Aglaophenia octodonta (Cnidaria, Hydrozoa) and the Associated Microbial Community: a Cooperative Alliance?

Recently, genetic approaches have revealed a surprising bacterial world as well as a growing knowledge of the enormous distribution of animal-bacterial interactions. In the present study, the diversity of the microorganisms associated to the hydroid Aglaophenia octodonta was studied with epifluorescence, optical, and scanning electron microscopy. Small subunit ribosomal RNA gene sequencing with "universal" and taxon-specific primers allowed the assignment of the microalgae to Symbiodinium and the peritrich ciliates to Pseudovorticella, while the luminous vibrios were identified as Vibrio jasicida of the Harvey clade. To understand the possible relationships among Vibrio jasicida, Symbiodinium, A. octodonta, and Pseudovorticella, specific treatments were conducted in microcosm experiments, with the antibiotic ampicillin and other substances that interfere with bacterial and hydroid metabolism. Treatment of A. octodonta with ampicillin resulted in a decrease of bacterial luminescence followed by Pseudovorticella detachment and Symbiodinium expulsion and suggesting that these microorganisms form a "consortium" with beneficial metabolic interdependence. This hypothesis was reinforced by the evidence that low concentrations of hydrogen peroxide, which stimulate the bacterial oxidative metabolism and luminescence by releasing oxygen, were able to counteract the detrimental effect of ampicillin on the stability of the studied A. octodonta association. A model is proposed in which microalgae that release oxygen during photosynthesis are useful to luminous bacteria for their metabolism and for establishing/maintaining symbiosis leading to a close alliance and mutual benefit of the system A. octodonta-Vibrio jasicida-Pseudovorticella sp.-Symbiodinium sp.

Shared Epizoic Taxa and Differences in Diatom Community Structure Between Green Turtles (Chelonia mydas) from Distant Habitats.

The first reports of diatoms growing on marine mammals date back to the early 1900s. However, only recently has direct evidence been provided for similar associations between diatoms and sea turtles. We present a comparison of diatom communities inhabiting carapaces of green turtles Chelonia mydas sampled at two remote sites located within the Indian (Iran) and Atlantic (Costa Rica) Ocean basins. Diatom observations and counts were carried out using scanning electron microscopy. Techniques involving critical point drying enabled observations of diatoms and other microepibionts still attached to sea turtle carapace and revealed specific aspects of the epizoic community structure. Species-poor, well-developed diatom communities were found on all examined sea turtles. Significant differences between the two host sea turtle populations were observed in terms of diatom abundance and their community structure (including growth form structure). A total of 12 and 22 diatom taxa were found from sea turtles in Iran and Costa Rica, respectively, and eight of these species belonging to Amphora, Chelonicola, Cocconeis, Navicula, Nitzschia and Poulinea genera were observed in samples from both locations. Potential mechanisms of diatom dispersal and the influence of the external environment, sea turtle behaviour, its life stage, and foraging and breeding habitats, as well as epibiotic bacterial flora on epizoic communities, are discussed.

Serogroup-specific interaction of Neisseria meningitidis capsular polysaccharide with host cell microtubules and effects on tubulin polymerization.

We have previously shown that during late stages of the infectious process, serogroup B meningococci (MenB) are able to escape the phagosome of in vitro-infected human epithelial cells. They then multiply in the cytosolic environment and spread intracellularly and to surrounding cells by exploiting the microtubule cytoskeleton, as suggested by results of infections in the presence of microtubule inhibitors and evidence of nanotubes connecting neighboring cells. In this study, by using microtubule binding assays with purified microtubule asters and bundles and microtubule bundles synthesized in vitro, we demonstrate that the MenB capsule directly mediates the interaction between bacteria and microtubules. The direct interaction between the microtubules and the MenB capsular polysaccharide was confirmed by coimmunoprecipitation experiments. Unexpectedly, serogroup C meningococci (MenC), which have a capsular polysaccharide that differs from that of MenB only by its anomeric linkage, α(2→9) instead of α(2→8), were not able to interact with the microtubules, and the lack of interaction was not due to capsular polysaccharide O-acetylation that takes place in most MenC strains but not in MenB strains. Moreover, we demonstrate that the MenB capsular polysaccharide inhibits tubulin polymerization in vitro. Thus, at variance with MenC, MenB may interfere with microtubule dynamics during cell infection.

Multiple-pathways light modulation in Pleurosigma strigosum bi-raphid diatom.

Ordered, quasi-ordered, and even disordered nanostructures can be identified as constituent components of several protists, plants and animals, making possible an efficient manipulation of light for intra- and inter- species communication, camouflage, or for the enhancement of primary production. Diatoms are ubiquitous unicellular microalgae inhabiting all the aquatic environments on Earth. They developed, through tens of millions of years of evolution, ultrastructured silica cell walls, the frustules, able to handle optical radiation through multiple diffractive, refractive, and wave-guiding processes, possibly at the basis of their high photosynthetic efficiency. In this study, we employed a range of imaging, spectroscopic and numerical techniques (including transmission imaging, digital holography, photoluminescence spectroscopy, and numerical simulations based on wide-angle beam propagation method) to identify and describe different mechanisms by which Pleurosigma strigosum frustules can modulate optical radiation of different spectral content. Finally, we correlated the optical response of the frustule to the interaction with light in living, individual cells within their aquatic environment following various irradiation treatments. The obtained results demonstrate the favorable transmission of photosynthetic active radiation inside the cell compared to potentially detrimental ultraviolet radiation.

Hexagonal Voronoi pattern detected in the microstructural design of the echinoid skeleton.

Repeated polygonal patterns are pervasive in natural forms and structures. These patterns provide inherent structural stability while optimizing strength-per-weight and minimizing construction costs. In echinoids (sea urchins), a visible regularity can be found in the endoskeleton, consisting of a lightweight and resistant micro-trabecular meshwork (stereom). This foam-like structure follows an intrinsic geometrical pattern that has never been investigated. This study aims to analyse and describe it by focusing on the boss of tubercles-spine attachment sites subject to strong mechanical stresses-in the common sea urchin Paracentrotus lividus. The boss microstructure was identified as a Voronoi construction characterized by 82% concordance to the computed Voronoi models, a prevalence of hexagonal polygons, and a regularly organized seed distribution. This pattern is interpreted as an evolutionary solution for the construction of the echinoid skeleton using a lightweight microstructural design that optimizes the trabecular arrangement, maximizes the structural strength and minimizes the metabolic costs of secreting calcitic stereom. Hence, this identification is particularly valuable to improve the understanding of the mechanical function of the stereom as well as to effectively model and reconstruct similar structures in view of future applications in biomimetic technologies and designs.

Bioremoval of Yttrium (III), Cerium (III), Europium (III), and Terbium (III) from Single and Quaternary Aqueous Solutions Using the Extremophile Galdieria sulphuraria (Galdieriaceae, Rhodophyta).

The lanthanides are among the rare earth elements (REEs), which are indispensable constituents of modern technologies and are often challenging to acquire from natural resources. The demand for REEs is so high that there is a clear need to develop efficient and environmentally-friendly recycling methods. In the present study, living cells of the extremophile Galdieria sulphuraria were used to remove four REEs, Yttrium, Cerium, Europium, and Terbium, from single- and quaternary-metal aqueous solutions. Two different strains, SAG 107.79 and ACUF 427, were exposed to solutions buffered at pH 2.5, 3.5, 4.5, and 5.5. Our data demonstrated that the removal performances were strain and pH dependent for all metal ions. At lower pH, ACUF 427 outperformed SAG 107.79 considerably. By increasing the pH of the solutions, there was a significant surge in the aqueous removal performance of both strains. The same trend was highlighted using quaternary-metal solutions, even if the quantities of metal removed were significantly lower. The present study provided the first insight into the comparative removal capacity of the Galdieria sulphuraria strains. The choice of the appropriate operational conditions such as the pH of the metal solutions is an essential step in developing efficient, rapid, and straightforward biological methods for recycling REEs.

Cyanidiophyceae (Rhodophyta) Tolerance to Precious Metals: Metabolic Response to Palladium and Gold.

Polyextremophilic red algae, which belong to the class Cyanidiophyceae, are adapted to live in geothermal and volcanic sites. These sites often have very high concentrations of heavy and precious metals. In this study, we assessed the capacity of three strains of Galdieria (G. maxima, G. sulphuraria, and G. phlegrea) and one strain of Cyanidiumcaldarium to tolerate different concentrations of precious metals, such as palladium (Cl4K2Pd) and gold (AuCl4K) by monitoring algal growths in cultures exposed to metals, and we investigated the algae potential oxidative stress induced by the metals. This work provides further understanding of metals responses in the Cyanidiophyceae, as this taxonomic class is developed as a biological refinement tool.

Underwater Light Manipulation by the Benthic Diatom Ctenophora pulchella: From PAR Efficient Collection to UVR Screening.

Several species of diatoms, unicellular microalgae which constitute the main component of phytoplankton, are characterized by an impressive photosynthetic efficiency while presenting a noticeable tolerance versus exposure to detrimental UV radiation (UVR). In particular, the growth rate of the araphid diatom Ctenophora pulchella is not significantly affected by harsh treatments with UVR, even in absence of detectable, specific UV-absorbing pigments and even if it is not able to avoid high UV exposure by motility. In this work we applied a multi-disciplinary approach involving numerical computation, photonics, and biological parameters in order to investigate the possible role of the frustule, micro- and nano-patterned silica shell which encloses the cell, in the ability of C. pulchella to efficiently collect photosynthetic active radiation (PAR) and to simultaneously screen the protoplasm from UVR. The characterization of the photonic properties of the frustule has been accompanied by in vivo experiments conducted in water in order to investigate its function as optical coupler between light and plastids.

The HrpB-HrpA two-partner secretion system is essential for intracellular survival of Neisseria meningitidis.

In this study we used HeLa cells to investigate the role of the HrpB-HrpA two-partner secretion (TPS) system in the meningococcal infection cycle. Although there is evidence that several pathogenic microorganisms may use TPS systems to colonize epithelial surfaces, the meningococcal HrpB-HrpA TPS system was not primarily involved in adhesion to or invasion of HeLa cells. Instead, this system was essential for intracellular survival and escape from infected cells. Gentamicin protection assays, immunofluorescence and transmission electron microscopy analyses demonstrated that, in contrast to the wild-type strain, HrpB-HrpA-deficient mutants were primarily confined to late endocytic vacuoles and trapped in HeLa cells. Haemolytic tests using human erythrocytes suggested that the secreted HrpA proteins could act as manganese-dependent lysins directly involved in mediating vacuole escape. In addition, we demonstrated that escape of wild-type meningococci from infected cells required the use of an intact tubulin cytoskeleton and that the hrpB-hrpA genes, which are absent in other Neisseria spp., were upregulated during infection.

Effects of tellurite on growth of Saccharomyces cerevisiae.

The effects of potassium tellurite on growth and survival of rho(+) and rho(0) Saccharomyces cerevisiae strains were investigated. Both rho(+) and rho(0) strains grew on a fermentable carbon source with up to 1.2 mM K(2)TeO(3), while rho(+) yeast cells grown on a non-fermentable carbon source were inhibited at tellurite levels as low as 50 muM suggesting that this metalloid specifically inhibited mitochondrial functions. Growth of rho(+) yeast cells in the presence of increasing amount of tellurite resulted in dose-dependent blackening of the culture, a phenomenon not observed with rho(0) cultures. Transmission electron microscopy of S. cerevisiae rho(+) cells grown in the presence of tellurite showed that blackening was likely due to elemental tellurium (Te(0)) that formed large deposits along the cell wall and small precipitates in both the cytoplasm and mitochondria.

The microbial community of Vetiver root and its involvement into essential oil biogenesis.

Vetiver is the only grass cultivated worldwide for the root essential oil, which is a mixture of sesquiterpene alcohols and hydrocarbons, used extensively in perfumery and cosmetics. Light and transmission electron microscopy demonstrated the presence of bacteria in the cortical parenchymatous essential oil-producing cells and in the lysigen lacunae in close association with the essential oil. This finding and the evidence that axenic Vetiver produces in vitro only trace amounts of oil with a strikingly different composition compared with the oils from in vivo Vetiver plants stimulated the hypothesis of an involvement of these bacteria in the oil metabolism. We used culture-based and culture-independent approaches to analyse the microbial community of the Vetiver root. Results demonstrate a broad phylogenetic spectrum of bacteria, including alpha-, beta- and gamma-Proteobacteria, high-G+C-content Gram-positive bacteria, and microbes belonging to the Fibrobacteres/Acidobacteria group. We isolated root-associated bacteria and showed that most of them are able to grow by using oil sesquiterpenes as a carbon source and to metabolize them releasing into the medium a large number of compounds typically found in commercial Vetiver oils. Several bacteria were also able to induce gene expression of a Vetiver sesquiterpene synthase. These results support the intriguing hypothesis that bacteria may have a role in essential oil biosynthesis opening the possibility to use them to manoeuvre the Vetiver oil molecular structure.

Reverse transcriptase-PCR differential display analysis of meningococcal transcripts during infection of human cells: up-regulation of priA and its role in intracellular replication.

BACKGROUND: In vitro studies with cell line infection models are beginning to disclose the strategies that Neisseria meningitidis uses to survive and multiply inside the environment of the infected host cell. The goal of this study was to identify novel virulence determinants that are involved in this process using an in vitro infection system. RESULTS: By using reverse transcriptase-PCR differential display we have identified a set of meningococcal genes significantly up-regulated during residence of the bacteria in infected HeLa cells including genes involved in L-glutamate transport (gltT operon), citrate metabolism (gltA), disulfide bond formation (dsbC), two-partner secretion (hrpA-hrpB), capsulation (lipA), and DNA replication/repair (priA). The role of PriA, a protein that in Escherichia coli plays a central role in replication restart of collapsed or arrested DNA replication forks, has been investigated. priA inactivation resulted in a number of growth phenotypes that were fully complemented by supplying a functional copy of priA. The priA-defective mutant exhibited reduced viability during late logarithmic growth phase. This defect was more severe when it was incubated under oxygen-limiting conditions using nitrite as terminal electron acceptors in anaerobic respiration. When compared to wild type it was more sensitive to hydrogen peroxide and the nitric oxide generator sodium nitroprusside. The priA-defective strain was not affected in its ability to invade HeLa cells, but, noticeably, exhibited severely impaired intracellular replication and, at variance with wild type and complemented strains, it co-localized with lysosomal associated membrane protein 1. CONCLUSION: In conclusion, our study i.) demonstrates the efficacy of the experimental strategy that we describe for discovering novel virulence determinants of N. meningitidis and ii.) provides evidence for a role of priA in preventing both oxidative and nitrosative injury, and in intracellular meningococcal replication.

UV-shielding and wavelength conversion by centric diatom nanopatterned frustules.

Diatoms can represent the major component of phytoplankton and contribute massively to global primary production in the oceans. Over tens of millions of years they developed an intricate porous silica shell, the frustule, which ensures mechanical protection, sorting of nutrients from harmful agents, and optimization of light harvesting. Several groups of microalgae evolved different strategies of protection towards ultraviolet radiation (UVR), which is harmful for all living organisms mainly through the formation of dimeric photoproducts between adjacent pyrimidines in DNA. Even in presence of low concentrations of UV-absorbing compounds, several diatoms exhibit significant UVR tolerance. We here investigated the mechanisms involved in UVR screening by diatom silica investments focusing on single frustules of a planktonic centric diatom, Coscinodiscus wailesii, analyzing absorption by the silica matrix, diffraction by frustule ultrastructure and also UV conversion into photosynthetically active radiation exerted by nanostructured silica photoluminescence. We identified the defects and organic residuals incorporated in frustule silica matrix which mainly contribute to absorption; simulated and measured the spatial distribution of UVR transmitted by a single valve, finding that it is confined far away from the diatom valve itself; furthermore, we showed how UV-to-blue radiation conversion (which is particularly significant for photosynthetic productivity) is more efficient than other emission transitions in the visible spectral range.

Mitochondrial ribosomal proteins involved in tellurite resistance in yeast Saccharomyces cerevisiae.

A considerable body of evidence links together mitochondrial dysfunctions, toxic action of metalloid oxyanions, and system and neurodegenerative disorders. In this study we have used the model yeast Saccharomyces cerevisiae to investigate the genetic determinants associated with tellurite resistance/sensitivity. Nitrosoguanidine-induced K2TeO3-resistant mutants were isolated, and one of these mutants, named Sc57-Te5R, was characterized. Both random spore analysis and tetrad analysis and growth of heterozygous (TeS/Te5R) diploid from Sc57-Te5R mutant revealed that nuclear and recessive mutation(s) was responsible for the resistance. To get insight into the mechanisms responsible for K2TeO3-resistance, RNA microarray analyses were performed with K2TeO3-treated and untreated Sc57-Te5R cells. A total of 372 differentially expressed loci were identified corresponding to 6.37% of the S. cerevisiae transcriptome. Of these, 288 transcripts were up-regulated upon K2TeO3 treatment. About half of up-regulated transcripts were associated with the following molecular functions: oxidoreductase activity, structural constituent of cell wall, transporter activity. Comparative whole-genome sequencing allowed us to identify nucleotide variants distinguishing Sc57-Te5R from parental strain Sc57. We detected 15 CDS-inactivating mutations, and found that 3 of them affected genes coding mitochondrial ribosomal proteins (MRPL44 and NAM9) and mitochondrial ribosomal biogenesis (GEP3) pointing out to alteration of mitochondrial ribosome as main determinant of tellurite resistance.

Folding mechanisms steer the amyloid fibril formation propensity of highly homologous proteins.

Significant advances in the understanding of the molecular determinants of fibrillogenesis can be expected from comparative studies of the aggregation propensities of proteins with highly homologous structures but different folding pathways. Here, we fully characterize, by means of stopped-flow, T-jump, CD and DSC experiments, the unfolding mechanisms of three highly homologous proteins, zinc binding Ros87 and Ml153-149 and zinc-lacking Ml452-151. The results indicate that the three proteins significantly differ in terms of stability and (un)folding mechanisms. Particularly, Ros87 and Ml153-149 appear to be much more stable to guanidine denaturation and are characterized by folding mechanisms including the presence of an intermediate. On the other hand, metal lacking Ml452-151 folds according to a classic two-state model. Successively, we have monitored the capabilities of Ros87, Ml452-151 and Ml153-149 to form amyloid fibrils under native conditions. Particularly, we show, by CD, fluorescence, DLS, TEM and SEM experiments, that after 168 hours, amyloid formation of Ros87 has started, while Ml153-149 has formed only amorphous aggregates and Ml452-151 is still monomeric in solution. This study shows how metal binding can influence protein folding pathways and thereby control conformational accessibility to aggregation-prone states, which in turn changes aggregation kinetics, shedding light on the role of metal ions in the development of protein deposition diseases.

Lensless light focusing with the centric marine diatom Coscinodiscus walesii.

In this work, we report on the light focusing ability exploited by the microshell of a marine organism: the Coscinodiscus wailesii diatom. A 100 microm spot size of a red laser beam is narrowed up to less than 10 microm at a distance of 104 microm after the transmission through the regular geometry of the diatom structure, which thus acts as a microlens. Numerical simulations of the electromagnetic field propagation show a good qualitative agreement with the experimental results. The focusing effect is due to the superposition of the waves scattered by the holes present on the surface of the diatom valve. Very interesting applications in micro-optic devices are feasible due to the morphological and biological characteristic of these unicellular organisms.

Diatom communities on commercial biocidal fouling control coatings after one year of immersion in the marine environment.

Little is known about the effect of commercial biocidal fouling control coatings on fouling diatom communities and their growth forms after long periods of exposure in the marine tropical environment. The current study investigated the abundance and composition of fouling diatom communities developed on 11 commercially available biocidal antifouling coatings, covering the three main technology types in recent historic use (Self-Polishing Copolymers, Self-Polishing Hybrid and Controlled Depletion Polymers) after one year of static immersion at two locations in Muscat, Oman (Marina Shangri La and Marina Bandar Rowdha). Light microscopy demonstrated that the total abundance of diatoms and the relative abundance of growth forms were significantly affected by the choice of biocidal antifouling coating and experimental location. Using scanning electron microscopy, a total of 21 diatom genera were identified which were grouped into adnate, motile, plocon and erect growth forms. The adnate growth forms, mainly the genera Amphora, Cocconeis and Mastogloia, dominated the other growth forms in terms of their relative abundance. Current results revealed the importance of exposure location and choice of biocidal antifouling coating on the relative abundance of diatom growth forms.

Summer Epiphytic Diatoms from Terra Nova Bay and Cape Evans (Ross Sea, Antarctica)--A Synthesis and Final Conclusions.

Despite recent advances in polar marine biology and related fields, many aspects of the ecological interactions that are crucial for the functioning of Antarctic shallow water habitats remain poorly understood. Although epiphytic diatoms play an essential role in the Antarctic marine food web, basic information regarding their ecology, biodiversity and biogeography is largely unavailable. Here, we synthesise studies on Ross Sea epiphytic diatoms collected during 11 summer Antarctic expeditions between the years 1989/90 and 2011/12, presenting a full list of diatom taxa associated with three macroalgal species (Iridaea cordata, Phyllophora antarctica, and Plocamium cartilagineum) and their epiphytic sessile fauna. Diatom communities found during the three summer months at various depths and sampling stations differed significantly in terms of species composition, growth form structure and abundances. Densities ranged from 21 to >8000 cells mm-2, and were significantly higher on the surface of epiphytic micro-fauna than on any of the macroalgal species examined. Generally, host organisms characterized by higher morphological heterogeneity (sessile microfauna, ramified Plocamium) supported richer diatom communities than those with more uniform surfaces (Iridaea). Differences between epiphytic communities associated with different macroalgae were reflected better in species composition than in growth form structure. The latter changed significantly with season, which was related strongly to the changing ice conditions. A general trend towards an increasing number of erect forms in deeper waters and tube-dwelling diatoms in the shallowest sites (2-5 m) was also observed. This study explores further important and largely previously unknown aspects of relationships and interactions between Antarctic epiphytic diatoms and their micro- and macro-environments.

Chlamydomonas pitschmannii Ettl, a little known species from thermoacidic environments.

Three Chlamydomonas strains were isolated from the soils of a hot spring located in the Campi Flegrei Caldera (Naples, Italy). Ecophysiological, morpho-cytological and molecular features were used to characterize these isolates and to compare them with chlamydomonax acidophila strains from algal culture collections. The strains were collected from three points of the volcanic site, differing in their physico-chemical conditions. Among the examined Chlamydomonas strains, only the isolates from Campi Flegrei could grow optimally at pH values < or =3.0. These isolates also showed a high tolerance to desiccation and high temperatures, not evidenced by the other Chlamydomonas strains included in the study. 18S rDNA phylogeny indicates that the isolates from Campi Flegrei are closely related to Chlamydomonas pitschmannii and two strains isolated in Canada and Europe, that have been designated as Chlamydomonas acidophila. A Chlamydomonas acidophila strain isolated from the type locality in Japan is less closely related according to its molecular phylogeny, and can also be discerned by light and electron microscopy. Moreover, vegetative cells and sporangia of Chlamydomonas acidophila from Japan showed a median trilaminar structure not observed in the other strains. Our results show that Chlamydomonas pitschmannii could represent a hitherto unknown extremophilic Chlamydomonas species.

Nanostructures in diatom frustules: functional morphology of valvocopulae in Cocconeidacean monoraphid taxa.

Nanopositioning is one of the most intriguing challenges in nanoelectronics and photonics. As the optoelectronic circuit components become more and more complex, the importance of nanopositioning becomes critical. For example, in single-mode fiber telecommunications it is easy to show that an alignment deviation as small as a few tens of nanometers can yield to 50 percent of loss performance. Diatoms are unicellular plants occurring in almost every aquatic environment. Their main morphological character is the frustule, a silica cell wall that consists of two valves, encasing the protoplasm, joined together by a girdle. The girdle is composed of a series of silica bands (copulae) linked together along their margins. In several diatom species, the first girdle bands (valvocopulae), which associate the valves with the rest of the girdle, appear different in shape and bring specific nanostructures devoted to facilitate this linkage. The species of the diatom family Cocconeidaceae show elaborate linkage systems between the valves in which functionally complex valvocopulae are involved. Different levels of complexity and functional efficiency are recognizable in the valve-valvocopula and valvocopula-valvocopula linkages of different cocconeidacean taxa. This work briefly reviews the morphological characteristics and the functional role of the nanostructures involved in the linkages; in fact, we firmly believe that they are stimulating models for design and manufacturing engineers working in the technological nanoworld.