Acetylcholinesterase activity in Clytia hemisphaerica (Cnidaria)

Cholinesterase activity is known in representatives of all living organisms phyla but the origin of the cholinergic system as known in bilaterian animals is still undeciphered. In particular the implication of cholinesterases in the nervous system of non-bilaterian Metazoa is not well known. We thus chose to investigate this activity in the Clytia hemisphaerica (Cnidaria) medusa. In toto histochemical staining revealed an acetylcholinesterase activity in the tentacle bulbs but not in the nervous system. Sequences homologous to acetylcholinesterase were searched within Clytia ESTs and compared to other sequences found in public databases.     

Spatiotemporal characteristics and mechanisms of intracellular Ca(2+) increases at fertilization in eggs of jellyfish (Phylum Cnidaria, Class Hydrozoa)

We have clarified, for the first time, the spatiotemporal patterns of intracellular Ca(2+) increases at fertilization and the Ca(2+)-mobilizing mechanisms in eggs of hydrozoan jellyfish, which belong to the evolutionarily old diploblastic phylum, Cnidaria. An initial Ca(2+) increase just after fertilization took the form of a Ca(2+) wave starting from one cortical region of the egg and propagating to its antipode in all of four hydrozoan species tested: Cytaeis uchidae, Cladonema pacificum, Clytia sp., and Gonionema vertens. The initiation site of the Ca(2+) wave was restricted to the animal pole, which is known to be the only area of sperm-egg fusion in hydrozoan eggs, and the wave propagating velocity was estimated to be 4.2-5.9 mum/s. After a Ca(2+) peak had been attained by the initial Ca(2+) wave, the elevated Ca(2+) gradually declined and returned nearly to the resting value at 7-10 min following fertilization. Injection of inositol 1,4,5-trisphosphate (IP(3)), an agonist of IP(3) receptors (IP(3)R), was highly effective in inducing a Ca(2+) increase in unfertilized eggs; IP(3) at a final intracellular concentration of 12-60 nM produced a fully propagating Ca(2+) wave equivalent to that observed at fertilization. In contrast, a higher concentration of cyclic ADP-ribose (cADPR), an agonist of ryanodine receptors (RyR), only generated a localized Ca(2+) increase that did not propagate in the egg. In addition, caffeine, another stimulator of RyR, was completely without effect. Sperm-induced Ca(2+) increases in Gonionema eggs were severely affected by preinjection of heparin, an inhibitor of Ca(2+) release from IP(3)R. These results strongly suggest that there is a well-developed IP(3)R-, but not RyR-mediated Ca(2+) release mechanism in hydrozoan eggs and that the former system primarily functions at fertilization. Our present data also demonstrate that the spatial characteristics and mechanisms of Ca(2+) increases at fertilization in hydrozoan eggs resemble those reported in higher triploblastic animals.     

Association of a luminous Vibrio sp., taxonomically related to Vibrio harveyi, with Clytia linearis (Thornely, 1900) (Hydrozoa, Cnidaria)

A previously unknown association between a luminous Vibrio sp., taxonomically related to the species Vibrio harveyi and a common member of the shallow/mid water communities of the Mediterranean Sea, the hydrozoan Clytia linearis is described. All the specimens of C. linearis observed under blue light excitation showed both a natural luminescence appearing as a series of fine dots due to clytin, and a clear fluorescence on the external side of the perisarc around the colonies due to the presence of luminous bacteria. Luminous bacteria were isolated from the surface of C. linearis, their phenotypic characterization as isolates was performed by several morphological, biochemical, and cultural tests, completed with 16S rDNA sequence analysis. All the isolates were referred to a Vibrio sp. taxonomically related to V. harveyi. The association of the V. harveyi-related species with C. linearis, as already suggested for another hydroid, Aglaophenia octodonta, could be explained with the activity of these bacteria of feeding on the chitinous structures present in these hydroids. Moreover, the adhesion of the luminous bacterium (here referred to as Vibrio sp. CL1) on C. linearis may contribute to the survival of this Vibrio species in the marine environment providing a suitable growth habitat.     

The distribution of hydroids (Cnidaria, Hydrozoa) from micro- to macro-scale: Spatial patterns on habitat-forming algae

Scaling up from local, short-term experiments to larger-area and longer-term ones is crucial to address the role of scale in ecology. Few studies, however, examined large-scale spatial variability in the distribution and abundance of marine organisms, with rare attempts to directly compare spatial variation at local (centimetres-metres) vs. regional (1000's of kilometres) scale. Here, we used a hierarchical design to describe the spatial distribution of the hydroids epiphitic of the brown alga Cystoseira amentacea, a habitat-forming species that provides a continuous, extensive settling substrate at regional scale along the rocky coasts in the Mediterranean Sea. This continuity provides the potential to deal with scale-related variability, increasing area of investigation without adding differences deriving from habitat heterogeneity or changes in topographic complexity. Hydroids were selected for their abundance and for their life cycle features (rapid growth, small body size, early sexual or asexual reproduction and short life span), allowing rapid responses to changes in environmental conditions. The aim of this study was to analyse whether the structure of hydroid assemblages living on C. amentacea had a consistent pattern of variation among three portions of the algal thallus (i.e., basal, middle, and distal) across a spectrum of scales and whether having or not a pelagic stage could exert a significant influence on the distribution patterns of the species. A total of 32 species were identified. Multivariate analyses showed that hydroid colonization of Cystoseira occurs differently along each thallus, with patterns of variation in the structure of assemblages differing at an even smaller spatial scale than that of single plants. However, such differences varied from patch to patch. Among the 14 species identified as “important” to define the hydroid assemblage inhabiting Cystoseira, only one (Clytia hemisphaerica) has free medusae, the other species reproducing by fixed gonophores or by short-lived medusoids. Univariate analysis showed significant differences among portions of thalli in terms of spatial variability at the various scales investigated, thus suggesting that patterns of multivariate variation along the three portions of thalli might vary across scale. Overall, our results suggest that patterns of distribution of hydroids along C. amentacea thalli significantly vary across spatial scales but that the observed differences can be hardly interpreted on the basis of life-cycle patterns.     

Migration and differentiation potential of stem cells in the cnidarian Hydractinia analysed in eGFP-transgenic animals and chimeras

To analyse cell migration and the differentiation potential of migratory stem cells in Hydractinia, we generated animals with an eGFP reporter gene stably expressed and transmitted via the germline. The transgene was placed under the control of two different actin promoters and the promoter of elongation factor-1α. One actin promoter (Act-II) and the EF-1α promoter enabled expression of the transgene in all cells, the other actin promoter (Act-I) in epithelial and gametogenic cells, but not in the pluripotent migratory stem cells. We produced chimeric animals consisting of histocompatible wild type and transgenic parts. When the transgene was under the control of the epithelial cell specific actin-I promoter, non-fluorescent transgenic stem cells immigrated into wild type tissue, stopped migration and differentiated into epithelial cells which then commenced eGFP-expression. Migratory stem cells are therefore pluripotent and can give rise not only to germ cells, nematocytes and nerve cells, but also to epithelial cells. While in somatic cells expression of the act-I promoter was restricted to epithelial cells it became also active in gametogenesis. The act-I gene is expressed in spermatogonia, oogonia and oocytes. In males the expression pattern showed that migratory stem cells are the precursors of both the spermatogonia and their somatic envelopes. Comparative expression studies using the promoters of the actin-II gene and the elongation factor-1α gene revealed the potential of transgenic techniques to trace the development of the nervous system.     

Induction of metamorphosis from the larval to the polyp stage is similar in Hydrozoa and a subgroup of Scyphozoa (Cnidaria, Semaeostomeae)

Larvae of cnidarians need an external cue for metamorphosis to start. The larvae of various hydrozoa, in particular of Hydractinia echinata, respond to Cs⁺, Li⁺, NH₄ ⁺ and seawater in which the concentration of Mg²⁺ ions is reduced. They further respond to the phorbolester, tetradecanoyl-phorbol-13-acetate (TPA) and the diacylglycerol (DAG) diC8, which both are argued to stimulate a protein kinase C. The only well-studied scyphozoa, Cassiopea spp., respond differently, i.e. to TPA and diC8 only. We found that larvae of the scyphozoa Aurelia aurita, Chrysaora hysoscella and Cyanea lamarckii respond to all the compounds mentioned. Trigonelline (N-methylnicotinic acid), a metamorphosis inhibitor found in Hydractinia larvae, is assumed to act by delivering a methyl group for transmethylation processes antagonising metamorphosis induction in Chrysaora hysoscella and Cyanea lamarckii. The three species tested are scyphozoa belonging to the subgroup of semaeostomeae, while Cassiopea spp. belong to the rhizostomeae. The results obtained may contribute to the discussion concerning the evolution of cnidarians and may help to clarify whether the way metamorphosis can be induced in rhizostomeae as a whole is different from that in hydrozoa and those scyphozoa belonging to the subgroup semaeostomeae. Electronic Publication     

Development of the rhopalial nervous system in <Emphasis Type="Italic">Aurelia</Emphasis> sp.1 (Cnidaria,Scyphozoa)

We examined the development of the nervous system in the rhopalium, a medusa-specific sensory structure, in Aurelia sp.1 (Cnidaria, Scyphozoa) using confocal microscopy. The rhopalial nervous system appears primarily ectodermal and contains neurons immunoreactive to antibodies against tyrosinated tubulin, taurine, GLWamide, and FMRFamide. The rhopalial nervous system develops in an ordered manner: the presumptive gravity-sensing organ, consisting of the lithocyst and the touch plate, differentiates first; the “marginal center,” which controls swimming activity, second; and finally, the ocelli, the presumptive photoreceptors. At least seven bilaterally arranged neuronal clusters consisting of sensory and ganglion cells and their neuronal processes became evident in the rhopalium during metamorphosis to the medusa stage. Our analysis provides an anatomical framework for future gene expression and experimental studies of development and functions of scyphozoan rhopalia. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.

A new genus of soft coral (Cnidaria,Octocorallia) from the Republic of Congo (Pointe-Noire Region)

A new genus of soft coral from the Republic of Congo is described, Complexum gen. n. Nine West African octocoral species previously described in the genus Alcyonium by Tixier-Durivault (1955) are referred to this new genus, and a new species is described and figured, Complexum pusillum sp. n. The new species is characterized by having encrusting growth form and abundant spiny clubs in the surface of the polyparium. It colonizes shallow calcareous rocky banks (5 to 20 m depth) existing in coastal water of the region of Pointe-Noire. Based on molecular phylogeny this new genus is well separated from Alcyonium species.     

Zoanthids (Cnidaria: Hexacorallia: Zoantharia) from shallow waters of the southern Chilean fjord region, with descriptions of a new genus and two new species

The taxonomy of the order Zoantharia (=Zoanthidea=Zoanthiniaria) is greatly hampered by the paucity of diagnostic morphological features. To facilitate discrimination between similar zoanthids, a combination of morphological and molecular analyses is applied here. The three most abundant zoanthid species in shallow waters of the southern Chilean fjord region are described. Comparison with other zoanthids using molecular markers reveals that two of them are new to science; these are described as Mesozoanthus fossii gen. n., sp. n. and Epizoanthus fiordicus sp. n. Their representatives grow on rocky substratum and do not live in symbiosis with demosponges. In the less abundant M. fossii, animals are greyish in colour and resemble members of Parazoanthus in growth form. Individual polyps can be up to 35 mm long. The more abundant E. fiordicus are also greyish; the polyps arise from thin stolons and reach only 12 mm in length. The third species studied is Parazoanthus elongatus McMurrich, 1904. For these three Chilean zoanthid species, in-situ photographs are presented as well as information on distribution, habitat and associated species. Establishment of the Mesozoanthus gen. n. is of particular importance to taxonomy in the chaotic suborder Macrocnemina.     

p53 regulates the proliferation, differentiation and spontaneous transformation of mesenchymal stem cells

Mesenchymal stem cells (MSC) have been extensively studied and gained wide popularity due to their therapeutic potential. Spontaneous transformation of MSC, from both human and murine origin, has been reported in many studies. MSC transformation depends on the culture conditions, the origin of the cells and the time on culture; however, the precise biological characteristics involved in this process have not been fully defined yet. In this study, we investigated the role of p53 in the biology and transformation of murine bone marrow (BM)-derived MSC. We demonstrate that the MSC derived from p53KO mice showed an augmented proliferation rate, a shorter doubling time and also morphologic and phenotypic changes, as compared to MSC derived from wild-type animals. Furthermore, the MSC devoid of p53 had an increased number of cells able to generate colonies. In addition, not only proliferation but also MSC differentiation is controlled by p53 since its absence modifies the speed of the process. Moreover, genomic instability, changes in the expression of c-myc and anchorage independent growth were also observed in p53KO MSC. In addition, the absence of p53 implicates the spontaneous transformation of MSC in long-term cultures. Our results reveal that p53 plays a central role in the biology of MSC.     

Temporal expression and steroidal regulation of piRNA pathway genes (mael, piwi, vasa) during Silurana (Xenopus) tropicalis embryogenesis and early larval development

It has been extensively documented that exposure of amphibians and teleost fish to exogenous steroid hormones like estrogen, androgen, xenoestrogen or steroid biosynthesis inhibitors can impair their gonadal development or induce sex reversal against genotypic sex. However, the molecular pathways underlying sexual development and the effects of sex steroids or other exogenous hormones in these aquatic vertebrates remain elusive. Recently, a germ plasm-associated piRNA (piwi-interacting RNA) pathway has been shown to be a determinant in the development of animal gonadal germline cells. In the current study, we examined whether this piRNA pathway is involved in the regulation of sex steroid hormones in gonadal development. We firstly established developmental expression patterns of three key piRNA pathway genes (mael, piwi and vasa), during Silurana (Xenopus) tropicalis embryogenesis and early larval development. All three genes exhibit high expression at early developmental stages and have significantly decreased expression thereafter, indicating a very active involvement of piRNA pathway at the beginning of embryogenesis. We further examined gene expression changes of those genes in frog larvae exposed to two sex steroid biosynthesis inhibitors, fadrozole and finasteride, both of which are known to result in male-biased or female-biased phenotypes, respectively. We found that fadrozole and finasteride exposures increased the expression of piRNA pathway genes such as mael and vasa at the larval stage when the expression of piRNA pathway genes is programmed to be very low. Therefore, our results indicate that the piRNA pathway is likely a common pathway by which different sex steroid hormones regulate gonadal sex differentiation.     

Stilbenoids from the stem of Gnetum latifolium (Gnetaceae)

An acetone extract of the stem of Gnetum latifolium Blume afforded the stilbene trimer (latifolol) together with five known stilbenoids (gnetin E, gnetin D, gnetin C, (-)epsilon -viniferin and resveratrol). Their structures were elucidated on the basis of spectral evidence, in particular by using 2D NMR methods.     

Alterations of rx1 and pax6 expression levels at neural plate stages differentially affect the production of retinal cell types and maintenance of retinal stem cell qualities

rx1 and pax6 are necessary for the establishment of the vertebrate eye field and for the maintenance of the retinal stem cells that give rise to multiple retinal cell types. They also are differentially expressed in cellular layers in the retina when cell fates are being specified, and their expression levels differentially affect the production of amacrine cell subtypes. To determine whether rx1 and pax6 expression after the eye field is established simply maintains stem cell-like qualities or affects cell type differentiation, we used hormone-inducible constructs to increase or decrease levels/activity of each protein at two different neural plate stages. Our results indicate that rx1 regulates the size of the retinal stem cell pool because it broadly affected all cell types, whereas pax6 regulates more restricted retinal progenitor cells because it selectively affected different cell types in a time-dependent manner. Analysis of rx1 and pax6 effects on proliferation, and expression of stem cell or differentiation markers demonstrates that rx1 maintains cells in a stem cell state by promoting proliferation and delaying expression of neural identity and differentiation markers. Although pax6 also promotes proliferation, it differentially regulates neural identity and differentiation genes. Thus, these two genes work in parallel to regulate different, but overlapping aspects of retinal cell fate determination.     

Boselaphines (Artiodactyla, Ruminantia, Bovidae) from the Middle Siwaliks of Hasnot, Pakistan

In this paper, boselaphine material from several localities in the area of the Hasnot Pakistan, is described, identified, and discussed. Four species that belong to three different genera of the tribe Boselaphini have been found: Selenoportax vexillarius, S. lydekkeri, Pachyportax latidens and Eotragus sp. Eotragus sp. is reported for the first time from the Hasnot and consequently from other Upper Middle Siwalik sediments of Pakistan and equivalent strata of the world, extending the range of the genus from the Lower to the Middle Siwaliks. Reviewing the Siwaliks’ Selenoportax species, S. dhokpathanensis Akhtar and S. tatrotensis Akhtar are synonymized with S. lydekkeri and S. vexillarius, respectively.     

Reproduction of the colonial hydroid Obelia geniculata (L., 1758) (Cnidaria,Hydrozoa) in the White Sea

Populations of the colonial hydroid Obelia geniculata in the White Sea reproduce asexually by frustule formation. Young medusae appear in the plankton during July and August. The number of medusae rarely exceeds 36 per m³, and the average number varies every year from 0.4 to 10 per m³. The size of medusae is smaller than reported from other regions. The umbrella of the largest recorded medusa was only 0.57 mm in diameter and the specimen had just 35 tentacles. Only a few mature medusae were found during the study. The colonies in the White Sea are epiphytic and grow only on laminarian thalli. At the beginning of July there are no colonies on thalli from the upper subtidal zone. By the end of August, colonies of O.␣geniculata had increased in density to 30 per m². Hydroid recruitment was attributed to active frustule production by colonies living below that zone. The frustules detach from the stems of the hydroids and are found in plankton. Production of frustules on branches occurs continuously during colony growth until water temperatures climb above 0 °C. We found that water temperature in this Arctic environment is generally too low for medusa maturation and planula development in the species. Propagation by frustule formation is the principal means of reproduction in Obelia geniculata within the White Sea, and this phenomenon accounts for the species being a dominant epiphyte on laminarian thalli there.     

New taxa and revisionary systematics of alcyonacean octocorals from the Pacific coast of North America (Cnidaria,Anthozoa)

A taxonomic assessment of four species of octocorals from the northeastern Pacific Ocean (British Columbia to California) is provided. Included here are a new species of clavulariid stolonifieran Cryptophyton, a new species of the nephtheid soft coral Gersemia, an undetermined species of soft coral in the genus Alcyonium that has been referred in the literature by several other names, and a new genus is named for a plexaurid sea fan originally described in the Indo-Pacific genus Euplexaura. Discussions are included that compare the species to related taxa, or provide revisionary assessments.     

Production of (2S,3S)-2,3-butanediol and (3S)-acetoin from glucose using resting cells of Klebsiella pneumonia and Bacillus subtilis

Production of highly pure (2S,3S)-2,3-butanediol ((2S,3S)-2,3-BD) and (3S)-acetoin ((3S)-AC) in high concentrations is desirable but difficult to achieve. In the present study, glucose was first transformed to a mixture of (2S,3S)-2,3-BD and meso-2,3-BD by resting cells of Klebsiella pneumoniae CICC 10011, followed by biocatalytic resolution of the mixture by resting cells of Bacillus subtilis 168. meso-2,3-BD was transformed to (3S)-AC, leaving (2S,3S)-2,3-BD in the reaction medium. Using this approach, 12.5 g l(-1) (2S,3S)-2,3-BD and 56.7 g l(-1) (3S)-AC were produced. Stereoisomeric purity of (2S,3S)-2,3-BD and enantiomeric excess of (3S)-AC was 96.9 and 96.2%, respectively.