Marine Ascidian Research Articles

Anti-Trichomonas vaginalis Activity of Marine Ascidians (Tunicates; Ascidiacea) from the Bushehr Province, Iran.

The aim of the current research is to evaluate the antiparasite effects of compounds isolated from marine ascidian tunicates on Trichomonas vaginalis. Ascidian tunicates after collection were cut into small pieces, freeze-dried, and powdered. The resulting material was subjected to extraction in double-distilled water, ethanol, n-hexane, and dichloromethane. To fractionate the extracts and identify the most bioactive compound, silica gel column chromatography and GC-M/S analysis were used. Fraction 18 of silica gel column chromatography of ethanol extract was the most effective against T. vaginalis. The respective IC50, CC50, and SI values for fraction 18 were 28.62 μg/mL, ˃800 μg/mL, and ˃27.95. GC-M/S analysis of this fraction identified a major phenolic compound (2, 4-bis (1, 1-dimethyl ethyl), whose toxicity against vero cells was only 10.15%. The ethanolic fraction containing phenol-2,4-bis (1,1-dimethylethyl), which has a potent lethality effect on T. vaginalis, may be considered as an antiparasite drug candidate.

The unknown unknown: A framework for assessing environmental DNA assay specificity against unsampled taxa.

Taxon-specific quantitative PCR (qPCR) assays are commonly used for environmental DNA sampling-based inference of animal presence. These assays require thorough validation to ensure that amplification truly indicates detection of the target taxon, but a thorough validation is difficult when there are potentially many non-target taxa, some of which may have incomplete taxonomies. Here, we use a previously published, quantitative model of cross-amplification risk to describe a framework for assessing qPCR assay specificity when there is missing information and it is not possible to assess assay specificity for each individual non-target confamilial. In this framework, we predict assay specificity against unsampled taxa (non-target taxa without sequence data available) using the sequence information that is available for other confamilials. We demonstrate this framework using four case study assays for: (1) An endemic, freshwater arthropod (meltwater stonefly; Lednia tumana), (2) a globally distributed, marine ascidian (Didemnum perlucidum), (3) a continentally distributed freshwater crustacean (virile crayfish; Faxonius virilis, deanae and nais species complex) and (4) a globally distributed freshwater teleost (common carp; Cyprinus carpio and its close relative C. rubrofuscus). We tested the robustness of our approach to missing information by simulating application of our framework for all possible subsamples of 20-all non-target taxa. Our results suggest that the modelling framework results in estimates which are largely concordant with observed levels of cross-amplification risk using all available sequence data, even when there are high levels of data missingness. We explore potential limitations and extensions of this approach for assessing assay specificity and provide users with an R Markdown template for generating reproducible reports to support their own assay validation efforts.

Anti-Mycobacterial Activity of Polycarpine and Polycarpaurine A from an Indonesia Marine Ascidian Polycarpa sp

Anti-Mycobacterial Activity of Polycarpine and Polycarpaurine A from an Indonesia Marine Ascidian Polycarpa sp

Anti-Mycobacterial Activity of Polycarpine and Polycarpaurine A from an Indonesia Marine Ascidian Polycarpa sp.

Anti-Mycobacterial Activity of Polycarpine and Polycarpaurine A from an Indonesia Marine Ascidian Polycarpa sp.

Rare Caulamidine Hexacyclic Alkaloids from the Marine Ascidian Polyandrocarpa sp.

Two new caulamidines C (2) and D (4) and three isocaulamidines B, C, and D (1, 3, and 5) along with the known compound caulamidine B (6) were isolated from the marine ascidian Polyandrocarpa sp. Their structures were elucidated by analysis of nuclear magnetic resonance (NMR) and electronic circular dichroism (ECD) data. Isocaulamidines have an altered pattern of N-methyl substitution (N-15 vs N-13 in the caulamidines) with a concomitant double-bond rearrangement to provide a new C-14/N-13 imine functionality. Caulamidine C (2) and isocaulamidine C (3) are the first members of this alkaloid family with two chlorine substituents in the core 6H-2,6-naphthyridine ring system.

Import and Implications of Vanadium in Live Aspects

In Earth’s regions accessible for living organisms (Earth’s crust, crude oil, water sanctuaries and lower atmosphere), vanadium is present in the oxidation states +III and—essentially—+IV (cationic) and +V (cationic and anionic), with the redox interchange and biochemical recycling often monitored by bacteria. Organisms having available vanadium-containing (bio)molecules with essential functions for life include marine brown algae (haloperoxidases), ascidians and fan worms, as well as terrestrial organisms, viz., nitrogen-fixing bacteria (associated with the roots of legumes), and the fly agaric mushroom. The hypohalite generated by the algal haloperoxidases in turn is involved in the emission of bromoform into the atmosphere. Nitrogen fixation (N2 ε NH4+) is a process of immanent importance for life on our planet. Other bacterial issues include the reduction of vanadate to VO2+. Medicinal applications of vanadium coordination compounds are directed towards the treatment of diabetes mellitus (vanadium complexes with hypoglycemic activity) and cancer—although boundaries are set due to side effects such as oxidative damage elicited by vanadium-induced hyperoxide formation. Physiological actions of vanadium are often invoked due to the structural and physiological similarity between vanadate and phosphate. An additional field of medicinal applications addresses the treatment of cancer, such as leukaemia, malignant melanoma and bone cancer.

Short-Term Exposure to Nanoplastics Does Not Affect Bisphenol A Embryotoxicity to Marine Ascidian Ciona robusta.

Plastic pollution is recognized as a global environmental threat and concern is increasing regarding the potential interactions of the smallest fragments, nanoplastics (1 µm), with either physical and chemical entities encountered in the natural environment, including toxic pollutants. The smallest size of nanoplastics (<100nm) rebounds to their safety associated with remarkable biological, chemical and physical reactivity that allow them to interact with cellular machinery by crossing biological barriers and causing damage to living beings. Recent findings on nanoplastic occurrence in marine coastal waters, including the Mediterranean Sea, leave open the question on their ability to act as a vector of other contaminants of emerging concerns (CECs) concomitantly released by wastewater treatment plants and reaching marine coastal waters. Here, we assess for the first time the role of non-functionalized polystyrene nanoparticles (PS NPs, 20 nm) as a proxy for nanoplastics (1 and 10 µg/mL) alone and in combination with bisphenol A (BPA) (4.5 and 10 µM) on Ciona robusta embryos (22 h post fertilization, hpf) by looking at embryotoxicity through phenotypic alterations. We confirmed the ability of BPA to impact ascidian C. robusta embryo development, by affecting sensory organs pigmentation, either alone and in combination with PS NPs. Our findings suggest that no interactions are taking place between PS NPs and BPA in filtered sea water (FSW) probably due to the high ionic strength of seawater able to trigger the sorption surface properties of PS NPs. Further studies are needed to elucidate such peculiarities and define the risk posed by combined exposure to BPA and PS NPs in marine coastal waters.

Meridianins Inhibit GSK3β In Vivo and Improve Behavioral Alterations Induced by Chronic Stress.

Major depression disorder (MDD) is a severe mental alteration with a multifactorial origin, and chronic stress is one of the most relevant environmental risk factors associated with MDD. Although there exist some therapeutical options, 30% of patients are still resistant to any type of treatment. GSK3β inhibitors are considered very promising therapeutic tools to counteract stress-related affectations. However, they are often associated with excessive off-target effects and undesired secondary alterations. Meridianins are alkaloids with an indole framework linked to an aminopyrimidine ring from Antarctic marine ascidians. Meridianins could overcome several of the aforementioned limitations since we previously demonstrated that they can inhibit GSK3β activity without the associated neurotoxic or off-target effects in rodents. Here, we show that meridianins delivered into the lateral ventricle inhibited GSK3β in several brain regions involved with stress-related symptoms. We also observed changes in major signaling pathways in the prefrontal cortex (Akt and PKA) and hippocampus (PKC and GluR1). Moreover, meridianins increased synaptic activity, specifically in the CA1 but not in the CA3 or other hippocampal subfields. Finally, we chronically treated the mice subjected to an unpredictable mild chronic stress (CUMS) paradigm with meridianins. Our results showed improvements produced by meridianins in behavioral alterations provoked by CUMS. In conclusion, meridianins could be of therapeutic interest to patients with stress-related disorders such as MDD.

Cultivation of gut microorganisms of the marine ascidian Halocynthia roretzi reveals their potential roles in the environmental adaptation of their host

It has long been known that abundant symbiotic bacteria exist in the tunic and gut of marine ascidians, and that these play crucial roles in host development, physiological metabolism, and environmental adaptation. However, the identity, roles and functions of these symbiotic bacteria are known for only a few strains. In this study, we isolated and cultivated 263 strains of microorganisms from the intestine of the marine ascidian Halocynthia roretzi through a combination of aerobic and anaerobic culture approaches. Most cultivated species, both aerobic and anaerobic, from ascidian stool samples belonged to the genus Bacillus based on 16S rDNA sequencing identification and phylogenetic assays. The distribution of cultured bacteria varied with seasonal changes in environmental conditions. To explore the functions of cultured bacteria, we screened out a strain of Serratia sp. whose extracts showed high antibacterial activity against aquatic pathogens. These findings revealed the potential roles of gut microorganisms in ascidian defense and environmental adaptation, thus providing insights into the interaction and co-evolution between gut bacteria and their hosts.

Aqueous Extract of Sea Squirt (Halocynthia roretzi) with Potent Activity against Human Cancer Cells Acts Synergistically with Doxorubicin.

Marine ascidian is becoming one of the main sources of an antitumor drug that has shown high bioactivity and extensive application in cancer treatment. Halocynthia roretzi, an edible marine sea squirt, has been demonstrated to have various kinds of biological activities, such as anti-diabetic, anti-hypertension, and enhancing immunity. In this study, we reported that aqueous extracts from the edible parts of H. roretzi presented significantly inhibiting the efficiency on HepG-2 cell viability. The separate mixed compound exhibited strong effects of inhibitory proliferation and induced apoptosis via the generation of ROS along with the concurrent loss of mitochondrial membrane potential on tumor cells. Furthermore, we found that there existed a significantly synergistic effect of the ascidian-extracted compound mixture with the anti-cancer drug doxorubicin. In the presence of the extracts from H. roretzi, the dose of doxorubicin at the cellular level could be reduced by a half dose. The extracts were further divided by semipreparative-HPLC and the active ingredients were identified as a mixture of fatty amide, which was composed of hexadecanamide, stearamide, and erucamide by UHPLC-MS/MS. Our results suggest that the potential toxicity of ascidian H. roretzi in tumor cells, and the compounds extracted from H. roretzi could be potentially utilized on functional nutraceuticals or as an adjunct in combination with chemotherapy.

One-Pot Green Synthesis of Acridine Alkaloid Derivatives and Screening of in vitro Anti-cancer Activity Against Cdc25b and SHP1.

To develop anti-cancer active pharmaceutical intermediates. Acridone derivatives possess a wide range of pharmacological activities: 1) they intercalate DNA and 2) form a covalent bond with DNA. To screen in vitro anti-cancer activity against Cdc25b and SHP1 of new acridone derivatives and preliminary study on the structure-activity relationship. The synthesis of new acridone derivatives and in vitro evaluation of their anti-cancer activity on Cdc25b and SHP1 was achieved. Natural products that contain acridine structures, such as cystodytin A and acronycine, are isolated from certain marine (tunicates & ascidians, sponges, sea anemones) and plant (bark of Australian scrub ash tree) species. Herein, we report the efficient one-pot green synthesis of twelve novel 3,4-dihydro-1 (2H) acridone derivatives, using montmorillonite K10 as the catalyst and iron/citric acid in water. Also, their inhibitory activity against Cdc25B and SHP1 is examined, in which specific derivatives show enhanced inhibitory activity compared to others. Twelve new acridone derivatives were prepared, starting from 2-nitrobenzaldehyde derivatives and 1, 3-cyclohexanedione derivatives, which exhibited substantial anti-cancer activity against Cdc25b and SHP1 cells. Preliminary studies on the structure-activity relationship have shown the influence of the structural parameters and, in particular, the nature of the substituent on aromatic ring structure and cyclohexanone. Other: Further study on the structure-activity relationship is required.

Research Progress on Secondary Metabolites from the Ascidians of Aplidium Genus

Abstract: Ascidians (tunicates) are widely recognized as one of the most prolific producers of bioactive natural products in the marine environment. This present study reviewed the chemical diversity of marine ascidians from the Aplidium genus and their pharmacological applications since Jan 2005. The resources of this genus from China Seas, including the changes of their names in the family Polydinidae were also summarized in this paper. In addition, a concise outlook on their chemical and pharmaceutical investigation is made to support further development.

Existence in cellulose shelters: industrial and pharmaceutical leads of symbiotic actinobacteria from ascidian Phallusia nigra, Andaman Islands.

The diversity of actinobacteria associated with marine ascidian Phallusia nigra from Andaman Islands was investigated. A total of 10 actinobacteria were isolated and based on the biochemical and molecular characterization, the isolates were assigned to 7 different actinobacterial genera. Eight putatively novel species belonging to genera Rhodococcus, Kineococcus, Kocuria, Janibacter, Salinispora and Arthrobacter were identified based on 16S rDNA sequence similarity with the NCBI database. The organic extracts of ten isolates displayed considerable bioactivity against test pathogens, which were Gram-positive and Gram-negative in nature. PCR-based screening for type I and type II polyketide synthases (PKS-I, PKS-II) and nonribosomal peptide synthetases (NRPS) revealed that, 10 actinobacterial isolates encoded at least one type of polyketide synthases biosynthesis gene. Majority of the isolates found to produce industrially important enzymes; amylase, protease, gelatinase, lipase, DNase, cellulase, urease, phosphatase and L-asparaginase. The present study emphasized that, ascidians are a prolific resource for novel bioactive actinobacteria with potential for novel drug discovery. This result expands the scope to functionally characterize the novel ascidian associated marine actinobacteria and their metabolites could be a source for the novel molecules of commercial interest.

Taxonomic Composition and Biological Activity of Bacterial Communities Associated with Marine Ascidians from Andaman Islands, India.

Marine invertebrates, particularly ascidians, constitute an important source of potential active and biofunctional natural products. The microbial diversity associated with ascidians is little recognized, although these microorganisms play a vital role in marine ecosystems. The objective of this study was to investigate bacterial population diversity in four ascidian samples: Phallusia nigra, Phallusia fumigata, Eudistoma viride, and Rhopalaea macrothorax, collected from the North Bay, Andaman and Nicobar Islands. Microbial strains identified up to the species level revealed 236 distinct species/ribotypes out of 298 bacterial strains. Of 298 ascidian-associated bacteria, 72 isolates belong to the class Gammaproteobacteria and the genus Endozoicomonas. The results from this investigation will contribute a broaden knowledge of microbial diversity associated to marine ascidians, and as a promising source for the discovery of new natural products.

Self-organized optimal packing of kinesin-5-driven microtubule asters scales with cell size.

Radial microtubule (MT) arrays or asters determine cell geometry in animal cells. Multiple asters interacting with motors, such as those in syncytia, form intracellular patterns, but the mechanical principles behind this are not clear. Here, we report that oocytes of the marine ascidian Phallusia mammillata treated with the drug BI-D1870 spontaneously form cytoplasmic MT asters, or cytasters. These asters form steady state segregation patterns in a shell just under the membrane. Cytaster centers tessellate the oocyte cytoplasm, that is divide it into polygonal structures, dominated by hexagons, in a kinesin-5-dependent manner, while inter-aster MTs form 'mini-spindles'. A computational model of multiple asters interacting with kinesin-5 can reproduce both tessellation patterns and mini-spindles in a manner specific to the number of MTs per aster, MT lengths and kinesin-5 density. Simulations predict that the hexagonal tessellation patterns scale with increasing cell size, when the packing fraction of asters in cells is ∼1.6. This self-organized in vivo tessellation by cytasters is comparable to the 'circle packing problem', suggesting that there is an intrinsic mechanical pattern-forming module that is potentially relevant to understanding the role of collective mechanics of cytoskeletal elements in embryogenesis. This article has an associated First Person interview with the first author of the paper.

Anticancer activity in HeLa and MCF-7 cells via apoptopic cell death by a sterol molecule Cholesta-4,6-dien-3-ol (EK-7), from the marine ascidian Eudistoma kaverium

ObjectiveEudistoma kaverium, an ascidian belonging to the family Polycitoridae is previously recorded from Indian waters. The work done on this species is limited only to taxonomy and ecology. As ascidians are potential source of anticancer compounds, we have purified fractions of E. kaverium using HPLC and evaluated its anticancer activity. MethodChromatography based fractionation of the crude extract was carried out and mass spectrometry was used to identify the active molecule. Cell viability was checked using MTT assay. Flow cytometry, ROS generation, Hoechst staining and DNA fragmentation was performed to confirm the anticancer activity of the compound. ResultsA column fraction EK-7, which was identified as a sterol Cholesta-4,6-dien-3-ol using GC–MS analysis, exhibited potent cytotoxicity with IC50 value of 6.5 μM in MCF-7 and 10.2 μM against HeLa cells. Moreover, EK-7 triggered the cell cycle arrest in the sub G0/G1 stage and induced apoptosis twenty-four hours post-treatment. Hoechst staining of EK-7-treated HeLa cells portrayed apoptotic events such as changes in cell morphology, chromatin condensation, membrane swelling, and development of apoptotic bodies. The changes in light scattering by the EK-7-treated HeLa cells indicate the general characteristics of cell death as a result of apoptosis. The EK-7-treated HeLa cells show cell cycle arrest in G0/G1, G2/M and S phases and the increased amount of sub G0/G1 population indicates the increase in the apoptotic induced cell population. The isolated compound Cholesta-4,6-dien-3-ol (EK-7), based on the GC–MS analysis and library search from E. kaverium displayed potent anticancer activity which is mediated through apoptosis targeting the G0/G1 phase. ConclusionBased onthe experimental data, it shows that EK-7(Cholesta-4,6-dien-3-ol) a sterol derivative has potent anticancer activity mediated through apoptosis. If this research is continued further with preclinical and clinical trials, EK-7(Cholesta-4,6-dien-3-ol) could be possibly used as a chemotherapeutic agent in treating cancer.

MiR-92 Family Members Form a Cluster Required for Notochord Tubulogenesis in Urochordate Ciona savignyi.

MicroRNAs are frequently clustered in the genome and polycistronically transcribed, regulating targeted genes in diverse signaling pathways. The miR-17-92 cluster is a typical miRNA cluster, playing crucial roles in the organogenesis and homeostasis of physiological processes in vertebrates. Here, we identified three miRNAs (csa-miR-92a, csa-miR-92b, and csa-miR-92c) that belonged to the miR-92 family and formed a miRNA cluster in the genome of a urochordate marine ascidian Ciona savignyi. Except for miR-92a and miR-92b, other homologs of the vertebrate miR-17-92 cluster members could not be identified in the Ciona genome. We further found that the mature sequences of urochordate miR-92 family members were highly conserved compared with the vertebrate species. The expression pattern revealed that three miR-92 family members had consistent expression levels in adult tissues and were predominantly expressed in heart and muscle tissue. We further showed that, at the embryonic and larval stages, csa-miR-92c was expressed in the notochord of embryos during 18–31 h post fertilization (hpf) by in situ hybridization. Knockout of csa-miR-92c resulted in the disorganization of notochord cells and the block of lumen coalescence in the notochord. Fibroblast growth factor (FGF), mitogen-activated protein kinase (MAPK), and wingless/integrated (Wnt)/planar cell polarity (PCP) signaling pathways might be involved in the regulatory processes, since a large number of core genes of these pathways were the predicted target genes of the miR-92 family. Taken together, we identified a miR-92 cluster in urochordate Ciona and revealed the expression patterns and the regulatory roles of its members in organogenesis. Our results provide expression and phylogenetic data on the understanding of the miR-92 miRNA cluster’s function during evolution.

The Ascidian-Derived Metabolites with Antimicrobial Properties.

Among the sub-phylum of Tunicate, ascidians represent the most abundant class of marine invertebrates, with 3000 species by heterogeneous habitat, that is, from shallow water to deep sea, already reported. The chemistry of these sessile filter-feeding organisms is an attractive reservoir of varied and peculiar bioactive compounds. Most secondary metabolites isolated from ascidians stand out for their potential as putative therapeutic agents in the treatment of several illnesses like microbial infections. In this review, we present and discuss the antibacterial activity shown by the main groups of ascidian-derived products, such as sulfur-containing compounds, meroterpenes, alkaloids, peptides, furanones, and their derivatives. Moreover, the direct evidence of a symbiotic association between marine ascidians and microorganisms shed light on the real producers of many extremely potent marine natural compounds. Hence, we also report the antibacterial potential, joined to antifungal and antiviral activity, of metabolites isolated from ascidian-associate microorganisms by culture-dependent methods.

Marine Organisms from the Yucatan Peninsula (Mexico) as a Potential Natural Source of Antibacterial Compounds.

A total of 51 sponges (Porifera) and 13 ascidians (Chordata) were collected on the coast of the Yucatan Peninsula (Mexico) and extracted with organic solvents. The resulting extracts were screened for antibacterial activity against four multidrug-resistant (MDR) bacterial pathogens: the Gram-negative Acinetobacter baumannii, Klebsiella pneumoniae, and Pseudomonas aeruginosa and the Gram-positive Staphylococcus aureus. The minimum inhibitory concentrations (MICs) of the organic extracts of each marine organism were determined using a broth microdilution assay. Extracts of eight of the species, in particular the Agelas citrina and Haliclona (Rhizoniera) curacaoensis, displayed activity against some of the pathogens tested. Some of the extracts showed similar MIC values to known antibiotics such as penicillins and aminoglycosides. This study is the first to carry out antimicrobial screening of extracts of marine sponges and ascidians collected from the Yucatan Peninsula. Bioassay-guided fractionation of the active extracts from the sponges Amphimedon compressa and A. citrina displayed, as a preliminary result, that an inseparable mixture of halitoxins and amphitoxins and (-)-agelasine B, respectively, are the major compounds responsible for their corresponding antibacterial activities. This is the first report of the antimicrobial activity of halitoxins and amphitoxins against major multidrug-resistant human pathogens. The promising antibacterial activities detected in this study indicate the coast of Yucatan Peninsula as a potential source of a great variety of marine organisms worthy of further research.

Synthesis and Cytotoxic Activity of Novel Mono- and Bis-Indole Derivatives: Analogues of Marine Alkaloid Nortopsentin.

The oceans cover more than 70% of the earth's surface, which represents over 95% of the biosphere. Therefore, oceans provide a wealth of marine invertebrates, especially sponges, ascidians, bryozoans and molluscs that produce structurally unique bioactive metabolites such as alkaloids. The bioactive scaffolds of marine alkaloids exhibit cytotoxic activities against human cancer cell lines. To prepare analogues of the marine alkaloid nortopsentin [having 2,4-bis(3'- indolyl)imidazole scaffold] as cytotoxic agents via structural modification of the core imidazole ring and one of the side indole rings. Four series of nortopsentin analogues were synthesized in which the imidazole ring was replaced by pyrazole, pyrido[2,3-d]pyrimidinone and pyridine rings. Furthermore, one of the side indole rings was replaced by substituted phenyl moiety. The target compounds were tested for their in vitro cytotoxic activity against HCT-116 cell-line and the most potent compound was subjected to further investigation on its effect on HCT-116 cell cycle progression. The cytotoxic screening of the synthesized compounds revealed that bis-indolylpyridinedicarbonitriles 8a-d exhibited the most potent cytotoxic activity with IC50=2.6-8.8 μM. Compound 8c was further tested by flow cytometry analysis to explore its effect on HCT-116 cell cycle progression that, in turn, indicated its anti-proliferative effect. Marine-derived bis-indole alkaloids (nortopsentins) have emerged as a new class of indole-based antitumor agents. The design of new analogues involved several modifications in order to obtain more selective and potent cytotoxic agents. Indole derivatives bearing a pyridine core displayed more potent cytotoxic activity than those containing pyrido[2,3-d]pyrimidin-4(1H)-one moiety.