Natural Products from Antarctic Colonial Ascidians of the Genera Aplidium and Synoicum: Variability and Defensive Role

Ascidians have developed multiple defensive strategies mostly related to physical, nutritional or chemical properties of the tunic. One of such is chemical defense based on secondary metabolites. We analyzed a series of colonial Antarctic ascidians from deep-water collections belonging to the genera Aplidium and Synoicum to evaluate the incidence of organic deterrents and their variability. The ether fractions from 15 samples including specimens of the species A. falklandicum, A. fuegiense, A. meridianum, A. millari and S. adareanum were subjected to feeding assays towards two relevant sympatric predators: the starfish Odontaster validus, and the amphipod Cheirimedon femoratus. All samples revealed repellency. Nonetheless, some colonies concentrated defensive chemicals in internal body-regions rather than in the tunic. Four ascidian-derived meroterpenoids, rossinones B and the three derivatives 2,3-epoxy-rossinone B, 3-epi-rossinone B, 5,6-epoxy-rossinone B, and the indole alkaloids meridianins AG, along with other minoritary meridianin compounds were isolated from several samples. Some purified metabolites were tested in feeding assays exhibiting potent unpalatabilities, thus revealing their role in predation avoidance. Ascidian extracts and purified compound-fractions were further assessed in antibacterial tests against a marine Antarctic bacterium. Only the meridianins showed inhibition activity, demonstrating a multifunctional defensive role. According to their occurrence in nature and within our colonial specimens, the possible origin of both types of metabolites is discussed.

Natural Products from Antarctic Colonial Ascidians of the Genera Aplidium and Synoicum: Variability and Defensive Role

Ascidians have developed multiple defensive strategies mostly related to physical, nutritional or chemical properties of the tunic. One of such is chemical defense based on secondary metabolites. We analyzed a series of colonial Antarctic ascidians from deep-water collections belonging to the genera Aplidium and Synoicum to evaluate the incidence of organic deterrents and their variability. The ether fractions from 15 samples including specimens of the species A. falklandicum, A. fuegiense, A. meridianum, A. millari and S. adareanum were subjected to feeding assays towards two relevant sympatric predators: the starfish Odontaster validus, and the amphipod Cheirimedon femoratus. All samples revealed repellency. Nonetheless, some colonies concentrated defensive chemicals in internal body-regions rather than in the tunic. Four ascidian-derived meroterpenoids, rossinones B and the three derivatives 2,3-epoxy-rossinone B, 3-epi-rossinone B, 5,6-epoxy-rossinone B, and the indole alkaloids meridianins AG, along with other minoritary meridianin compounds were isolated from several samples. Some purified metabolites were tested in feeding assays exhibiting potent unpalatabilities, thus revealing their role in predation avoidance. Ascidian extracts and purified compound-fractions were further assessed in antibacterial tests against a marine Antarctic bacterium. Only the meridianins showed inhibition activity, demonstrating a multifunctional defensive role. According to their occurrence in nature and within our colonial specimens, the possible origin of both types of metabolites is discussed.

Natural Products from Antarctic Colonial Ascidians of the Genera Aplidium and Synoicum: Variability and Defensive Role

Ascidians have developed multiple defensive strategies mostly related to physical, nutritional or chemical properties of the tunic. One of such is chemical defense based on secondary metabolites. We analyzed a series of colonial Antarctic ascidians from deep-water collections belonging to the genera Aplidium and Synoicum to evaluate the incidence of organic deterrents and their variability. The ether fractions from 15 samples including specimens of the species A. falklandicum, A. fuegiense, A. meridianum, A. millari and S. adareanum were subjected to feeding assays towards two relevant sympatric predators: the starfish Odontaster validus, and the amphipod Cheirimedon femoratus. All samples revealed repellency. Nonetheless, some colonies concentrated defensive chemicals in internal body-regions rather than in the tunic. Four ascidian-derived meroterpenoids, rossinones B and the three derivatives 2,3-epoxy-rossinone B, 3-epi-rossinone B, 5,6-epoxy-rossinone B, and the indole alkaloids meridianins A-G, along with other minoritary meridianin compounds were isolated from several samples. Some purified metabolites were tested in feeding assays exhibiting potent unpalatabilities, thus revealing their role in predation avoidance. Ascidian extracts and purified compound-fractions were further assessed in antibacterial tests against a marine Antarctic bacterium. Only the meridianins showed inhibition activity, demonstrating a multifunctional defensive role. According to their occurrence in nature and within our colonial specimens, the possible origin of both types of metabolites is discussed.

Spatial and temporal variation of natural toxicity in cnidarians, bryozoans and tunicates in Mediterranean caves

The natural toxicity of cnidarians, bryozoans and tunicates in two caves was assessed using the Microtox® technique in spring and autumn. One cave was located in the Cabrera Archipelago (Balearic Islands) and the other in the Medes Islands (Catalan littoral). The organisms analysed were good representatives of the coverage of each Phylum in the communities; however, these Phyla are less abundant than sponges which are the dominant group in these caves. Seventy-one percent of the species of cnidarians and bryozoans analysed were toxic in one of the caves, communities or seasons, which indicates the relevance of bioactive species in these groups. The tunicate Lissoclinum perforatum was the most toxic species. Although all three Phyla had some highly toxic species, a common pattern that related the caves, communities and seasons was not found. Seasonal variation of toxicity in cnidarians and bryozoans was higher in the Cabrera than in the Medes cave. Moreover, variation in toxicity either between communities or between seasons was a common trait for most cnidarians and bryozoans, whereas tunicates remained toxic throughout communities and seasons.

Tough adults, frail babies: an analysis of stress sensitivity across early life-history stages of widely introduced marine invertebrates

All ontogenetic stages of a life cycle are exposed to environmental conditions so that population persistence depends on the performance of both adults and offspring. Most studies analysing the influence of abiotic conditions on species performance have focussed on adults, while studies covering early life-history stages remain rare. We investigated the responses of early stages of two widely introduced ascidians, Styela plicata and Microcosmus squamiger, to different abiotic conditions. Stressors mimicked conditions in the habitats where both species can be found in their distributional ranges and responses were related to the selection potential of their populations by analysing their genetic diversity. Four developmental stages (egg fertilisation, larval development, settlement, metamorphosis) were studied after exposure to high temperature (30°C), low salinities (26 and 22 ) and high copper concentrations (25, 50 and 100 µg/L). Although most stressors effectively led to failure of complete development (fertilisation through metamorphosis), fertilisation and larval development were the most sensitive stages. All the studied stressors affected the development of both species, though responses differed with stage and stressor. S. plicata was overall more resistant to copper, and some stages of M. squamiger to low salinities. No relationship was found between parental genetic composition and responses to stressors. We conclude that successful development can be prevented at several life-history stages, and therefore, it is essential to consider multiple stages when assessing species' abilities to tolerate stress. Moreover, we found that early development of these species cannot be completed under conditions prevailing where adults live. These populations must therefore recruit from elsewhere or reproduce during temporal windows of more benign conditions. Alternatively, novel strategies or behaviours that increase overall reproductive success might be responsible for ensuring population survival.

Recent advances in lamellarin alkaloids: isolation, synthesis and activity

Lamellarins are a large family of marine alkaloids with potential anticancer activity that have been isolated from diverse marine organisms, mainly ascidians and sponges. All lamellarins feature a 3,4-diarylpyrrole system. Pentacyclic lamellarins, whose polyheterocyclic system has a pyrrole core, are the most active compounds. Some of these alkaloids are potently cytotoxic to various tumor cell lines. To date, Lam-D and Lam-H have been identified as lead compounds for the inhibition of topoisomerase I and HIV-1 integrase, respectively nuclear enzymes which are over-expressed in deregulation disorders. Moreover,these compounds have been reported for their efficacy in treatment of multi-drug resistant (MDR) tumors cells without mediated drug efflux, as well as their immunomodulatory activity and selectivity towards melanoma cell lines. This article is an overview of recent literature on lamellarins, encompassing their isolation, recent synthetic strategies for their total synthesis, the preparation of their analogs, studies on their mechanisms of action, and their structure-activity relationships (SAR).

Defensive Metabolites from Antarctic Invertebrates: Does Energetic Content Interfere with Feeding Repellence?

Many bioactive products from benthic invertebrates mediating ecological interactions have proved to reduce predation, but their mechanisms of action, and their molecular identities, are usually unknown. It was suggested, yet scarcely investigated, that nutritional quality interferes with defensive metabolites. This means that antifeedants would be less effective when combined with energetically rich prey, and that higher amounts of defensive compounds would be needed for predator avoidance. We evaluated the effects of five types of repellents obtained from Antarctic invertebrates, in combination with diets of different energetic values. The compounds came from soft corals, ascidians and hexactinellid sponges; they included wax esters, alkaloids, a meroterpenoid, a steroid, and the recently described organic acid, glassponsine. Feeding repellency was tested through preference assays by preparing diets (alginate pearls) combining different energetic content and inorganic material. Experimental diets contained various concentrations of each repellent product, and were offered along with control compound-free pearls, to the Antarctic omnivore amphipod Cheirimedon femoratus. Meridianin alkaloids were the most active repellents, and wax esters were the least active when combined with foods of distinct energetic content. Our data show that levels of repellency vary for each compound, and that they perform differently when mixed with distinct assay foods. The natural products that interacted the most with energetic content were those occurring in nature at higher concentrations. The bioactivity of the remaining metabolites tested was found to depend on a threshold concentration, enough to elicit feeding repellence, independently from nutritional quality.

The whereabouts of an ancient wonderer: Global phylogeography of the solitary ascidian Styela plicata

Genetic tools have greatly aided in tracing the sources and colonization history of introduced species. However, recurrent introductions and repeated shuffling of populations may have blurred some of the genetic signals left by ancient introductions. Styela plicata is a solitary ascidian distributed worldwide. Although its origin remains unclear, this species is believed to have spread worldwide by travelling on ship's hulls. The goals of this study were to infer the genetic structure and global phylogeography of S. plicata and to look for present-day and historical genetic patterns. Two genetic markers were used: a fragment of the mitochondrial gene Cytochrome Oxidase subunit I (COI) and a fragment of the nuclear gene Adenine Nucleotide Transporter/ADP-ATP Translocase (ANT). A total of 368 individuals for COI and 315 for ANT were sequenced from 17 locations worldwide. The levels of gene diversity were moderate for COI to high for ANT. The Mediterranean populations showed the least diversity and allelic richness for both markers, while the Indian, Atlantic and Pacific Oceans had the highest gene and nucleotide diversities. Network and phylogenetic analyses with COI and ANT revealed two groups of alleles separated by 15 and 4 mutational steps, respectively. The existence of different lineages suggested an ancient population split. However, the geographic distributions of these groups did not show any consistent pattern, indicating different phylogeographic histories for each gene. Genetic divergence was significant for many population-pairs irrespective of the geographic distance among them. Stochastic introduction events are reflected in the uneven distribution of COI and ANT allele frequencies and groups among many populations. Our results confirmed that S. plicata has been present in all studied oceans for a long time, and that recurrent colonization events and occasional shuffling among populations have determined the actual genetic structure of this species.

Cyanobacterial diversity and a new Acaryochloris-like symbiont from Bahamian sea-squirts

Symbiotic interactions between ascidians (sea-squirts) and microbes are poorly understood. Here we characterized the cyanobacteria in the tissues of 8 distinct didemnid taxa from shallow-water marine habitats in the Bahamas Islands by sequencing a fragment of the cyanobacterial 16S rRNA gene and the entire 16S-23S rRNA internal transcribed spacer region (ITS) and by examining symbiont morphology with transmission electron (TEM) and confocal microscopy (CM). As described previously for other species, Trididemnum spp. mostly contained symbionts associated with the Prochloron-Synechocystis group. However, sequence analysis of the symbionts in Lissoclinum revealed two unique clades. The first contained a novel cyanobacterial clade, while the second clade was closely associated with Acaryochloris marina. CM revealed the presence of chlorophyll d (chl d) and phycobiliproteins (PBPs) within these symbiont cells, as is characteristic of Acaryochloris species. The presence of symbionts was also observed by TEM inside the tunic of both the adult and larvae of L. fragile, indicating vertical transmission to progeny. Based on molecular phylogenetic and microscopic analyses, Candidatus Acaryochloris bahamiensis nov. sp. is proposed for this symbiotic cyanobacterium. Our results support the hypothesis that photosymbiont communities in ascidians are structured by host phylogeny, but in some cases, also by sampling location.