Protopalythoa Variabilis Research Articles

Novel neurotoxic peptides from Protopalythoa variabilis virtually interact with voltage-gated sodium channel and display anti-epilepsy and neuroprotective activities in zebrafish

We previously reported a novel toxic peptide identified from the anthozoan Protopalythoa variabilis transcriptome which is homologous to a novel structural type of sodium channel toxin isolated from a parental species (Palythoa caribaeorum). The peptide was named, according to its homologous, as Pp V-shape α-helical peptide (PpVα) in the present study. Through molecular docking and dynamics simulation, linear and hairpin folded PpVα peptides were shown to be potential voltage-gated sodium channel blockers. Nowadays, sodium channel blockers have been the mainstream of the pharmacological management of epileptic seizures. Also, sodium channel blockers could promote neuronal survival by reducing sodium influx and reducing the likelihood of calcium importation resulting in suppressing microglial activation and protecting dopaminergic neurons from degeneration. The folded PpVα peptide could decrease pentylenetetrazol (PTZ)-induced c-fos and npas4a expression level leading to reverse PTZ-induced locomotor hyperactivity in zebrafish model. In vitro, the folded PpVα peptide protected PC12 cells against 6-hydroxydopamine (6-OHDA)-induced neurotoxicity via activating heme oxygenase-1 (HO-1) and attenuating inducible nitric oxide synthase (iNOS) expression. In vivo, PpVα peptide suppressed the 6-OHDA-induced neurotoxicity on the locomotive behavior of zebrafish and, importantly, prevented the 6-OHDA-induced excessive ROS generation and subsequent dopaminergic neurons loss. This study indicates that the single S-S bond folded PpVα peptide arises as a new structural template to develop sodium channel blockers and provides an insight on the peptide discovery from cnidarian transcriptome to potentially manage epilepsy and neurodegenerative disorders.

The Holo-Transcriptome of the Zoantharian Protopalythoa variabilis (Cnidaria: Anthozoa): A Plentiful Source of Enzymes for Potential Application in Green Chemistry, Industrial and Pharmaceutical Biotechnology.

Marine invertebrates, such as sponges, tunicates and cnidarians (zoantharians and scleractinian corals), form functional assemblages, known as holobionts, with numerous microbes. This type of species-specific symbiotic association can be a repository of myriad valuable low molecular weight organic compounds, bioactive peptides and enzymes. The zoantharian Protopalythoa variabilis (Cnidaria: Anthozoa) is one such example of a marine holobiont that inhabits the coastal reefs of the tropical Atlantic coast and is an interesting source of secondary metabolites and biologically active polypeptides. In the present study, we analyzed the entire holo-transcriptome of P. variabilis, looking for enzyme precursors expressed in the zoantharian-microbiota assemblage that are potentially useful as industrial biocatalysts and biopharmaceuticals. In addition to hundreds of predicted enzymes that fit into the classes of hydrolases, oxidoreductases and transferases that were found, novel enzyme precursors with multiple activities in single structures and enzymes with incomplete Enzyme Commission numbers were revealed. Our results indicated the predictive expression of thirteen multifunctional enzymes and 694 enzyme sequences with partially characterized activities, distributed in 23 sub-subclasses. These predicted enzyme structures and activities can prospectively be harnessed for applications in diverse areas of industrial and pharmaceutical biotechnology.

Microcystins production and antibacterial activity of cyanobacterial strains of Synechocystis, Synechococcus and Romeria isolated from water and coral reef organisms of Brazilian coast

Cyanobacteria are widely distributed in terrestrial, freshwater and marine environments, and over the past decades have been recognized as a powerful source of bioactive compounds. In this study, some cyanobacterial strains were isolated from samples of seawater, brackish water and tissue of reef benthic invertebrates (zoanthid Protopalythoa variabilis, the sponges Cynachrella sp. and Haliclona sp., the coral Siderastrea stellata, and ascidians), collected at the states of Paraiba and Rio Grande do Norte (Northeast of Brazil), during the period between July 2010 and February 2014. After standard isolation methods, the cultivation of the strains was carried out in acclimatized culture chamber (25 °C) under constant aeration, for 15 days at 12-hour photoperiod, using Conway and BG11 media made with filtered seawater. After ethanolic and methanolic extracts, the strains were analysed for the microcystin production by the ELISA technique and for the antibacterial activity against Staphylococcus aureus and Pseudomonas aeruginosa by the agar well diffusion method. The detection of the mcy B gene, one of the genes related to the microcystin synthesis, was done by the Polymerase Chain Reaction (PCR) technique. The majority of the eighteen cyanobacterial strains belonged to Synechococcaceae Family. The genera of Synechocystis, Synechococcus and Romeria were represented by ten, six and two strains, respectively. The production of microcystins was observed in five strains belonging to the genus Synechocystis . The presence of mcy B gene was detected in 12 strains of cyanobacteria: Synechocystis (three strains), Synechococcus (six strains) and Romeria (two strains). Only one strain ( Synechocystis aquatilis ) showed both the microcystin production and the mcy B gene presence. The antibacterial activity was observed for one strain of Romeria gracilis , one strain of Synechocystis aquatilis and two strains of Synechococcus sp. The ethanolic extracts of R. gracilis strain and two Synechococcus spp. strains inhibited the growth of P. aeruginosa . Among methanolic extracts of cyanobacteria, only one strain of S. aquatilis showed activity against S. aureus, and one R. gracilis strain against P. aeruginosa . Some cyanobacterial strains studied were positive for the microcystin production and antibacterial activity against pathogenic bacteria S. aureus and P. aeuruginosa , and may be further explored for additional biotechnological applications.

Microcystins production and antibacterial activity of cyanobacterial strains of Synechocystis, Synechococcus and Romeria isolated from water and coral reef organisms of Brazilian coast

Cyanobacteria are widely distributed in terrestrial, freshwater and marine environments, and over the past decades have been recognized as a powerful source of bioactive compounds. In this study, some cyanobacterial strains were isolated from samples of seawater, brackish water and tissue of reef benthic invertebrates (zoanthid Protopalythoa variabilis, the sponges Cynachrella sp. and Haliclona sp., the coral Siderastrea stellata, and ascidians), collected at the states of Paraíba and Rio Grande do Norte (Northeast of Brazil), during the period between July 2010 and February 2014. After standard isolation methods, the cultivation of the strains was carried out in acclimatized culture chamber (25 °C) under constant aeration, for 15 days at 12-hour photoperiod, using Conway and BG11 media made with filtered seawater. After ethanolic and methanolic extracts, the strains were analysed for the microcystin production by the ELISA technique and for the antibacterial activity against Staphylococcus aureus and Pseudomonas aeruginosa by the agar well diffusion method. The detection of the mcyB gene, one of the genes related to the microcystin synthesis, was done by the Polymerase Chain Reaction (PCR) technique. The majority of the eighteen cyanobacterial strains belonged to Synechococcaceae Family. The genera of Synechocystis, Synechococcus and Romeria were represented by ten, six and two strains, respectively. The production of microcystins was observed in five strains belonging to the genus Synechocystis. The presence of mcyB gene was detected in 12 strains of cyanobacteria: Synechocystis (three strains), Synechococcus (six strains) and Romeria (two strains). Only one strain (Synechocystis aquatilis) showed both the microcystin production and the mcyB gene presence. The antibacterial activity was observed for one strain of Romeria gracilis, one strain of Synechocystis aquatilis and two strains of Synechococcus sp. The ethanolic extracts of R. gracilis strain and two Synechococcus spp. strains inhibited the growth of P. aeruginosa. Among methanolic extracts of cyanobacteria, only one strain of S. aquatilis showed activity against S. aureus, and one R. gracilis strain against P. aeruginosa. Some cyanobacterial strains studied were positive for the microcystin production and antibacterial activity against pathogenic bacteria S. aureus and P. aeuruginosa, and may be further explored for additional biotechnological applications.

The Transcriptome of the Zoanthid Protopalythoa variabilis (Cnidaria, Anthozoa) Predicts a Basal Repertoire of Toxin-like and Venom-Auxiliary Polypeptides.

Protopalythoa is a zoanthid that, together with thousands of predominantly marine species, such as hydra, jellyfish, and sea anemones, composes the oldest eumetazoan phylum, i.e., the Cnidaria. Some of these species, such as sea wasps and sea anemones, are highly venomous organisms that can produce deadly toxins for preying, for defense or for territorial disputes. Despite the fact that hundreds of organic and polypeptide toxins have been characterized from sea anemones and jellyfish, practically nothing is known about the toxin repertoire in zoanthids. Here, based on a transcriptome analysis of the zoanthid Protopalythoa variabilis, numerous predicted polypeptides with canonical venom protein features are identified. These polypeptides comprise putative proteins from different toxin families: neurotoxic peptides, hemostatic and hemorrhagic toxins, membrane-active (pore-forming) proteins, protease inhibitors, mixed-function venom enzymes, and venom auxiliary proteins. The synthesis and functional analysis of two of these predicted toxin products, one related to the ShK/Aurelin family and the other to a recently discovered anthozoan toxin, displayed potent in vivo neurotoxicity that impaired swimming in larval zebrafish. Altogether, the complex array of venom-related transcripts that are identified in P. variabilis, some of which are first reported in Cnidaria, provides novel insight into the toxin distribution among species and might contribute to the understanding of composition and evolution of venom polypeptides in toxiferous animals.

Distribution pattern of zoanthids (Cnidaria: Zoantharia) on a tropical reef

Coastal reef environments support a great diversity of benthic species, which are physically arrayed according to their adaptations to biotic factors such as competition and to abiotic factors such as salinity and desiccation. Few studies have described adaptive strategies on rocky shores in tropical regions. This contribution deals with the spatial distribution of three species of zoanthids (Palythoa caribaeorum, Protopalythoa variabilis and Zoanthus sociatus) on flat sandstone reefs in north-eastern Brazil. Zoanthid distribution and abundances were determined as the percentage of cover along six transects, where 50 × 50 cm quadrats were sampled. Biotic data (influence of macroalgal cover) and abiotic data (relief, substrate type, temperature and salinity) were also taken along the transects to attempt to explain the zoanthid distribution. These species tended to colonize the middle and low intertidal zones, which are least exposed to air. Zoanthus sociatus apparently resists desiccation better than P. caribaeorum. Protopalythoa variabilis showed the highest abundance and frequency, occurring in the entire middle intertidal zone and colonized areas with high proportions of unconsolidated sediment. Desiccation was the main factor limiting the spatial distributions of the three species, although substrate composition and probably interspecific competition with macroalgae were also important. The results suggest that the zoanthids use a variety of these adaptive strategies, which account for their success on consolidated substrates. Continuous monitoring of changes in distribution patterns can be useful for indicating human impacts on marine biodiversity, providing insights for monitoring and conservation programmes.

Symbiodinium diversity associated with zoanthids (Cnidaria: Hexacorallia) in Northeastern Brazil

Some marine animals have a dynamic mutualistic association involving heterotrophic coral animals (hosts) and autotrophic dinoflagellate endosymbionts in the genus Symbiodinium, known as zooxanthellae, which are directly involved in their physiology. Currently, there is limited knowledge of the Symbiodinium diversity associated with zoanthids from the Brazilian coast. Hence, this study assessed the diversity of zooxanthellae associated with Palythoa caribaeorum, Zoanthus sociatus and Protopalythoa variabilis commonly found on the northeast coast. Zoanthids samples were collected from sandstone reefs, and the total DNA from the samples was extracted. Diversity was assessed by denaturing gradient gel electrophoresis (DGGE) and ITS region cloning library. Data from ITS region sequences showed that zoanthids hosted two phylogenetic subclades or subgenus (C1 and A3) closely related to Symbiodinium from previous studies. Subclade C1 was found in Pa. caribaeorum, Pr. variabilis and Z. sociatus, while subclade A3 was found exclusively in Z. sociatus. This suggests that Pa. caribaeorum and Pr. variabilis are extremely selective, whereas Z. sociatus shows greater flexibility in the selection of its symbionts, and these differences may be involved with living in different reef environments. Knowledge of zooxanthellae diversity associated with zoanthids can explain their large distribution in Brazilian coast and contribute to a better understanding of zoanthid’s sensitivity to thermal stress.

Osmoregulation in tropical zoanthid Protopalythoa variabilis (Cnidaria: Anthozoa)

This study evaluated the physiological, morphological and ethological effects of salinity variations on the tropical zoanthid Protopalythoa variabilis. This ...

Competitive interactions among zoanthids (cnidaria: zoanthidae) in an intertidal zone of northeastern Brazil

Sessile organisms that live in consolidated substrates frequently compete for space. Coral species have many strategies to face this competition, including harming their opponents or hindering their growth. In the present study, the competitive interactions between three species of zoanthids were investigated in the intertidal zone of a sandstone reef environment in northeastern Brazil. The competitive abilities of the three species were evaluated by periodic observation of the natural fringes of contact and experimental evaluation of their growth rate through removal of 100 cm² of colonies of each species. Palythoa caribaeorum and Zoanthus sociatus had similar growth rates, and both species grew faster than Protopalythoa variabilis. The recolonization strategy seems to differ among species. The contact fringes between P. caribaeorum and Z. sociatus remained unchanged over time, without any type of aggressive interaction between them, suggesting that stand-off was the strategy used by these organisms. Palythoa caribaeorum and Z. sociatus grew among the polyps of P. variabilis, often killing its colonies. The coexistence of zoanthids reveals a capacity for survival in the face of competition for limited resources such as free substrate, which led to the colonization and establishment of zoanthids in intertidal environments.

Palyosulfonoceramides A and B: Unique Sulfonylated Ceramides from the Brazilian Zoanthids Palythoa caribaeorum and Protopalyhtoa variabilis

The zoanthids Palythoa caribaeorum and Protopalythoa variabilis are among the most abundant marine species along the Brazilian coast. We now report the isolation and structure elucidation of two unprecedented sulfonylated ceramides, palyosulfonoceramide A (1) and palyosulfonoceramide B (2) from specimens collected off Brazil’s northeastern coast. The structures of 1 and 2 were established using a combination of NMR analyses, including: evaluation of 1H, 13C, 1H–1H COSY, 1H–13C HSQC, 1H–13C HMBC, and 1H–15N HMBC NMR spectra, high-resolution mass spectrometry and chemical degradation. In addition, we also isolated the corresponding known ceramides, N-((2S,3R,4E,8E)-1,3-dihydroxyoctadeca-4,8-dien-2-yl)-hexadecanamide (3) and N-((2S,3R,4E)-1,3-dihydroxyoctadeca-4-en-2-yl)-hexadecanamide (4), which provided further support for the assignments of 1 and 2.

Pro-apoptotic activity of lipidic α-amino acids isolated from Protopalythoa variabilis

Lipidic α-amino acids (LAAs) have been described as non-natural amino acids with long saturated or unsaturated aliphatic chains. In the continuing prospect to discover anticancer agents from marine sources, we have obtained a mixture of two cytotoxic LAAs ( 1a and 1b) from the zoanthid Protopalythoa variabilis. The anti-proliferative potential of 14 synthetic LAAs and 1a/ 1b were evaluated on four tumor cell lines (HCT-8, SF-295, MDA-MB-435, and HL-60). Five of the synthetic LAAs showed high percentage of tumor cell inhibition, while 1a/ 1b completely inhibited tumor cell growth. Additionally, apoptotic effects of 1a/ 1b were studied on HL-60 cell line. 1a/ 1b-treated cells showed apoptosis morphology, loss of mitochondrial potential, and DNA fragmentation.

Cytotoxic lipidic α-amino acids from the zoanthid Protopalythoa variabilis from the Northeastern coast of Brazil

Two lipidic α-amino acids 1a and 1b were isolated from the zoanthid Protopalythoa variabilis using a bioguided fractionation for cytotoxic activity. The structures of the metabolites were determined by spectroscopic methods, including NMR (nuclear magnetic resonance) 1H e 13C, IR (infrared) and high resolution mass spectrometry (positive mode). The cytotoxic activity of the crude extract, as well as of the mixture of 1a and 1b were measured in vitro using the MTT assay for four human tumor cell lines. This finding has important biological and chemical implications for this type of compound. This is the first report of lipidic α-amino acids from natural sources, as well as of their cytotoxic activity.

A new cytotoxic 2-amino-n-alkyl-carboxylic acid mixture obtained from the zoanthid Protopalythoa variabilis collected at Paracuru beach, Ceará State, Brazil

This work performed a cytotoxicity-guided fractionation of a hydromethanolic extract obtained from the zoanthid Protopalythoa variabilis (PV-HM) collected at Paracuru beach (Ceará State, Brazil). The cytotoxicity of samples was evaluated by the MTT assay against 4 human tumor cell lines: HL-60 (leukemia), MDA MB-435 (melanoma), SF-295 (CNS glioblastoma) and HCT-8 (colon). The PV-HM and its 4 derived fractions (PV-MeOH; -Hex; -EtOAc; -DCM) were cytotoxic against all cell lines tested (IC50 ranging from 0.36 to 4.24µg/mL for PV-HM and from 0.09 to 2.9 for fractions). PV-Hex, the most potent, was purified by successive partitions with acetonitrile:water and pyridine, yielding PV-Hex-SIIS. For structure elucidation, NMR, MS and IR analysis were performed. PV-Hex-SIIS correspond to a new 2-amino-n-alkyl-carboxylic acid mixture of two compounds differing from each other by one methylene among their aliphatic chains (figure1). Preliminary analysis by flow cytometry and morphological observations of HL-60 cells treated with PV-Hex-SIIS indicates that cells are suffering apoptosis by the intrinsic pathway and showed intense cell membrane disruption. Further studies are necessary to elucidate the specific mechanism of action of these compounds.

Reproductive biology of Palythoa caribaeorum and Protopalythoa variabilis (Cnidaria, Anthozoa, Zoanthidea) from the southeastern coast of Brazil

The reproductive biology of Palythoa caribaeorum (Duchassaing & Michelotti 1860) and Protopalythoa variabilis (Duerden 1898) was studied through monthly samples from tagged colonies from June 1996 to June 1997, in Sao Sebastiao channel, São Paulo, Brazil (45 degrees 26'W, 23 degrees 50'S). The gametogenesis was similar to that of other zoanthids as shown by histological preparations. Oocyte diameters and maturation stages of testis vesicles were evaluated on squash preparations. Both species showed sequential protogynic hermaphroditism, with high frequency of fertile polyps (83% in P. variabilis and 72% in P. caribaeorun), high frequency of colonies in female sex condition (65.3% of P variabilis and 41.7% of P caribaeorum), and apparently continuous gametogenesis. In P. caribaribaeonrum, egg release was continuous and sperm release took place during half of the analyzed period. In P. variabilis, egg and sperm release occurred in April-May and February-March 1997, respectively.