Ascidian Species Research Articles

Tunicates Illuminate the Enigmatic Evolution of Chordate Metallothioneins by Gene Gains and Losses, Independent Modular Expansions, and Functional Convergences.

To investigate novel patterns and processes of protein evolution, we have focused in the metallothioneins (MTs), a singular group of metal-binding, cysteine-rich proteins that, due to their high degree of sequence diversity, still represents a “black hole” in Evolutionary Biology. We have identified and analyzed more than 160 new MTs in nonvertebrate chordates (especially in 37 species of ascidians, 4 thaliaceans, and 3 appendicularians) showing that prototypic tunicate MTs are mono-modular proteins with a pervasive preference for cadmium ions, whereas vertebrate and cephalochordate MTs are bimodular proteins with diverse metal preferences. These structural and functional differences imply a complex evolutionary history of chordate MTs—including de novo emergence of genes and domains, processes of convergent evolution, events of gene gains and losses, and recurrent amplifications of functional domains—that would stand for an unprecedented case in the field of protein evolution.

Substrate Selection of Ascidian Larva: Wettability and Nano-Structures

Ascidians are marine sessile chordates that comprise one of the major benthic animal groups in marine ecosystems. They sometimes cause biofouling problems on artificial structures underwater, and non-indigenous, invasive ascidian species can potentially and seriously alter native faunal communities. Ascidian larvae are usually tadpole-shaped, negatively phototactic, and adhere on substrates by secreting a glue from their adhesive organs. Although larvae often prefer hydrophobic surfaces, such as a silicone rubber, for settlement, hydrophobic materials are often used to reduce occurrence of fouling organisms on artificial structures. This inconsistency may indicate that an attractive surface for larvae is not always suitable for settlement. Micro-scale structures or roughness may enhance the settlement of ascidian larvae, but settlement is significantly reduced by a nano-scale nipple array (or moth-eye structure), suggesting functional properties of similar structures found on the body surfaces of various invertebrates. The substrate preferences of larvae should be one of the important bases in considering measures against biofouling, and this review also discusses the potential uses of materials to safely reduce the impacts of invasive species.

Evaluation of lipid profiles in three species of ascidians using UPLC-ESI-Q-TOF-MS-based lipidomic study

Ascidians are excellent, yet underused, marine sources of unique bioactive compounds of high nutritive content. However, reports regarding the lipid composition of ascidians are rare. In this study, using ultra-high-performance liquid chromatography-electrospray ionization-quadrupole time-of-flight mass spectrometry, we aimed to characterize the lipid profile of the tunics and inner body tissues of three species of ascidians, including Ciona intestinalis, Halocynthia roretzi, and Styela clava. We identified over 245 molecular species from 13 major lipid subclasses; glycerophospholipids (GP) and glycerolipids were the dominant lipid components in these three ascidian species (accounted for 66.30–90.60% of total lipids). Importantly, GP enriched in polyunsaturated fatty acids mainly existed in the inner body tissues of ascidians, which accounted for 18.17–32.47% of total lipids. Considering the high level of GP, we proposed that ascidians can be potentially used as health-promoting food for humans.

Current status (as of end of 2020) of marine alien species in Turkey

The 2020’s update of marine alien species list from Turkey yielded a total of 539 species belonging to 18 taxonomic groups, 404 of which have become established in the region and 135 species are casual. A total of 185 new alien species have been added to the list since the previous update of 2011. The present compilation includes reports of an ascidian species (Rhodosoma turcicum) new to the marine fauna of Turkey and range extensions of six species. Among the established species, 105 species have invasive characters at least in one zoogeographic region, comprising 19% of all alien species. Mollusca ranked first in terms of the number of species (123 species), followed by Foraminifera (91 species), Pisces (80 species) and Arthropoda (79 species). The number of alien species found in seas surrounding Turkey ranged from 28 (Black Sea) to 413 (Levantine Sea). The vectoral importance of the Suez Canal diminishes when moving from south to north, accounting for 72% of species introductions in the Levantine Sea vs. only 11% of species introductions in the Black Sea. Most alien species on the coasts of Turkey were originated from the Red Sea (58%), due to the proximity of the country to the Suez Canal. Shipping activities transported 39% of alien species, mainly from the Indo-Pacific area (20%) and the Atlantic Ocean (10%). Misidentified species (such as Pterois volitans, Trachurus declivis, etc.) and species those classified as questionable or cryptogenic were omitted from the list based on new data gathered in the last decade and expert judgements. The documented impacts of invasive species on socio-economy, biodiversity and human health in the last decade as well as the legislation and management backgrounds against alien species in Turkey are presented.

Phylogenomic and morphological relationships among the botryllid ascidians (Subphylum Tunicata, Class Ascidiacea, Family Styelidae)

Ascidians (Phylum Chordata, Class Ascidiacea) are a large group of invertebrates which occupy a central role in the ecology of marine benthic communities. Many ascidian species have become successfully introduced around the world via anthropogenic vectors. The botryllid ascidians (Order Stolidobranchia, Family Styelidae) are a group of 53 colonial species, several of which are widespread throughout temperate or tropical and subtropical waters. However, the systematics and biology of this group of ascidians is not well-understood. To provide a systematic framework for this group, we have constructed a well-resolved phylogenomic tree using 200 novel loci and 55 specimens. A Principal Components Analysis of all species described in the literature using 31 taxonomic characteristics revealed that some species occupy a unique morphological space and can be easily identified using characteristics of adult colonies. For other species, additional information such as larval or life history characteristics may be required for taxonomic discrimination. Molecular barcodes are critical for guiding the delineation of morphologically similar species in this group.

Diversity and abundance of native and non-native ascidians in Puerto Rican harbors and marinas

Ascidians are an ideal taxon to study invasion processes: they require anthropogenic introduction vectors for long-distance dispersal, are easy to collect and monitor, and are abundant on artificial substrates. In March 2019 we surveyed 11 harbors around Puerto Rico and recorded 47 ascidian species. Eleven of these were only identified to the genus level or above based on morphological or genetic characterization. The remaining 36 species were classified as: 11 introduced (7 with worldwide distributions), 13 cryptogenic, and 12 native. We report the occurrence of Phallusia cf. philippinensis in the Atlantic for the first time. Ascidian community structure did not differ significantly across geographic locations and distances between marinas, while marina size had a significant effect on species richness and composition. Stakeholder involvement and periodic monitoring efforts are essential to detect the arrival of new species and the spread of already introduced ones to natural habitats.

Glycoside hydrolases from the tunics of two Antarctic ascidians (Ascidia challengeri and Pyura bouvetensis) and the tropical species Phallusia nigra

Tunicates are the only metazoans capable of synthesising cellulose, which is present in the tunic. Cellulases (endo and exoglucanases) that can degrade or remodel cellulose might be necessary during the growth phase and during scarring after injury to the tunic. However, the occurrence of these enzymes in the tunic has not been reported. Hence, the aim of the study was to investigate the presence of cellulases as well as other glycoside hydrolases, such as xyloglucanases and mannanases, in the tunic of two Antarctic ascidian species, Ascidia challengeri and Pyura bouvetensis, and in the tropical species Phallusia nigra. Endoglucanases and xyloglucanase were found in all three species. Galactomannanases were detected in P. nigra and P. bouvetensis but were not found in A. challengeri. Cellulases that can degrade Avicel were found in P. bouvetensis. To the best of our knowledge, this is the first report detailing the presence of these enzymes in the tunic of ascidians. In A. challengeri, the optimal temperature for the endoglucanase activity was 9 °C, higher than their habitat temperature, but within the temperature tolerance of the species, while that of P. nigra was at 25–30 °C, which is close to the maximum temperature of its habitat. The temperature at which the endoglucanase has optimal activity is discussed as a proxy to the limit of thermal tolerance of the species that express this enzyme.

Cryptic genetic lineages of a colonial ascidian host distinct microbiomes

AbstractColour variation in marine invertebrates is typically attributed to intraspecific variability; however, the use of molecular tools to investigate its genetic basis has uncovered many instances of cryptic speciation. The colonial ascidian Distaplia bermudensis occurs in a wide array of colour morphs, ranging from all white to a dark blue tunic with yellow siphon apertures; yet careful observations of zooid and larval morphologies have not revealed any consistent difference among these colour morphs. In this study, we sampled the 5 main colour morph groupings of D. bermudensis from Wilmington, North Carolina (USA) and sequenced fragments of the mitochondrial cytochrome c oxidase subunit I (COI) and the nuclear 18S rRNA genes. Analyses of both genes revealed the presence of two distinct genetic lineages that corresponded to unique colour morphology groupings. We further characterized symbiotic microbial communities for four samples from each lineage by high‐throughput sequencing of partial 16S rRNA genes. Each genetic lineage also harboured unique symbiont communities that differed significantly in diversity and structure. Thus, the two genetic lineages detected herein corresponded to unique colour morphs and microbial communities, indicating that two distinct species coexist under the same scientific name. Further, our study reports the utility of symbiont profiling in resolving the taxonomic status of phenotypically variable ascidian species.

Antarctic ascidians under increasing sedimentation: Physiological thresholds and ecosystem hysteresis

Glacier melting sediment inputs affect coastal ecosystems on the Antarctic Peninsula. In Potter Cove (South Shetland Islands, Antarctica), the shift from an “ascidian dominated” to a “mixed” assemblage has been linked to sedimentation. However, in recently described newly ice-free areas ascidians became dominant in spite of total suspended particulate matter (TSPM) concentrations, which are the highest measured in Potter Cove. Here, we compared the gut content and energy reserve of three ascidian species at three stations under different TSPM regimes. All analysed species had a higher gut content with lower %OM at these newly areas. A theoretical relationship between the scope for growth for the targeted ascidians and TSPM explained assemblages' recorded change but failed to explain current ascidians distribution. The results may indicate the existence of a TSPM threshold that allows the spatial coexistence of alternative stable states at benthic Potter Cove system.

Ascidian biodiversity in the shallow waters of the Madeira Archipelago: Fouling studies on artificial substrates and new records

Ascidians are recognized as major invaders on a global scale, found from the poles to the tropics and from shallow to deep sea waters with approximately 3000 known described species worldwide. However, to date only a few opportunistic studies focusing on the ascidian diversity colonizing natural and artificial substrates have been conducted in the Madeira Archipelago (NE Atlantic). Furthermore, little is known about the community composition and distribution of native and non-indigenous ascidians in the islands of the archipelago. In this context, we conducted a 3-phase study that allowed us to report an updated list of ascidian records found on artificial substrates in the Madeira Archipelago. First, to assess ascidian biodiversity colonizing artificial substrates present in the Madeira Archipelago, we conducted standardized monitoring campaigns in four marinas and hull vessel surveys during dry dock inspections. Second, we complemented this inventory with a comprehensive literature search and Museum collection focused on ascidians colonizing both artificial and natural substrates. Third, we investigated the temporal and spatial variability on the recruitment and succession of young vs mature ascidian assemblages. We found 24 ascidian species in all four marinas, from which 13 are NIS and 11 are new records for Madeira Archipelago. In addition, a literature survey found a total of 27 ascidians occurring on both artificial and natural substrates. This study now reveals a total of 39 ascidian species colonizing both artificial and natural substrates and comprises the most up-to-date list for all shallow-water ascidians in the Madeira Archipelago. A total of 15 NIS are now currently known, 9 exclusively from Madeira Island, 1 exclusively from Porto Santo Island and 5 on the two main islands of the archipelago. This study also shows that the spatial and temporal variability of fouling ascidian assemblages are driven by recruitment and succession of NIS rather than by presence of native species. Finally, total ascidian richness and cover was significantly higher in 4-month-old communities than in 8 to 16-month-old communities across all marinas, suggesting that “bare” artificial substrates are the main drivers of NIS success in the region.

The influence of invasive ascidian diets on the growth of the sea star Henricia sanguinolenta

AbstractInvasive species can disrupt food webs by altering the abundance of prey species or integrating into the food web themselves. In the Gulf of Maine, there have been a suite of invasions that have altered the composition of the benthic ecosystem. These novel prey species can potentially benefit native predators depending on their nutritional value and relative abundance. We measured feeding instances of the native blood star, Henricia sanguinolenta, and changes in the seasonal abundances of invasive ascidian prey species. Results indicate that H. sanguinolenta forages optimally, as the blood star will prey on invasive ascidians when in high abundance, but feed on other species during periods of scarcity. Further, our study shows that blood stars prey on a wider variety of species than was previously known, such as small bivalves and barnacles. Additionally, we compared growth and reproduction of sea stars fed different combinations of invasive ascidians (Diplosoma listerianum or Botrylloides violaceus) or a native sponge (Haliclona oculata). Sea stars grew more on the native diet when compared with the invasive ascidian species, and D. listerianum appeared to be a superior quality food source when compared with B. violaceus. By comparing our data with historical data, we determined that there was a dramatic increase in sea star populations between 1980 and 2011, but then populations decreased by almost half from 2011 to 2016–2017. These data suggest that while invasive ascidians may have helped sea star populations at one point, sea stars are declining without their native food source.

En masse DNA Electroporation for in vivo Transcriptional Assay in Ascidian Embryos.

Ascidian embryos are powerful models for functional genomics, in particular, due to the ease of generating a large number of transgenic embryos by electroporation. In addition, the small size of their genome makes them an attractive model for studying cis-regulatory elements that control gene expression during embryonic development. Here, I describe the adaptation of the seminal method developed 25 years ago in Ciona robusta for en masse DNA electroporation for in vivo transcription to additional species belonging to three genera. It is likely that similar optimizations would make electroporation successful in other ascidian species, where in vitro fertilization can be performed on a large number of eggs.

Morphology, genetics, and historical records support the synonymy of two ascidian species and suggest their spread throughout areas of the Southern Hemisphere

Taxonomic uncertainties and the lack of ecological knowledge can hinder the correct identification and the assignment of biogeographic status of marine species. The ascidian Asterocarpa humilis (Heller, 1878), originally described from New Zealand, has a broad distribution in shallow temperate areas of the Southern Hemisphere, having recently colonised areas of the Northern Hemisphere. A closely related species, Cnemidocarpa robinsoni Hartmeyer, 1916, has been reported in the South-Eastern Pacific and the South-Western Atlantic, and several authors considered it a junior synonym of A. humilis. We gathered for the first time morphological and genetic data from specimens from distant areas. We studied the morphology of specimens collected at seven locations of South America. We also re-examined specimens from museum collections and revised the available literature on these species. Genetic data were obtained from specimens from Argentina and compared with available sequences of A. humilis from Chile, New Zealand, England and France. Morphological and genetic analyses showed that all compared specimens were conspecific. Furthermore, specimens from different continents shared haplotypes and exhibited low genetic distance among them. These results, the biological characteristics of this ascidian, and its longstanding presence in different habitats from disjoint areas, allow us to question its native range. We support the idea that A. humilis is a cryptogenic and neocosmopolitan species that has been transported by maritime traffic through the Southern Hemisphere, revealing frequent processes of exchange through this wide area for more than a century, with presumably associated alterations in the marine biota.

Effects of temperature and salinity on egg development and larval settlement of an invasive ascidian species, <i>Herdmania momus</i> (Savigny, 1816)

The effects of temperature and salinity on egg development and settlement of the ascidian Herdmania momus were investigated. Adult specimens were collected from the Dodu Yacht facility in Jeju Island, Korea (33°30′30.54″N, 126°27′55.46″E) in August 2018. Egg development and larval settlement were observed and recorded at 8 h intervals using a stereomicroscope, under nine temperature (10, 13, 16, 19, 22, 25, 28, 31, and 34°C), and four salinity regimens (28, 30, 32, and 34 psu). The highest hatching rate (82.8±7%) was observed at 32 psu and 25°C and the lowest hatching rate (1.0±2%) was at 34 psu and 13°C. The developmental rate (0.222±0.0994) was highest at 28 psu and 28°C, and lowest (0.016±0.008) at 30 psu and 13°C. The highest settlement success rate (77.1±5%) was at 32 psu and 25°C and the lowest (0.1±1.0%) was at 30 psu, and 13°C. The rate of settlement (0.080±0.000) was highest at 28 psu and 28°C, and lowest (0.013±0.000) at 30 psu and 13°C. Both hatching and settlement success rates increased as temperature increased and tended to decrease beyond an optimal temperature range. Herdmania momus preferred 30-34 psu salinity and 22-25°C temperature. This study provides baseline information about the life history of H. momus, and important data to control the damage caused by the increase in number and distribution of this invasive ascidian.

The Pericardial Body of Ciona intestinalis Contains Hemocytes and Degenerating Muscle Cells, But No Parasites

PurposeA ventral heart positioned posterior to the branchial basket and equipped with a pericardium is homologous in tunicates and their sister group, the craniates, yet the tunicate model organism Ciona intestinalis features a pericardial body, a structure peculiar to few ascidian species. Here, we set out to distinguish between two competing hypotheses regarding the function of the pericardial body found in the literature: (H1) The pericardial body performs a role in the removal of dysfunctional myocardial cells, and (H2) it is a specialized niche of the immune system involved in defense against parasites.MethodsWe used histological techniques, transmission electron microscopy, and PCR-based gene sequencing to investigate whether individual ascidians parasitized with apicomplexan protists show signs of infections within the pericardial body.ResultsIn individuals of C. intestinalis from the German North Sea infested with apicomplexan protists, the pericardial body contains numerous myocardial cells in various stages of degeneration while no remnants of parasitic cells could be identified.ConclusionThus, we conclude that H2—the pericardial body is a specialized niche of the immune system involved in defense against parasites—can be refuted. Rather, our observations support H1, the hypothesis that the pericardial body performs a role in the removal of dysfunctional myocardial cells.

First occurrence of fouling ascidian species Microcosmus squamiger Michaelsen, 1927 and Didemnum ahu Monniot C. & Monniot F., 1987 in İzmir Bay (Eastern Aegean Sea)

ABSTRACT The aim of the present study was to determine species composition of ascidians and to detect the occurrence of alien ascidian species in İzmir Bay (Eastern Aegean Sea). The study was conducted at seven stations situated in recreational marinas and frequently used places for boating, fishing, and shipping activities in İzmir Bay between 25 and 27 August 2015 and between 23 and 26 November 2015. According to the findings, the non-native ascidian species such as Microcosmus squamiger Michaelsen, 1927, Didemnum ahu Monniot C. & Monniot F., 1987, and Polyclinum sp. were recorded for the first time on natural and artificial substrates at the sampling stations. The results indicate an increase in the number of alien ascidian species in the region. Also, the findings of this study showed that global marine invasive species M. squamiger has spread from the western Mediterranean to the eastern Mediterranean.

A Nodal/Eph signalling relay drives the transition from apical constriction to apico-basal shortening in ascidian endoderm invagination.

Gastrulation is the first major morphogenetic event during animal embryogenesis. Ascidian gastrulation starts with the invagination of 10 endodermal precursor cells between the 64- and late 112-cell stages. This process occurs in the absence of endodermal cell division and in two steps, driven by myosin-dependent contractions of the acto-myosin network. First, endoderm precursors constrict their apex. Second, they shorten apico-basally, while retaining small apical surfaces, thereby causing invagination. The mechanisms that prevent endoderm cell division, trigger the transition between step 1 and step 2, and drive apico-basal shortening have remained elusive. Here, we demonstrate a conserved role for Nodal and Eph signalling during invagination in two distantly related ascidian species, Phallusia mammillata and Ciona intestinalis Specifically, we show that the transition to step 2 is triggered by Nodal relayed by Eph signalling. In addition, our results indicate that Eph signalling lengthens the endodermal cell cycle, independently of Nodal. Finally, we find that both Nodal and Eph signals are dispensable for endoderm fate specification. These results illustrate commonalities as well as differences in the action of Nodal during ascidian and vertebrate gastrulation.

DNA bar coding of Aplousobranchiata and Phlebobranchiata Ascidians (Phylum:Chordata) inferred from mitochondrial cytochrome oxidase subunit I (COI) gene sequence approach in Andaman and Nicobar Islands, India: a first report

Ascidians (Phylum: Chordata) are sessile and filter-feeding marine animal, species identification of ascidians is possible by observing various morphological and anatomical features in various stages of life span. However, this method is labor intensive, time-consuming and very difficult for non-specialists particularly when dealing with field collections. Suborder Aplousobranchiata and Phlebobranchiata is the largest group of tunicates within, morphological and molecular data suggest that Didemnidae and Ascidiidae are monophyletic, but the monophyly of each genus and their phylogenetic relationships are still poorly understood. Therefore, this study was aimed to develop DNA barcodes of ascidians belonging to the orders of Aplousobranchiata and Phlebobranchiata species namely Diplosoma listerianum, Lissoclinum fragile, Didemnum psammatode, Phallusia fumigata and Phallusia ingeria collected from Andaman and Nicobar Islands were sequenced and submitted in Gen Bank. Colony structure, Scanning Electron Microscope (SEM) for spicules of colonial ascidians, larval type and zooids formation were found to be the most useful morphological characters for discriminating the species. Our BLAST results proved D. Listerianum KP842724 (98%) L. fragile KP842726 (100%) D. psammatode KP779902 (99%), P. fumigata KP779904 (99%) and P. ingeria KP842727 (100%) similarity and this is the first report of mitochondrial COI gene of these ascidians from Andaman and Nicobar Islands. We explored the usefulness of CO1 gene sequences for molecular level identification and mtDNA data in assessing a phylogenetic relationship of ascidian species.

Hidden diversity in Antarctica: Molecular and morphological evidence of two different species within one of the most conspicuous ascidian species.

The Southern Ocean is one of the most isolated marine ecosystems, characterized by high levels of endemism, diversity, and biomass. Ascidians are among the dominant groups in Antarctic benthic assemblages; thus, recording the evolutionary patterns of this group is crucial to improve our current understanding of the assembly of this polar ocean. We studied the genetic variation within Cnemidocarpa verrucosa sensu lato, one of the most widely distributed abundant and studied ascidian species in Antarctica. Using a mitochondrial and a nuclear gene (COI and 18S), the phylogeography of fifteen populations distributed along the West Antarctic Peninsula and Burdwood Bank/MPA Namuncurá (South American shelf) was characterized, where the distribution of the genetic distance suggested the existence of, at least, two species within nominal C. verrucosa. When reevaluating morphological traits to distinguish between genetically defined species, the presence of a basal disk in one of the genotypes could be a diagnostic morphological trait to differentiate the species. These results are surprising due to the large research that has been carried out with the conspicuous C. verrucosa with no differentiation between species. Furthermore, it provides important tools to distinguish species in the field and laboratory. But also, these results give new insights into patterns of differentiation between closely related species that are distributed in sympatry, where the permeability of species boundaries still needs to be well understood.

Shotgun Proteomics of Ascidians Tunic Gives New Insights on Host-Microbe Interactions by Revealing Diverse Antimicrobial Peptides.

Ascidians are marine invertebrates associated with diverse microbial communities, embedded in their tunic, conferring special ecological and biotechnological relevance to these model organisms used in evolutionary and developmental studies. Next-generation sequencing tools have increased the knowledge of ascidians’ associated organisms and their products, but proteomic studies are still scarce. Hence, we explored the tunic of three ascidian species using a shotgun proteomics approach. Proteins extracted from the tunic of Ciona sp., Molgula sp., and Microcosmus sp. were processed using a nano LC-MS/MS system (Ultimate 3000 liquid chromatography system coupled to a Q-Exactive Hybrid Quadrupole-Orbitrap mass spectrometer). Raw data was searched against UniProtKB – the Universal Protein Resource Knowledgebase (Bacteria and Metazoa section) using Proteome Discoverer software. The resulting proteins were merged with a non-redundant Antimicrobial Peptides (AMPs) database and analysed with MaxQuant freeware. Overall, 337 metazoan and 106 bacterial proteins were identified being mainly involved in basal metabolism, cytoskeletal and catalytic functions. 37 AMPs were identified, most of them attributed to eukaryotic origin apart from bacteriocins. These results and the presence of “Biosynthesis of antibiotics” as one of the most highlighted pathways revealed the tunic as a very active tissue in terms of bioactive compounds production, giving insights on the interactions between host and associated organisms. Although the present work constitutes an exploratory study, the approach employed revealed high potential for high-throughput characterization and biodiscovery of the ascidians’ tunic and its microbiome.