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14 pages, 6712 KiB

Open AccessArticle

Nanoplastic-Induced Developmental Toxicity in Ascidians: Comparative Analysis of Chorionated and Dechorionated Phallusia mammillata Embryos

by Maria Concetta Eliso, Ilaria Corsi, Antonietta Spagnuolo and Rémi Dumollard

J. Xenobiot. 2025, 15(1), 10; https://doi.org/10.3390/jox15010010 - 10 Jan 2025

Viewed by 557

Abstract

Nanoplastics pose a growing threat to marine ecosystems, particularly affecting the early developmental stages of marine organisms. This study investigates the effects of amino-modified polystyrene nanoparticles (PS-NH₂, 50 nm) on the embryonic development of Phallusia mammillata, a model ascidian species. [...] Read more.

Nanoplastics pose a growing threat to marine ecosystems, particularly affecting the early developmental stages of marine organisms. This study investigates the effects of amino-modified polystyrene nanoparticles (PS-NH₂, 50 nm) on the embryonic development of Phallusia mammillata, a model ascidian species. Both chorionated and dechorionated embryos were exposed to increasing concentrations of PS-NH₂ so morphological alterations could be assessed with a high-content analysis of the phenotypes and genotoxicity. PS-NH₂ induced the same morphological alterations in both chorionated and dechorionated embryos, with dechorionated embryos being more sensitive (EC₅₀ = 3.0 μg mL⁻¹) than chorionated ones (EC₅₀ = 6.26 μg mL⁻¹). Interestingly, results from the morphological analysis showed two concentration-dependent mechanisms of action: (i) at concentrations near the EC₅₀, neurodevelopmental abnormalities resembling the ones induced by exposure to known endocrine disruptors (EDs) were observed, and (ii) at higher concentrations (15 μg mL⁻¹ and 7.5 μg mL⁻¹ for chorionated and dechorionated embryos, respectively), a nonspecific toxicity was evident, likely due to general oxidative stress. The phenotypes resulting from the PS-NH₂ treatment were not related to DNA damage, as revealed by a genotoxicity assay performed on neurula embryos. Our data suggest that PS-NH₂-induced toxicity is primarily mediated through oxidative stress, probably triggered by interactions between the positive charges of the PS NPs and the negative charges on the cell membranes. The lack of a protective chorion further exacerbated these effects, highlighting its role in mitigating/protecting against NP-induced damage. Full article

(This article belongs to the Special Issue Feature Papers in Ecotoxicology)

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Figure 1
Percentage (%) of normal hatched larvae of P. mammillata upon 22 h exposure of chorionated embryos (A) and dechorionated embryos (B) to PS-NH2 in NSW. Bars represent mean ± SD (PS-NH2 N = 240). Asterisks indicate values that are significantly different compared to the control (Kruskal–Wallis test, Dunn’s post hoc test (* p < 0.05, ** p < 0.01, *** p < 0.001)). EC50 values are shown. Full article ">Figure 2
Light microscopy images of P. mammillata embryos exposed for 22 h to PS-NH2. (A–E) represent the phenotypes for embryos developed in a chorion: (A) control; (B) 5 μg mL−1; (C) 7.5 μg mL−1; (D) 10 μg mL−1; (E) 15 μg mL−1. White and red arrows represent a good and wrong shape of pigmented cells (PCs) and palps (P), respectively. Scale bar: 100 μm. Full article ">Figure 3
Light microscopy images of P. mammillata embryos exposed for 22 h to PS-NH2. (A–E) represent the phenotypes for embryos developed without a chorion: (A) control; (B) 2 μg mL−1; (C) 3 μg mL−1; (D) 3.5 μg mL−1; (E) 5 μg mL−1; (F) 7.5 μg mL−1. White and red arrows represent a good and wrong shape of pigmented cells (PCs) and palps (P), respectively. Scale bar: 100 μm. Full article ">Figure 4
Morphometric analysis of phenotypes induced in P. mammillata embryos (A) with a chorion and (B) without a chorion exposed to PS-NH2 (0; 7.5 and 15 μg mL−1 for chorionated embryos, 0; 3.5 and 7.5 μg mL−1 for dechorionated embryos). The radar charts summarize the following endpoints: ocellus (Oc) + otolith (Ot) area (µm2); Oc/Ot distance (µm); percentage of embryos with palps (%); trunk L/W (length/width) ratio; tail length (µm). All measurements were performed at 22 hpf. The values are normalized to the corresponding value of the same parameter in the control (stage 26) and presented as a percentage of the control value. Complete radar charts of all tested concentrations are shown in <a href="#app1-jox-15-00010" class="html-app">Figure S3</a>. Full article ">Figure 5
Genotoxicity assay performed analyzing 7 hpf embryos (neurula stage). (A) Control embryo (NSW), (B) 5 μg mL−1; (C) 7.5 μg mL−1. At the bottom of each image, the percentage of embryos not showing DNA aberrations is reported (N control = 152; 5 μg mL−1 = 175; 7.5 μg mL−1 = 160.). Full article ">

28 pages, 8919 KiB

Open AccessArticle

Fine-Tuned Expression of Evolutionarily Conserved Signaling Molecules in the Ciona Notochord

by Lenny J. Negrón-Piñeiro, Yushi Wu, Ravij Mehta, Julie E. Maguire, Cindy Chou, Joyce Lee, Chitra L. Dahia and Anna Di Gregorio

Int. J. Mol. Sci. 2024, 25(24), 13631; https://doi.org/10.3390/ijms252413631 - 20 Dec 2024

Viewed by 651

Abstract

The notochord is an axial structure required for the development of all chordate embryos, from sea squirts to humans. Over the course of more than half a billion years of chordate evolution, in addition to its structural function, the notochord has acquired increasingly [...] Read more.

The notochord is an axial structure required for the development of all chordate embryos, from sea squirts to humans. Over the course of more than half a billion years of chordate evolution, in addition to its structural function, the notochord has acquired increasingly relevant patterning roles for its surrounding tissues. This process has involved the co-option of signaling pathways and the acquisition of novel molecular mechanisms responsible for the precise timing and modalities of their deployment. To reconstruct this evolutionary route, we surveyed the expression of signaling molecules in the notochord of the tunicate Ciona, an experimentally amenable and informative chordate. We found that several genes encoding for candidate components of diverse signaling pathways are expressed during notochord development, and in some instances, display distinctive regionalized and/or lineage-specific patterns. We identified and deconstructed notochord enhancers associated with TGF-β and Ctgf, two evolutionarily conserved signaling genes that are expressed dishomogeneously in the Ciona notochord, and shed light on the cis-regulatory origins of their peculiar expression patterns. Full article

(This article belongs to the Special Issue Ascidian Early Development)

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11 pages, 739 KiB

Open AccessArticle

Single but Not Combined In Vitro Exposure to Bisphenol A and Nanoplastics Affects the Cholinergic Function of the Ascidian Ciona robusta

by Safa Melki, Emma Ferrari, Raja Ben Ahmed, Antonietta Spagnuolo and Ilaria Corsi

J. Xenobiot. 2024, 14(4), 1930-1940; https://doi.org/10.3390/jox14040103 - 5 Dec 2024

Cited by 1 | Viewed by 762

Abstract

Nanoplastics are known to represent a threat to marine ecosystems. Their combination with other contaminants of emerging concerns (CECs) may amplify ecotoxic effects, with unknown impacts on marine biodiversity. This study investigates the effects, single and combined, of bisphenol A (BPA)—one of the [...] Read more.

Nanoplastics are known to represent a threat to marine ecosystems. Their combination with other contaminants of emerging concerns (CECs) may amplify ecotoxic effects, with unknown impacts on marine biodiversity. This study investigates the effects, single and combined, of bisphenol A (BPA)—one of the most hazardous CECs—and polystyrene nanoparticles (PS NPs)—as a proxy for nanoplastics, being among the most commonly found asmarine debris—on cholinesterase (ChE) activities of the ascidian Ciona robusta. ChE activity was first measured in the siphons, tunic, and viscera of wild-caught adult specimens and exposed in vitro to BPA (0.01, 0.21, 0.69 mM) and PS NPs (0.0096–0.096 mM; 8.096 × 10⁹–10¹⁰ particles, respectively) alone and combined for 15 min of incubation. PS NPs’ behavior in milliQ water and in the ChE assay reaction buffer was characterized alone, combined with BPA, and analyzed through ζ-potential measurements via Dynamic Light Scattering. The results revealed that ChE activity was predominant in the viscera and siphons of C. robusta; PS NPs did not affect the ChE activity alone or combined, while BPA caused a concentration-dependent inhibition of ChE activity in the viscera. No changes in ζ-potential were observed for PS NPs alone or combined with BPA in the ChE buffer, suggesting no interaction. Further investigations are needed to understand the potential neurotoxic consequences for C. robusta and ecological risk scenarios due to exposure to BPA and nanoplastics in marine coastal waters. Full article

(This article belongs to the Special Issue Feature Papers in Ecotoxicology)

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15 pages, 4878 KiB

Open AccessArticle

Involvement of Metalloproteases in the Fertilization of the Ascidian Halocynthia roretzi

by Hitoshi Sawada, Ikuya Hattori, Noritaka Hashii and Takako Saito

Biomolecules 2024, 14(12), 1487; https://doi.org/10.3390/biom14121487 - 22 Nov 2024

Viewed by 658

Abstract

We previously reported that five astacin-like metalloproteases with thrombospondin type-1 repeats (Tasts) located on the sperm surface are a promising candidate as the protease involved in sperm penetration of the vitelline coat (VC) during fertilization of the ascidian Ciona intestinalis type A (Phlebobranchia). [...] Read more.

We previously reported that five astacin-like metalloproteases with thrombospondin type-1 repeats (Tasts) located on the sperm surface are a promising candidate as the protease involved in sperm penetration of the vitelline coat (VC) during fertilization of the ascidian Ciona intestinalis type A (Phlebobranchia). However, whether such a protease is involved in the fertilization of other ascidians is unknown. Here, we investigated the effects of four metalloprotease inhibitors on the fertilization of the ascidian Halocynthia roretzi (Stolidobranchia). Three metalloprotease inhibitors, GM6001, TAPI-0, and TAPI-1, strongly inhibited fertilization at 33 and 11 μM, whereas TAPI-2 weakly inhibited fertilization at 33 μM. In contrast, GM6001NC (negative control) had no effect on fertilization at 100 μM. Furthermore, GM6001 had no inhibitory effect on the fertilization of VC-deprived eggs. The metalloprotease appears to function at the middle or late stage of fertilization. Ten Tast genes were identified in the H. roretzi genome database, among which four genes (HrTast1, HrTast2b, HrTast2c, and HrTast3c) possessed a single transmembrane domain in the N-terminal region. These four genes are transcribed in the testis and ovary, as revealed by RT-PCR. Anti-HrTast2c IgG raised against a peptide corresponding to the Zn-binding consensus sequence weakly inhibited fertilization at 0.5 mg/mL. These results led us to propose that sperm astacin-like metalloproteases may be involved in sperm penetration of the VC during H. roretzi fertilization. Full article

(This article belongs to the Special Issue Gametogenesis and Gamete Interaction, 2nd Edition)

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14 pages, 11443 KiB

Open AccessFeature PaperArticle

Immunohistochemical Characterization of Spermatogenesis in the Ascidian Ciona robusta

by Haruka Sakurai, Kogiku Shiba, Katsumi Takamura and Kazuo Inaba

Cells 2024, 13(22), 1863; https://doi.org/10.3390/cells13221863 - 11 Nov 2024

Viewed by 860

Abstract

Animals show diverse processes of gametogenesis in the evolutionary pathway. Here, we characterized the spermatogenic cells in the testis of the marine invertebrate Ciona robusta. Ciona sperm differentiate in a non-cystic type of testis, comprising many follicles with various sizes and stages of [...] Read more.

Animals show diverse processes of gametogenesis in the evolutionary pathway. Here, we characterized the spermatogenic cells in the testis of the marine invertebrate Ciona robusta. Ciona sperm differentiate in a non-cystic type of testis, comprising many follicles with various sizes and stages of spermatogenic cells. In the space among follicles, we observed free cells that were recognized by antibody against Müllerian inhibiting substance, a marker for vertebrate Sertoli cells. We further categorized the spermatogenic cells into four round stages (RI to RIV) and three elongated stages (EI to EIII) by morphological and immunohistochemical criteria. An antibody against a vertebrate Vasa homolog recognized a few large spermatogonium-like cells (RI) near the basal wall of a follicle. Consistent with the period of meiosis, a synaptonemal complex protein SYCP3 was recognized from early spermatocytes (RII) to early spermatids (E1). Acetylated tubulins were detected in spermatids before flagellar elongation at the RIV stage and became distributed along the flagella. Electron microscopy showed that the free cells outside the testicular follicle possessed a characteristic of vertebrate Sertoli cells. These results would provide a basis for basic and comparative studies on the mechanism of spermatogenesis. Full article

(This article belongs to the Special Issue The Cell Biology of Fertilization)

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28 pages, 3510 KiB

Open AccessReview

Harnessing Ascidians as Model Organisms for Environmental Risk Assessment

by Amalia Rosner and Baruch Rinkevich

Environments 2024, 11(11), 232; https://doi.org/10.3390/environments11110232 - 23 Oct 2024

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Abstract

Environmental Risk Assessment (ERA) often relies on a restricted set of species as bio-indicators, introducing uncertainty when modeling complex environmental variables. This may lead to oversimplified or erroneous risk assessments. Ascidians, marine filter-feeding sessile chordates, are valuable models for scientific research in various [...] Read more.

Environmental Risk Assessment (ERA) often relies on a restricted set of species as bio-indicators, introducing uncertainty when modeling complex environmental variables. This may lead to oversimplified or erroneous risk assessments. Ascidians, marine filter-feeding sessile chordates, are valuable models for scientific research in various biological fields such as stem cell biology, embryogenesis, regeneration, innate immunity, and developmental biology. Their global distribution, sensitivity to pollutants, high abundance, mass sexual reproduction, and habitation in coastal areas impacted by anthropogenic pollution make them excellent indicators for monitoring marine pollution and global environmental changes, including biological invasions and species diversity diminution cases. Despite their potential as environmental bioindicators, ascidians remain underutilized in ERAs (≤0.13% of ERA studies), particularly in the field of chemical pollution impact assessment, primarily due to a lack of standardization. This underrepresentation poses a challenge for accurate modeling, especially in models relying on a broad range of species (e.g., Species Sensitivity Distributions). Given these constraints, expanding the use of ascidians in ERAs could improve the comprehension and precision of environmental changes and their assessments. This underscores the necessity for future research to establish standardized testing protocols and choose the most suitable ascidian species for inclusion in ERAs. Full article

(This article belongs to the Special Issue Environmental Risk Assessment of Aquatic Ecosystem)

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Figure 1
Phylogenetic tree based on 18S ribosomal RNA (18S rRNA) gene fragments of ascidian species reviewed in this study. A few species are not represented due to the absence of their 18S gene sequences in the database. The taxonomic order of the species is depicted, illustrating the clustering of species within the same order. The analysis was conducted using the EMBL-EBI T-Coffee program [<a href="#B33-environments-11-00232" class="html-bibr">33</a>]. The NCBI GenBank accession numbers for the 18S rRNA gene sequences used are as follows: Ascidiella scabra (AB811928.1), Botrylloides leachi (JN573237.1), Botrylloides_violaceus (AY903927.1), Botryllus_schlosseri (FM244858.1), Ciona intestinalis (AB013017.1), Ciona savignyi (LC547329.1), Didemnum molle (AB211071.1), Didemnum vexillum (JF738071.1), Halocynthia roretzi (AB013016.1), Herdmania momus (AF165827.1), Microcosmus exasperates (XR005567858.1), Molgula manhattensis (L12426.2), Phallusia nigra (FM244845.1), Polycarpa mytiligera (FM244860.1), Styela clava (XR_005567858.1), Styela_plicata (L12444.2). Full article ">Figure 2
Modes of life cycles in two ascidian model species: (a) a solitary ascidian (Ciona spp.) revealing classical sexual reproduction progressions of a broadcasting species; (b) asexual (outer cycle in the diagram) and sexual phases in a colonial ascidian (Botryllus schlosseri) highlighted by weekly astogenic rounds of zooidal life and death (each cycle is called blastogenesis). Each blastogenic cycle is divided into four stages (A–D) where three generations of colonial modules (the functional zooids and two cohorts of developing buds, primary and secondary buds) coexist side by side, depicting highly synchronized developmental statuses as the colony progresses through blastogenesis. At stage D, the functional zooids start their degeneration, first by closing the siphons, where simultaneously all zooids undergo cell apoptosis and phagocytosis processes within the next 24–36 h and are morphologically absorbed, while the primary buds mature to the zooidal level of development. Subsequently, stage A of the following blastogenic cycle begins as primary buds complete their development into zooids by opening the inhalant siphons and resuming water filtration, while secondary buds (budlets) develop to the primary bud’s state, starting the generation of new sets of secondary buds. Gametogenesis is highly synchronized with the blastogenic cycle among modules of the same generation. The sexual cycle is typified as brooding, where gametes start differentiation within the budlets. Gametes maturation and egg fertilization occur within the zooids (at the onset of stage A). Embryos differentiation is synchronized with the blastogenic stage and continues within the zooids (inner cycle; showing for each blastogenic stage a single large zooid with its bud/s and budlet/s), culminating in the release of the larvae into the surrounding waters at late blastogenic stage C. The larvae swim for a very short period until they settle, undergo metamorphosis starting with the absorption of the tail, and new juveniles (oozooids) are formed, and a colony is formed by repeated blastogenic cycles. Full article ">Figure 3
Pie charts depicting taxa percentages for search hits in the ‘Web of Science’ and ‘Google Scholar’ databases, filtered by the terms ‘environmental risk assessments’ and ‘marine’ environment, as compared to the total hits for ‘environmental risk assessment’ (marine and terrestrial). Each taxon (a group of different bioindicators) is represented by a specific-colored descriptor detailed in the caption. The black sections encompass the residual hits for taxa with the smaller number of hits. The magnified pie sections on the right provide a detailed breakdown for some of these taxa. <a href="#app1-environments-11-00232" class="html-app">Supplementary Table S1</a> details the specific queries used to obtain the data for each taxon. Full article ">Figure 4
Commonly used model ascidians: the solitary ascidians (a) Ciona robusta and (b) Ciona intestinalis (by John Bishop from the Marine Biological Association of the United Kingdom), once considered as a single species; (c,d) different color morphs of the colonial ascidian Botryllus schlosseri. (c) A colony reared in the laboratory at the Israel Oceanography and Limnological Research, Haifa, and maintained at a constant temperature of 20 °C with a regimen of 12:12 light:dark hours. This colony is a descendant of the Monterey, California, population; (d) a colony from New Zealand reared on a glass slide. Full article ">Figure 5
Pictures of additional solitary (a–c) and colonial ascidian (d–f) used in toxicity, environmental pollution monitoring tests, and biological invasions. (a) Phallusia spp.; (b) Polycarpa spp.; (c) Halocynthia spp.; (d) Botrylloides spp.; (e) Didemnum spp.; (f) Didemnum vexillum. Full article ">Figure 6
Pie charts depicting taxa percentages for search hits in ‘Google Scholar’ databases, filtered by the terms: (a) ‘environmental risk assessment’ and ‘marine’ environment, and ‘invasion’ (61.5% of hits) as compared to the total hits for ‘environmental risk assessments’ and ‘invasion’ (100%). The total number of hits involving individual taxa exceeds the number of hits for ‘environmental risk assessment’, ‘marine’ environment, and ‘invasion’, as many publications examine multiple taxa; (b) ‘environmental risk assessment’ and ‘marine’ environment, and ‘biodiversity’ (62.6%) as compared to the total hits for ‘environmental risk assessments’ and ‘biodiversity’ (100%). The total number of hits involving individual taxa exceeds the number of hits for ‘environmental risk assessment’ and ‘marine’ environment and ‘biodiversity’, as many publications examine multiple taxa. Each taxon is represented by a specific-colored descriptor detailed in the caption. The magnified pie sections on the right are for categories with smaller numbers of hits. <a href="#app1-environments-11-00232" class="html-app">Supplementary Table S2</a> details the specific queries used to obtain the data for each taxon. Full article ">Figure 7
A graphical representation illustrating the potential applications of ascidian-based bioassays within the framework of ERA. The bioassays utilize both solitary and colonial ascidians, which may offer unique advantages for studying different environmental impacts. MCR—multiple clonal ramets; Blue arrows indicate bioassays that have already been successfully employed in toxicity testing; Red arrows represent potential applications that have yet to be widely explored. Full article ">

19 pages, 4753 KiB

Open AccessArticle

New Clavelina (Ascidiacea) Species from the Bahamas

by Xavier Turon and Susanna López-Legentil

Taxonomy 2024, 4(3), 661-679; https://doi.org/10.3390/taxonomy4030034 - 3 Sep 2024

Viewed by 1746

Abstract

The ascidian fauna of the Bahamas remains grossly understudied. Here, we examined specimens of the genus Clavelina collected from four Bahamian islands using morphological observations and genetic barcoding. Only three species of Clavelina have been previously reported in the Caribbean: C. picta, [...] Read more.

The ascidian fauna of the Bahamas remains grossly understudied. Here, we examined specimens of the genus Clavelina collected from four Bahamian islands using morphological observations and genetic barcoding. Only three species of Clavelina have been previously reported in the Caribbean: C. picta, C. oblonga, and C. puertosecensis. Here, we report C. picta and three species new to science: C. rochae, C. pawliki, and C. erwinorum. C. picta and C. pawliki were found in the northernmost island surveyed, while C. rochae and C. erwinorum were particularly prevalent on the southeastern Bahamian islands. A complete review of genetic barcoding data and morphological characters of accepted Clavelina species was performed. The unique combination of in vivo coloration, morphological characteristics, haplotypes, and species distribution supported the establishment of three new species, significantly adding to the diversity of the genus Clavelina in the Caribbean. Full article

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73 pages, 18532 KiB

Open AccessReview

An Overview on the Synthesis of Lamellarins and Related Compounds with Biological Interest

by Vasiliki-Panagiota M. Mitsiou, Anastasia-Maria N. Antonaki, Matina D. Douka and Konstantinos E. Litinas

Molecules 2024, 29(17), 4032; https://doi.org/10.3390/molecules29174032 - 26 Aug 2024

Cited by 1 | Viewed by 1268

Abstract

Lamellarins are natural products with a [3,4]-fused pyrrolocoumarin skeleton possessing interesting biological properties. More than 70 members have been isolated from diverse marine organisms, such as sponges, ascidians, mollusks, and tunicates. There is a continuous interest in the synthesis of these compounds. In [...] Read more.

Lamellarins are natural products with a [3,4]-fused pyrrolocoumarin skeleton possessing interesting biological properties. More than 70 members have been isolated from diverse marine organisms, such as sponges, ascidians, mollusks, and tunicates. There is a continuous interest in the synthesis of these compounds. In this review, the synthetic strategies for the synthesis of the title compounds are presented along with their biological properties. Three routes are followed for the synthesis of lamellarins. Initially, pyrrole derivatives are the starting or intermediate compounds, and then they are fused to isoquinoline or a coumarin moiety. Second, isoquinoline is the starting compound fused to an indole moiety. In the last route, coumarins are the starting compounds, which are fused to a pyrrole moiety and an isoquinoline scaffold. The synthesis of isolamellarins, azacoumestans, isoazacoumestans, and analogues is also described. The above synthesis is achieved via metal-catalyzed cross-coupling, [3 + 2] cycloaddition, substitution, and lactonization reactions. The title compounds exhibit cytotoxic, multidrug resistance (MDR), topoisomerase I-targeted antitumor, anti-HIV, antiproliferative, anti-neurodegenerative disease, and anti-inflammatory activities. Full article

(This article belongs to the Special Issue Coumarin and Its Derivatives III)

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13 pages, 3820 KiB

Open AccessArticle

Updated and Corrected List of Hosts of European Pinnotherids (Crustacea, Decapoda, and Pinnotheridae): Relationship between Number of Hosts and Distribution

by Jose A. Cuesta, Sara Román, Isabel Muñoz, Ana Rodríguez de la Rúa, Carlos Farias, Luis Silva and Marina Delgado

Diversity 2024, 16(8), 470; https://doi.org/10.3390/d16080470 - 4 Aug 2024

Viewed by 698

Abstract

The present work provides an updated and corrected list of hosts for the five European species of pinnotherid crabs, taking into consideration all recent published works as well as new data obtained in the present study. In relation to the number of hosts, [...] Read more.

The present work provides an updated and corrected list of hosts for the five European species of pinnotherid crabs, taking into consideration all recent published works as well as new data obtained in the present study. In relation to the number of hosts, there are two groups of species, one composed by Pinnotheres pisum and Afropinnotheres monodi with the highest number of hosts, and a second group, with a reduced number of hosts and more specific taxa, consisting of three other species: Nepinnotheres pinnotheres, Pinnotheres bicristatus, and P. pectunculi. When studying the relationships between host preferences, host distributions, and pea crab distributions, we could not find a clear direct relationship between these parameters. Temperature is also probably an important influence and relevant factor responsible for the distribution of pinnotherid crabs, and this may be independent from the potential distribution due to the distribution of hosts. Full article

(This article belongs to the Section Marine Diversity)

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11 pages, 1990 KiB

Open AccessFeature PaperEditor’s ChoiceArticle

The Interplay of TLR-NFκB Signalling Pathway and Functional Immune-Related Enzymes in the Inflammatory Response of Ciona robusta

by Luca Bisanti, Claudia La Corte, Mariano Dara, Federica Bertini, Jacopo Vizioli, Maria Giovanna Parisi, Matteo Cammarata and Daniela Parrinello

Animals 2024, 14(15), 2169; https://doi.org/10.3390/ani14152169 - 25 Jul 2024

Cited by 1 | Viewed by 1060

Abstract

The close phylogenetic relationship between ascidians (Tunicata) and vertebrates makes them a powerful model for studying the innate immune system. To better understand the nature and dynamics of immune responses and the mechanisms through which bacterial infections are detected and translated into inflammation [...] Read more.

The close phylogenetic relationship between ascidians (Tunicata) and vertebrates makes them a powerful model for studying the innate immune system. To better understand the nature and dynamics of immune responses and the mechanisms through which bacterial infections are detected and translated into inflammation in Ciona robusta, we applied an approach combining in vivo lipopolysaccharide (LPS) stimulation, immune-labelling techniques and functional enzymatic analyses. The immunohistochemistry showed that Toll-like receptor 4 (TLR4) and nuclear factor kappa B (NFκB) were expressed during the inflammatory pharynx response 4 h post-LPS, with the formation of nodules in pharynx vessel lumen. Also, the endothelium vessels were involved in the inflammatory response. Observations of histological sections from naive and buffer-inoculated ascidians confirmed an immuno-positive response. Enzyme immune parameters—which included the activity of phenoloxidase, glutathione peroxidase, lysozyme, alkaline phosphatase and esterase—showed up-modulation 4 h after LPS injection, confirming their participation during ascidian inflammatory response. These findings provide new insights into the mechanisms underlying the LPS-induced C. robusta response and suggest that a broad innate immune mechanism, as in vertebrates, is involved in the regulation of inflammatory responses. Further findings in this direction are needed to cover knowledge gaps regarding the organized set of molecular and cellular networks involved in universal immune interactions with pathogens. Full article

(This article belongs to the Section Aquatic Animals)

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12 pages, 3226 KiB

Open AccessArticle

Conserved Signaling Pathways in the Ciona robusta Gut

by Marco Gerdol, Samuele Greco, Rita Marino, Annamaria Locascio, Michelina Plateroti and Maria Sirakov

Int. J. Mol. Sci. 2024, 25(14), 7846; https://doi.org/10.3390/ijms25147846 - 18 Jul 2024

Viewed by 1087

Abstract

The urochordate Ciona robusta exhibits numerous functional and morphogenetic traits that are shared with vertebrate models. While prior investigations have identified several analogies between the gastrointestinal tract (i.e., gut) of Ciona and mice, the molecular mechanisms responsible for these similarities remain poorly understood. [...] Read more.

The urochordate Ciona robusta exhibits numerous functional and morphogenetic traits that are shared with vertebrate models. While prior investigations have identified several analogies between the gastrointestinal tract (i.e., gut) of Ciona and mice, the molecular mechanisms responsible for these similarities remain poorly understood. This study seeks to address this knowledge gap by investigating the transcriptional landscape of the adult stage gut. Through comparative genomics analyses, we identified several evolutionarily conserved components of signaling pathways of pivotal importance for gut development (such as WNT, Notch, and TGFβ-BMP) and further evaluated their expression in three distinct sections of the gastrointestinal tract by RNA-seq. Despite the presence of lineage-specific gene gains, losses, and often unclear orthology relationships, the investigated pathways were characterized by well-conserved molecular machinery, with most components being expressed at significant levels throughout the entire intestinal tract of C. robusta. We also showed significant differences in the transcriptional landscape of the stomach and intestinal tract, which were much less pronounced between the proximal and distal portions of the intestine. This study confirms that C. robusta is a reliable model system for comparative studies, supporting the use of ascidians as a model to study gut physiology. Full article

(This article belongs to the Section Molecular Biology)

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Figure 1
RNA-seq approach and preliminary results. (A) The anatomical parts of the Ciona gastrointestinal tract analyzed separately. S = stomach, PI = proximal intestine, DI = distal intestine, blue. (B) Principal component analysis, constructed based on log-transformed Transcript Per Million (TPM) expression values of all genes, summarizing the relatedness of the transcriptional profiles of the three biological samples of S (SA, SB, and SH), PI (PIA, PIB, and PIH), and DI (DIA, DIB, and DIH). (C) Heat map representing the log-transformed TPM expression levels of all DEGs characterized by fold change (FC) > 10 in at least one pairwise comparison among S, PI, and ID. DEGs were hierarchically clustered based on Euclidean distance, using an average linkage method. (D) Heat map representing the log-transformed TPM expression levels of the Hox and paraHox genes, hierarchically clustered based on Euclidean distance, using an average linkage method. Genes displaying statistically significant over-expression in one of the three GI tracts are boxed. The terms A, B, and H refer to different individual animals; for details, see the Material and Methods (<a href="#sec4-ijms-25-07846" class="html-sec">Section 4</a>). Full article ">Figure 2
Go enrichment and validation of digestion-related genes. Dendrogram representation of the significantly enriched GO terms in the S vs. I pairwise comparison. (A) GO terms associated with DEGs upregulated in S. (B) GO terms associated with DEGs upregulated in I. The size of the GO terms indicates the observed/expected ratio, whereas the color scale represents the statistical significance. GO term clustering was based on the presence of shared DEGs. (C) Validation by real-time quantitative PCR (RT-qPCR) of the digestion-related genes. Violin plots represent relative expression calculated using the ΔCt method, normalization against ELF1. N = 4. ns: not significant, ** p < 0.01 *** p < 0.001, **** p < 0.0001. Full article ">Figure 3
Canonical WNT signaling reconstruction. (A) Schematic view of the WNT/βcatenin pathway realized using BioRender.com (accessed on 13 September 2023). (B) STRING networks represent the interactions among selected genes linked with the Wnt pathway, arbitrarily classified as ligands, receptors, nuclear effectors, modulators, targets, and related genes. Only mouse genes sharing at least one orthologous gene with C. robusta are shown. Gene interaction networks were constructed based on known or inferred relationships among mouse orthologs. (C) Heat map representing the log-transformed TPM expression levels of the main genes (excluding target and related genes) included in the STRING network, observed in C. robusta S, PI, and DI samples. Full article ">Figure 4
Canonical Notch signaling reconstruction. (A) Schematic view of the Notch pathway realized using BioRender.com (accessed on 13 September 2023). (B) STRING networks representing the interactions among selected genes linked with the Notch pathway, arbitrarily classified as ligands, receptors, nuclear effectors, modulators, targets, and related genes. Only mouse genes sharing at least one orthologous gene with C. robusta are shown. Gene interaction networks were constructed based on known or inferred relationships among mouse orthologs. (C) Heat map representing the log-transformed TPM expression levels of the main genes (excluding target and related genes) included in the STRING network observed in C. robusta S, PI, and DI samples. Full article ">Figure 5
Canonical TGFβ-BMP signaling reconstruction. (A) Schematic view of the TGFβ-BMP pathway realized using BioRender.com (accessed on 13 September 2023). (B) STRING networks representing the interactions among selected genes linked with the TGFβ-BMP pathway, arbitrarily classified as ligands, receptors, nuclear effectors, modulators, targets, and related genes. Only mouse genes sharing at least one orthologous gene with C. robusta are shown. Gene interaction networks were constructed based on known or inferred relationships among mouse orthologs. (C) Heat map representing the log-transformed TPM expression levels of the main genes (excluding target and related genes) included in the STRING network, observed in C. robusta S, PI, and DI samples. Full article ">

15 pages, 11374 KiB

Open AccessArticle

Colonial Ascidian Populations at Inshore Coral Reefs of Norfolk Island, South Pacific

by Shannon Eckhardt, Tracy D. Ainsworth, William Leggat and Charlotte E. Page

Diversity 2024, 16(7), 384; https://doi.org/10.3390/d16070384 - 30 Jun 2024

Viewed by 1069

Abstract

Subtropical coral reefs such as the lagoonal reefs of Norfolk Island in the remote South Pacific are vastly understudied, with many benthic species unrecorded in the scientific literature. Here we report on ascidian populations from 2021 to 2023 at Norfolk Islands inshore reefs. [...] Read more.

Subtropical coral reefs such as the lagoonal reefs of Norfolk Island in the remote South Pacific are vastly understudied, with many benthic species unrecorded in the scientific literature. Here we report on ascidian populations from 2021 to 2023 at Norfolk Islands inshore reefs. Quantitative assessments spanning that period were conducted to assess ascidian presence, cover, and benthic interactions. We show ascidian cover remained persistently low and stable across the lagoonal reefs during the study period. We find adjacent reef site, Cemetery Bay showed variation in ascidian cover over time, where we measure a 3.2-fold increase in cover over 2 years. Ascidians were associated primarily with sand and sediment substrates at all reef locations. Recorded interactions between hard corals and ascidians were low. This study provides valuable insights into the population dynamics of ascidians within subtropical reef ecosystems. Ongoing ascidian population monitoring can provide a comprehensive understanding of ascidian dynamics whilst also providing insight of theimpact of anthropogenic stressors on benthic communities. Together, this can aid in informing management and conservation strategies for subtropical reefs. Full article

(This article belongs to the Special Issue Patterns Of Marine Benthic Biodiversity)

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(A) Location of Norfolk Island 1400 km east of the Australian East Coast. (B) Map of Norfolk Island and location of the 3 study sites (red box). (C) Slaughter, Emily, and Cemetery Bay are located in the south of Norfolk Island. The KAVHA catchment creek flows directly into the lagoon at Emily Bay. (D) Timeline of the bleaching event in March 2020 [<a href="#B47-diversity-16-00384" class="html-bibr">47</a>], followed by benthic surveys in April 2021, April 2022, and April 2023. Disease was prevalent from December 2020 until April 2023 [<a href="#B48-diversity-16-00384" class="html-bibr">48</a>]. Red line: Austral summer, blue line: Austral winter. Picture credits: (A) Google Maps, 2023, (B,C) Google Earth, 2023. Full article ">Figure 2
Images of ascidian colonies observed at Norfolk Island. (A–C) Colonial ascidian suspected to be Diplosoma spp. (D) Colonial ascidian suspected to be Diplosoma spp. containing photosynthetic algae. Photo credits: (A–C) Charlotte Page, (D) Ashley Coutts, Biofouling Solutions Pty Ltd., used with permission. Full article ">Figure 3
Benthic categories ascidians, cyanobacteria, hard coral, hard substrate, macroalgae, other benthic invertebrates, sand and sediment, turf, and all other. (A) Mean percent cover-based frequency distribution of the benthic categories present at each site (SB = Slaughter Bay; EB = Emily Bay; CB = Cemetery Bay) averaged over April 2021, 2022, and 2023. Each bar represents the relative proportion for each benthic category, calculated based on its mean percent cover value. See mean percent cover values in <a href="#diversity-16-00384-t002" class="html-table">Table 2</a>. (B) Ascidians. (C) Red cyanobacteria in the “cyanobacteria” category. (D) Hard coral. (E) Dead coral in the “hard substrate” category. (F) Macroalgae. (G) Anemone belonging to the “other benthic invertebrates” category. (H) Sand in the “sand and sediment” category. (I) Green turf belonging to the “turf” category. Photo credits: Norfolk Island, Charlotte Page, used with permission. Full article ">Figure 4
Box plots showing percent cover of Diplosoma spp. at (A) Slaughter Bay, (B) Emily Bay, (C) Cemetery Bay over three time points in April 2021, 2022, and 2023. The box represents the interquartile range (IQR; 25th to 75th percentile), with the black line indicating the median. Whiskers extend to 1.5 × IQR from the quartiles, and points beyond are outliers. Full article ">Figure 5
Percentage-based frequency distribution of (A) substrate Diplosoma spp. were found growing on and (B) interactions Diplosoma spp. had with cyanobacteria, hard coral, hard substrate, macroalgae, sand and sediment, and/or turf averaged the sites (SB = Slaughter Bay; EB = Emily Bay; CB = Cemetery Bay) over April 2021, 2022, and 2023. Diplosoma spp. growing (C) on coral, (D) in between a bed of macroalgae and turf, (E) on sand and interacting with turf, macroalgae, and coral, (F) on coral and interacting with red cyanobacteria, (G) on sediment and interacting with macroalgae, (H) on sand and interacting with turf and red cyanobacteria. Photo credits: Norfolk Island, Charlotte Page and Sophie Vuleta, used with permission. Full article ">

14 pages, 6494 KiB

Open AccessArticle

Marine-Derived Phosphoeleganin and Its Semisynthetic Derivative Decrease IL6 Levels and Improve Insulin Signaling in Human Hepatocellular Carcinoma Cells

by Ayewa L. Agognon, Marcello Casertano, Alessio Vito, Sonia Orso, Serena Cabaro, Federica Mormone, Cristina Morelli, Giuseppe Perruolo, Pietro Formisano, Marialuisa Menna, Concetta Imperatore and Francesco Oriente

Int. J. Mol. Sci. 2024, 25(11), 6039; https://doi.org/10.3390/ijms25116039 - 30 May 2024

Cited by 1 | Viewed by 925

Abstract

Marine natural products constitute a great source of potential new antidiabetic drugs. The aim of this study was to evaluate the role of phosphoeleganin (PE), a polyketide purified from the Mediterranean ascidian Sidnyum elegans, and its derivatives PE/2 and PE/3 on insulin [...] Read more.

Marine natural products constitute a great source of potential new antidiabetic drugs. The aim of this study was to evaluate the role of phosphoeleganin (PE), a polyketide purified from the Mediterranean ascidian Sidnyum elegans, and its derivatives PE/2 and PE/3 on insulin sensitivity in human hepatocellular carcinoma (HepG2) cells. In our experiments, insulin stimulates the phosphorylation of its receptor (INSR) and AKT by 1.5- and 3.5-fold, respectively, whereas in the presence of PE, PE/2, and PE/3, the insulin induced INSR phosphorylation is increased by 2.1-, 2-, and 1.5-fold and AKT phosphorylation by 7.1-, 6.0-, and 5.1-fold, respectively. Interestingly, PE and PE/2 have an additive effect on insulin-mediated reduction of phosphoenolpyruvate carboxykinase (PEPCK) expression. Finally, PE and PE/2, but not PE/3, decrease interleukin 6 (IL6) secretion and expression before and after palmitic acid incubation, while in the presence of high glucose (HG), only PE reduces IL6. Levels of other cytokines are not significantly affected by PE and its derivates. All these data suggest that PE and its synthetic-derived compound, PE/2, significantly decrease IL6 and improve hepatic insulin signaling. As IL6 impairs insulin action, it could be hypothesized that PE and PE/2, by inhibiting IL6, may improve the hepatic insulin pathway. Full article

(This article belongs to the Special Issue New Pharmacological Insights on Natural Macromolecules and Natural Small Molecules)

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Figure 1
(a) The marine polyketide phosphoeleganin (PE), purified from the Mediterranean ascidian S. elegans. (b) Previously reported pharmacological effects of PE and its semisynthetic derivatives PE/2 and PE/3. For the oxidative cleavage, reactions and conditions are (i) NaIO4, MeOH, rt, 3 h; (ii) NaBH4, 0 °C, 1 h. Full article ">Figure 2
Effect of PE, PE/2, and PE/3 on insulin receptor (INSR), AKT, and ERK1/2 phosphorylation. HepG2 cells were incubated with PE, PE/2, and PE/3 (25 µM) for 4 h and then stimulated with insulin (100 nM) for 10 min. The protein expression of pINSR, pAKT, and pERK1/2 was analyzed by Western blotting. INSR, AKT, and ERK1/2 bands were taken from a parallel gel loaded with the same lysates. Vinculin antibody was used for normalization. The autoradiographs shown are representative of three different experiments and subjected to densitometric analysis. Bars represent the mean ± SD of three independent experiments. Asterisks denote statistical differences (* p < 0.05; ** p < 0.01; *** p < 0.001). Full article ">Figure 3
Effect of PE, PE/2, and PE/3 on PEPCK expression and glycogen content. (a) HepG2 cells were incubated with PE, PE/2, and PE/3 (25 µM) and insulin (100 nM) for 8 h. PEPCK mRNA levels were evaluated by real-time RT-PCR analysis. Data were normalized by the amount of beta-ACTIN, used as an internal control. Bars represent the mean ± SD of three independent experiments, each performed in triplicate. (b) HepG2 cells were incubated with PE, PE/2, and PE/3 (25 µM) and insulin (100 nM) for 3 h. Glycogen content was measured as described in the <a href="#sec4-ijms-25-06039" class="html-sec">Section 4</a>. Asterisks denote statistical differences (* p < 0.05; ** p < 0.01; *** p < 0.001). Full article ">Figure 4
Effect of PE, PE/2, and PE/3 on IL6 expression. HepG2 cells were incubated with PE, PE/2, and PE/3 (25 µM) for 4 h. IL6 mRNA levels were evaluated by real-time RT-PCR analysis. Data were normalized by the amount of beta-ACTIN, used as an internal control. Bars represent the mean ± SD of three independent experiments, each performed in triplicate. Asterisks denote statistical differences (* p < 0.05; *** p < 0.001). Full article ">Figure 5
Effect of PE, PE/2, and PE/3 on IL6 secretion and expression in the presence of palmitic acid (PA) (a,b) or high glucose (HG) (c,d). HepG2 cells were incubated with PA (0.5 mM) for 8 h or with HG (60 mM) for 24 h with or without PE, PE/2, and PE/3 (25 µM). IL6 secretion was determined in the conditioned media by using a custom Human Magnetic Luminex Assay. IL6 mRNA levels were evaluated by real-time RT-PCR analysis. Data were normalized by the amount of beta-ACTIN, used as an internal control. Bars represent the mean ± SD of three independent experiments, each performed in triplicate. Asterisks denote statistical differences (* p < 0.05; *** p < 0.001). Full article ">Figure 6
Scheme illustrating PE, PE/2, and PE/3 action in myoblast and hepatocyte. PE/3 exhibits a high inhibitory activity towards PTP1B, which enhances the INSR/AKT pathway and leads to increased glucose uptake in muscle cells. However, considering PE/2’s inability to act through PTP1B, we may suppose that its improvement of hepatic insulin resistance could be mediated by the inhibition of IL6 (black dashed arrow). Finally, PE possibly targets both IL6 and PTPB1, hence its amelioration of the insulin pathway in both muscle and hepatic cells (red arrows refer to negative regulation, black arrows refer to positive regulation). Full article ">

14 pages, 4833 KiB

Open AccessArticle

Environmentally Friendly and Efficient Methods for Mitigating the Density of Ascidian Fouling in Mediterranean Mussel Farming

by Dimitrios Tsotsios, Dimitrios K. Papadopoulos, Maria V. Alvanou, Ioannis Georgoulis, Athanasios Lattos, Basile Michaelidis, Konstantinos Feidantsis, Ioannis A. Giantsis and John A. Theodorou

Fishes 2024, 9(4), 135; https://doi.org/10.3390/fishes9040135 - 14 Apr 2024

Cited by 3 | Viewed by 1335

Abstract

Ascidian biofouling generates significant challenges to bivalve aquaculture. Their rapid spread across almost every available surface leads to increased maintenance costs and reduced yields in shellfish farming. In addition, ascidians may introduce pathogens or toxins, further compromising the health and marketability of bivalve [...] Read more.

Ascidian biofouling generates significant challenges to bivalve aquaculture. Their rapid spread across almost every available surface leads to increased maintenance costs and reduced yields in shellfish farming. In addition, ascidians may introduce pathogens or toxins, further compromising the health and marketability of bivalve stocks and thereby necessitating strict management strategies to manage these impacts. The aim of this study was the evaluation of different management practices for eliminating ascidian fouling and the identification of the best method for Mytilus galloprovincialis (Lamarck, 1819) aquaculture farms. The effects of different anti-fouling treatments as well as their interactive outcomes were examined by conducting two experiments. Various experimental procedures were applied, including the temporally differential washing of mussels, air exposure and immersion in 50 ppt and 70 ppt salinity solutions, as well as the combination of these applications. All treatments reduced the number of ascidian colonies on mussel socks but at varying proportions. Immersing mussel socks in a 70 ppt salinity solution followed by air exposure for 1 day was the most efficient method and led to a 93% eradication. Full article

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8 pages, 29126 KiB

Open AccessInteresting Images

Unique Aggregations of a Large Undescribed Solitary Tunicate in the Arabian Sea

by Kaveh Samimi-Namin, Tito Monteiro da Cruz Lotufo, Bert W. Hoeksema, Sarah M. Tweedt, Christopher Meyer and Gustav Paulay

Diversity 2024, 16(4), 221; https://doi.org/10.3390/d16040221 - 6 Apr 2024

Viewed by 1882

Abstract

We document aggregations of an undescribed benthic solitary tunicate of the family Pyuridae from the Arabian Sea. This new genus was found forming dense thickets in shallow rocky substrates around Masirah Island and the Dhofar area in Oman. Such aggregations of tunicates have [...] Read more.

We document aggregations of an undescribed benthic solitary tunicate of the family Pyuridae from the Arabian Sea. This new genus was found forming dense thickets in shallow rocky substrates around Masirah Island and the Dhofar area in Oman. Such aggregations of tunicates have not been reported before from coral reefs in the Indo-West Pacific region and the Atlantic. This observation contributes to our understanding of the ecology and biogeography of ascidians, setting the stage for a comprehensive species description and in-depth analysis of this species. Full article

(This article belongs to the Collection Interesting Images from the Sea)

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Locations where the undescribed pyurid tunicate species was observed along the Omani coast and around Masirah Island (red dots). Blue shading represents depth gradients. Full article ">Figure 2
Dense aggregations of the undescribed pyurid tunicate species around Masirah Island, Oman. Scales = 10 cm. Note overgrowth by other organisms. (A) Individuals at the north of the island at 5 m depth. (B) Individuals at the southwest of the island at 10 m depth. (C) Close-up of an individual animal. Full article ">Figure 3
(A,B) Tunic remaining from individual attached by putative predator; note the absence of the orange tunicate body and remaining white tunic (hatched rectangle). (C) Remnant of an individual, most likely removed by predation. (D) Space left on rock by several individuals that appear to have been recently removed judging by the clean, vacant surface. (E) The individual without its thick tunic. (F) Cross-section of the individual with the body separated, scale = 5 cm. Full article ">Figure 4
Anatomical section of the preserved specimen of the undescribed pyurid tunicate species from Oman. (A) Cross-section of the individual, arrow pointing at the body wall, scale = 2 cm. (B) Dissected pharynx and body, hatched rectangles, representing (C–F) macro images, scale = 1 cm. (C) Endocarps. (D) Pharynx wall with folds, showing longitudinal vessels and 2nd, 3rd, and 4th folds. (E) External view of the oral siphon and oral tentacles. (F) Internal view of the oral siphon and oral tentacles. Full article ">

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