activities which were being produced by this strain. Culture broths from Streptomyces sp. SM8 were extracted and purified by various techniques such as reverse–phase HPLC, MPLC and ash chromatography. Anti–bacterial activity was observed in a fraction which contained a hydroxylated saturated fatty acid and also another compound with a m/z 227 but further structural elucidation of these compounds proved unsuccessful. The anti–fungal fractions from SM8 were shown to contain antimycin–like compounds, with some of these compounds having different retention times from that of an antimycin standard. A high–throughput assay was developed to screen for novel calcineurin inhibitors using yeast as a model system and three putative bacterial extracts were found to be positive using this screen. One of these extracts from SM8 was subsequently analysed using NMR and the calcineurin inhibition activity was con rmed to belong to a butenolide type compound. A H. simulans metagenomic library was also screened using the novel calcineurin inhibitor high–throughput assay system and eight clones displaying putative calcineurin inhibitory activity were detected. The clone which displayed the best inhibitory activity was subsequently sequenced and following the use of other genetic based approaches it became clear that the inhibition was being caused by a hypothetical protein with similarity to a hypothetical Na+/Ca2+ exchanger protein. The Streptomyces sp. SM8 genome was sequenced from a fragment library using Roche 454 pyrosequencing technology to identify potential secondary metabolism clusters. The draft genome was annotated by IMG/ER using the Prodigal pipeline. The Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession AMPN00000000. The genome contains genes which appear to encode for several polyketide synthases (PKS), non–ribosomal peptide synthetases (NRPS), terpene and siderophore biosynthesis and ribosomal peptides. Transcriptional analyses led to the identification of three hybrid clusters of which one is predicted to be involved in the synthesis of antimycin, while the functions of the others are as yet unknown. Two NRPS clusters were also identified, of which one may be involved in gramicidin biosynthesis and the function of the other is unknown. A Streptomyces sp. SM8 NRPS antC gene knockout was constructed and extracts from the strain were shown to possess a mild anti–fungal activity when compared to the SM8 wild–type. Subsequent LCMS analysis of antC mutant extracts confirmed the absence of the antimycin in the extract proving that the observed anti–fungal activity may involve metabolite(s) other than antimycin. Anti–bacterial activity in the antC gene knockout strain against P. aeruginosa was reduced when compared to the SM8 wild–type indicating that antimycin may be contributing to the observed anti–bacterial activity in addition to the metabolite(s) already identified during the chemical analyses. This is the first report of antimycins exhibiting anti–bacterial activity against P. aeruginosa. One of the hybrid clusters potentially involved in secondary metabolism in SM8 that displayed high and consistent levels of gene–expression in RNA studies was analysed in an attempt to identify the metabolite being produced by the pathway. A number of unusual features were observed following bioinformatics analysis of the gene sequence of the cluster, including a formylation domain within the NRPS cluster which may add a formyl group to the growing chain. Another unusual feature is the lack of AT domains on two of the PKS modules. Other unusual features observed in this cluster is the lack of a KR domain in module 3 of the cluster and an aminotransferase domain in module 4 for which no clear role has been hypothesised.

Antimicrobial activities and diversity of sponge derived microbes

In this study, marine sponges collected in Irish waters were analysed for their associated microbiota. Of the approximately 240 bacterial isolates obtained from two sponges several showed antimicrobial activity; among them members of genera which have rarely been shown to produce antimicrobial compounds. Differences observed from the sponge-derived groups of isolates in terms of bioactivity suggests that S. carnosus isolates may be a better source of antibacterial compounds, while Leucosolenia sp. isolates appear to be a better source of antifungal compounds. More than 60% of fungal isolates obtained from 12 sponge samples proved to be bioactive. One of the isolates, which was closely related to Fusarium oxysporum and showed activity against bacteria and fungi, was investigated for its secondary metabolite genes. At least 5 different NRPS genes, with a sequence similarity as low as 50 % to known genes, were identified highlighting the likelihood that this isolate may be capable of producing novel secondary metabolites. A Micromonospora sp. was isolated from a Haliclona simulans sample collected in Irish waters. The isolate inhibited the growth of Gram positive bacterial test strains in three different antimicrobial assays. Employing preparative layer chromatography the compound responsible for the bioactivity could be isolated. According to LC-MS andNMR data the bioactive compound could indeed be novel. Finally, two deep water sponges were shown to host a remarkably different bacterial and archaeal diversity by application of 454 Pyrosequencing. The L. diversichela –proteobacterial community was dominated by a single ƴ-proteobacterial bacterium whereas the S. normani sample hosted a largely sponge specific microbial community, even more diverse than has been previously reported for shallow water sponges. Organisms potentially involved in nitrification, sulphate reduction and secondary metabolite production were found to be spatially distributed in the sponge. Furthermore, a deep sea specific population was implied.

Exploiting the bioactive potential of marine sponge microbiota

Endospore-forming bacteria are often isolated from different marine sponges, but their abundance varies, and they are frequently missed by culture-independent studies. Within endospore-formers, Bacillus are renowned for the production of antimicrobials and other compounds of medical and industrial importance. Although this group has been well studied in many different environments, very little is known about the actual diversity and properties of sporeformers associated with marine sponges. Identification of the endospore-forming bacteria associated with the marine sponges; Haliclona simulans, Amphilectus fucorum and Cliona celata, has uncovered an abundant and diverse microbial population composed of Bacillus, Paenibacillus, Solibacillus, Halobacillus and Viridibacillus species. This diversity appears to be overlooked by other non-targeted approaches where spore-formers are masked by more dominant species within the ecosystem. In addition to the identification of two antibiotic resistant plasmids, this bank of sporeformers produce a range of bioactive compounds. New antimicrobial compounds are urgently needed to combat the spread of multidrug resistant pathogens, as few new options are entering the drug discovery pipelines for clinical trials. Based on the results of this project, endospore-formers associated with marine sponges may hold the answer. The power of coupling functional based assays with genomic approaches has enabled us to identify a novel class 1 lantibiotic, subtilomycin, which is active against several clinically relevant pathogens. Subtilomycin is encoded in the genomes of all the marine sponge B. subtilis isolates analysed. They cluster together phylogenetically and form a distinct group from other sequenced B. subtilis strains. Regardless of its potential clinical relevance, subtilomycin may be providing these strains with a specific competitive advantage(s) within the stringent confines of the marine sponge environment. This work has outlined the industrial and biotechnological potential of marine sponge endospore-formers which appear to produce a cocktail of bioactive compounds. Genome sequencing of specific marine sponge isolates highlighted the importance of mining extreme environments and habitats for new lead compounds with potential therapeutic applications.

A systematic revision of Goniosomatinae (Arachnida : Opiliones : Gonyleptidae), with a cladistic analysis and biogeographical notes

Goniosomatine harvestmen have strongly armed pedipalps, generally large bodies and, commonly, very long legs (sometimes more than 20 cm), and are distributed in the Brazilian Atlantic forest, from southern Bahia to Santa Catarina. Since they are conspicuous animals and individuals of some species tend to concentrate in caves (and also under rock boulders), they have been (and still are) the target of several studies, especially those focusing on reproductive and defensive behavior, population ecology, physiology, chromosomes, etc. In spite of their importance for biological studies (some species constitute important and frequently used models for these studies), the taxonomy of Goniosomatinae has faced some problems, including misidentification, a large number of undescribed species and the lack of a phylogenetic hypothesis for the relationships among its species (which would allow evolutionary studies to be made). The last taxonomic changes in the subfamily were made 60 years ago. Considering a taxonomic revision and cladistic analysis of the subfamily to be of paramount importance, the main scope of the present paper is to provide a cladistic analysis and taxonomic revision of the species of Goniosomatinae and a new arrangement of genera (and species). The main taxonomic changes are given as follows. Six genera are recognised within the subfamily: Goniosoma; the newly described genus Pyatan; the reestablished genera Serracutisoma, Heteromitobates and Mitogoniella; and Acutisoma. New generic synonyms include: Glyptogoniosoma = Goniosomella = Lyogoniosoma = Metalyogoniosoma = Xulapona = Goniosoma, Acutisomelloides = Pygosomoides = Spelaeosoma = Serracutisoma; and Acutisomella = Heteromitobates. Newly described species include: Goniosoma capixaba; G. apoain; Pyatan insperatum DaSilva, Stefanini-Jim & Gnaspini; Serracutisoma pseudovarium; S. fritzmuelleri; S. guaricana; Heteromitobates anarchus; H. harlequin; H. alienus; Mitogoniella taquara; M. unicornis; and Acutisoma coriaceum. New combinations include: Goniosoma macracanthum (Mello-Leitao, 1922); G. unicolor (Mello-Leitao, 1932); G. carum (Mello-Leitao, 1936); Serracutisoma proximum (Mello-Leitao, 1922); S. banhadoae (Soares & Soares, 1947); S. molle (Mello-Leitao, 1933); S. thalassinum (Simon, 1879); S. catarina (Machado, Pinto-da-Rocha & Ramires, 2002); S. inerme (Mello-Leitao, 1927); S. spelaeum (MelloLeitao, 1933); Heteromitobates inscriptus (Mello-Leitao, 1922); H. albiscriptus (Mello-Leitao, 1932); Mitogoniella modesta (Perty, 1833); and M. badia (Koch, 1839). Reestablished combinations include: Mitogoniella indistincta MelloLeitao, 1936 and Acutisoma longipes Roewer, 1913. New speci. c synonyms include: Acutisomella cryptoleuca = Acutisomella intermedia = Goniosoma junceum = Goniosoma patruele = Goniosoma xanthophthalmum = Metalyogoniosoma unum = Goniosoma varium, Goniosoma geniculatum = Goniosoma venustum; Goniosomella perlata = Progoniosoma minense = Goniosoma vatrax, Glyptogoniosoma perditum = Progoniosoma cruciferum = Progoniosoma tijuca = Goniosoma dentipes; Leitaoius iguapensis = Leitaoius viridifrons = Serracutisoma proximum; Acutisoma marumbicola = Acutisoma patens = Serracutisoma thalassinum; Progoniosoma tetrasetae = Serracutisoma inerme; and Acutisoma monticola = Leitaoius nitidissimus = Leitaoius xanthomus = Mitogoniella mutila = Acutisoma longipes. The following species are considered species inquirenda: Goniosoma lepidum Gervais, 1844; G. monacanthum Gervais, 1844; G. obscurum Perty, 1833; G. versicolor Perty, 1833; and Mitogoniella badia (Koch, 1839). The monotpic genus Goniosomoides Mello-Leitao, 1932 (and its species, G. viridans Mello-Leitao, 1932) is removed from Goniosomatinae and considered incertae sedis.

Sponges as substrata and early life history of the tubulariid Zyzzyzus warreni (Cnidaria: Hydrozoa) in the Sao Sebastiao Channel, Brazil

The hydroid Zyzzyzus warreni is usually found in shallow coastal waters forming aggregations of solitary polyps embedded in demosponges. Early life history transformations and settlement responses by the actinulae of this hydroid were studied in the laboratory using 13 species of sponges and 2 species of algae collected in the Sao Sebastiao Channel (Brazil) as substrata. The absence of oral tentacles and mouth in the actinulae and early events of metamorphosis suggest that these larvae are unable to spend long periods in the plankton and attach quickly near conspecifics when a preferred substratum is encountered. The time required for settlement and the number of elicited settlings indicated four settlement responses: (a) frequent and short-time settlement, in actinulae exposed to Halichondria cebimarensis, Mycale angulosa, M. aff. americana, M. laxissima (skeleton) and Tedania ignis; (b) moderate and delayed settlement, in actinulae exposed to Aplysina caissara, A. fulva, Haliclona melana and M. microsigmatosa; (c) no settlement, in actinulae exposed to Suberites aurantiacus and to the algae Hypnea musciformis and Sargassum cymosum; and (d) lethal response, in actinulae exposed to Amphimedon viridis, Aplysilla rosea, Dragmacidon reticulatum and M. laxissima. These responses indicate a considerable degree of species discrimination by the actinulae and are consistent with substrata used by the hydroid in the natural environment.

Counts of benthos communities in Kongsfjorden in 2009

In order to reveal the structure of the sparsely known deeper sublittoral hard bottom communities of glacial Kongsfjorden, the macroepibenthos from six depth zones (30-200 m) was analysed. A total of 180 still images derived from 6-h video recorded at the Kongsfjordneset remotely operated vehicle station were assessed quantitatively. Overall 27 mainly suspension-feeding species/taxa were observed. Of these, two-thirds have an arcto-boreal distribution, while the remainder are cosmopolitan. The overall mean epibenthos abundance was 33 ind./m**2 with maximum values at 150 m depth (97.9 ind./m**2). The majority of the taxa inhabited the entire depth range. Encrusting red algae, an unidentified sponge and the sea anemone Urticina eques, characterized the assemblage of the shallow zone. The sea anemones Hormathia spp. were important below 30 m, the Serpulid polychaete Protula tubularia was characteristic for the community below 50 m and the demosponge Haliclona sp. was a key taxon between 100 and 200 m depth. Cluster analysis and non-metrical multidimensional scaling based on abundance data showed differences between the assemblages along the bathymetric gradient, but only in the shallower depths in relation to the substratum surface incline. As surface and tidal current impacts attenuate with increasing depth, there is a gradual trend from robust key species towards more fragile ones (i.e. P. tubularia), in line with the 'Physical control hypothesis'.

Succession of benthic hard-bottom communities in the shallow sublitoral of Comau Fjord, Chile

Succession was already studied over decades. The present thesis investigated the succession on hard substrate at two different study sites within the fjord Comau, Chile. Nine plates were installed at both sites (mouth of fjord and inner fjord) and photographed over three years. Additionally the natural community was recorded and a ground truthing was carried out to verify the analyzed species. Respectively at both sites over 50 different species were identified. Abundance data decreased with only one exception continuously, whereas the percentage cover increased. But the communities on the recruitment plates do still not reach the community structure of the natural environment. The present data showed that the hard-bottom succession in the fjord Comau is best described by the TOLERANCE MODEL (Connell & Slatyer, 1977). An important species of the natural community is the stony coral Desmophyllum dianthus, which normally (outside the fjord) grows beneath 1000 m water depth. The results of this work indicate that the mature community is not reached after 36 months.