<Emphasis Type="Italic">Ochrobactrum</Emphasis> sp. MPV1 from a dump of roasted pyrites can be exploited as bacterial catalyst for the biogenesis of selenium and tellurium nanoparticles

Background

Bacteria have developed different mechanisms for the transformation of metalloid oxyanions to non-toxic chemical forms. A number of bacterial isolates so far obtained in axenic culture has shown the ability to bioreduce selenite and tellurite to the elemental state in different conditions along with the formation of nanoparticles—both inside and outside the cells—characterized by a variety of morphological features. This reductive process can be considered of major importance for two reasons: firstly, toxic and soluble (i.e. bioavailable) compounds such as selenite and tellurite are converted to a less toxic chemical forms (i.e. zero valent state); secondly, chalcogen nanoparticles have attracted great interest due to their photoelectric and semiconducting properties. In addition, their exploitation as antimicrobial agents is currently becoming an area of intensive research in medical sciences.

Results

In the present study, the bacterial strain Ochrobactrum sp. MPV1, isolated from a dump of roasted arsenopyrites as residues of a formerly sulfuric acid production near Scarlino (Tuscany, Italy) was analyzed for its capability of efficaciously bioreducing the chalcogen oxyanions selenite (SeO₃ ^2?) and tellurite (TeO₃ ^2?) to their respective elemental forms (Se⁰ and Te⁰) in aerobic conditions, with generation of Se- and Te-nanoparticles (Se- and TeNPs). The isolate could bioconvert 2 mM SeO₃ ^2? and 0.5 mM TeO₃ ^2? to the corresponding Se⁰ and Te⁰ in 48 and 120 h, respectively. The intracellular accumulation of nanomaterials was demonstrated through electron microscopy. Moreover, several analyses were performed to shed light on the mechanisms involved in SeO₃ ^2? and TeO₃ ^2? bioreduction to their elemental states. Results obtained suggested that these oxyanions are bioconverted through two different mechanisms in Ochrobactrum sp. MPV1. Glutathione (GSH) seemed to play a key role in SeO₃ ^2? bioreduction, while TeO₃ ^2? bioconversion could be ascribed to the catalytic activity of intracellular NADH-dependent oxidoreductases. The organic coating surrounding biogenic Se- and TeNPs was also characterized through Fourier-transform infrared spectroscopy. This analysis revealed interesting differences among the NPs produced by Ochrobactrum sp. MPV1 and suggested a possible different role of phospholipids and proteins in both biosynthesis and stabilization of such chalcogen-NPs.

Conclusions

In conclusion, Ochrobactrum sp. MPV1 has demonstrated to be an ideal candidate for the bioconversion of toxic oxyanions such as selenite and tellurite to their respective elemental forms, producing intracellular Se- and TeNPs possibly exploitable in biomedical and industrial applications.

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The application of oligonucleotide probes and microarrays for the identification of freshwater diatoms

Diatoms are major contributors to global carbon fixation and constitute a significant portion of biofilms found in lotic ecosystems. Despite their widespread abundance and the fact that extensive studies have been performed on morphological features of frustules, molecular tools for the identification of diatoms are not commonly available. This study focuses on the development of oligonucleotide probes for the detection of diatom species relevant to water quality assessment. The selected panel of diatoms covers all the species found in water of varying quality from the rivers of central-East Apennine (Italy). Small subunit rRNA-targeted probes were applied to a microarray platform as well as to a new technique termed Primer–Probe, with the aim of obtaining a molecular tool suitable for accurate identification of both single and mixed species diatom populations. The Primer–Probe technique together with dot-blot assays proved to be ideal for the preliminary screening of a large set of DNA oligonucleotides designed by ARB software. It was shown that microarrays, as a promising technology for rapid and simultaneous detection of a wide range of species-specific genetic markers, can be adapted to monitor changes within a diatom community. It is suggested that microarrays will provide a molecular basis for microbial identification to support standard microscopy techniques used by ecologists and environmental scientists for monitoring water quality.     

Unraveling interactions of cell cycle-regulating proteins Sic1 and B-type cyclins in living yeast cells: a FLIM-FRET approach

Sic1, cyclin-dependent kinase inhibitor of budding yeast, is synthesized in anaphase and largely degraded at the S-phase onset to regulate timing of DNA synthesis. Sic1 interacts with phase-specific B-type cyclin (Clb)-kinase (Cdk1) complexes, central regulators in cell cycle control. Its appearance is timed to mediate reduction in kinase activities at appropriate stages. Clbs are unstable proteins with extremely short half-lives. Interactions of Sic1 with Clbs have been detected both in vitro and in vivo by high-throughput genome-wide screenings. Furthermore, we have recently shown that Sic1 regulates waves of Clbs, acting as a timer in their appearance, thus controlling Cdk1-Clbs activation. The molecular mechanism is not yet fully understood but is hypothesized to occur via stoichiometric binding of Sic1 to Cdk1-Clb complexes. Using F?rster resonance energy transfer (FRET) via fluorescence lifetime imaging microscopy (FLIM), we showed association of Sic1 to Clb cyclins in living yeast cells. This finding is consistent with the notion that inhibition of kinase activity can occur over the whole cell cycle progression despite variable Sic1 levels. Specifically, Sic1/Clb3 interaction was observed for the first time, and Sic1/Clb2 and Sic1/Clb5 pairs were confirmed, but no Sic1/Clb4 interaction was found, which suggests that, despite high functional homology between Clbs, only some of them can target Sic1 function in vivo.     

Immunotargeting of catalase to the pulmonary endothelium alleviates oxidative stress and reduces acute lung transplantation injury

Vascular immunotargeting may facilitate the rapid and specific delivery of therapeutic agents to endothelial cells. We investigated whether targeting of an antioxidant enzyme, catalase, to the pulmonary endothelium alleviates oxidative stress in an in vivo model of lung transplantation. Intravenously injected enzymes, conjugated with an antibody to platelet-endothelial cell adhesion molecule-1, accumulate in the pulmonary vasculature and retain their activity during prolonged cold storage and transplantation. Immunotargeting of catalase to donor rats augments the antioxidant capacity of the pulmonary endothelium, reduces oxidative stress, ameliorates ischemia-reperfusion injury, prolongs the acceptable cold ischemia period of lung grafts, and improves the function of transplanted lung grafts. These findings validate the therapeutic potential of vascular immunotargeting as a drug delivery strategy to reduce endothelial injury. Potential applications of this strategy include improving the outcome of clinical lung transplantation and treating a wide variety of endothelial disorders.     

Canopy height variation and environmental heterogeneity in the tropical dry forests of coastal Oaxaca,Mexico

Despite its importance for carbon storage and other ecosystem functions, the variation in vegetation canopy height is not yet well understood. We examined the relationship between this community attribute and environmental heterogeneity in a tropical dry forest of southern Mexico. We sampled vegetation in 15 sites along a 100‐km coastal stretch of Oaxaca State, and measured the heights of all woody plants (excluding lianas). The majority of the ca. 4000 individuals recorded concentrated in the 4–8 m height range. We defined three plant sets to describe overall community canopy height at each site: a set including all plants, a set made up by the tallest plants representing 10 percent of all individuals, and a set comprising the 10 tallest plants. For each site we computed maximum height and the mean and median heights of the three sets. Significant collinearity was observed between the seven resulting height variables, but null distributions constructed through bootstrap revealed their different behaviors as functions of species richness and density of individuals. Through linear modeling and a model selection procedure, we identified 21 models that best described the variation in canopy height variables. These models pointed out to soil (measured as PC1 of a principal component analysis performed on 10 soil variables), water stress, and elevation as the main drivers of canopy height variation in the region. In the event of increasing water stress resulting from global climate change, the studied tropical dry forests could become shorter and thus decrease their carbon storage potential.     

The yeast Ski complex is a hetero-tetramer

The yeast Ski complex assists the exosome in the degradation of mRNA. The Ski complex consists of three components; Ski2, Ski3, and Ski8, believed to be present in a 1:1:1 stoichiometry. Measuring the mass of intact isolated endogenously expressed Ski complexes by native mass spectrometry we unambiguously demonstrate that the Ski complex has a hetero-tetrameric stoichiometry consisting of one copy of Ski2 and Ski3 and two copies of Ski8. To validate the stoichiometry of the Ski complex, we performed tandem mass spectrometry. In these experiments one Ski8 subunit was ejected concomitant with the formation of a Ski2/Ski3/Ski8 fragment, confirming the proposed stoichiometry. To probe the topology of the Ski complex we disrupted the complex and mass analyzed the thus formed subcomplexes, detecting Ski8-Ski8, Ski2-Ski3, Ski8-Ski2, and Ski8-Ski8-Ski2. Combining all data we construct an improved structural model of the Ski complex.     

The MAPK pathway triggers activation of Nek2 during chromosome condensation in mouse spermatocytes

Chromosome condensation during the G2/M progression of mouse pachytene spermatocytes induced by the phosphatase inhibitor okadaic acid (OA) requires the activation of the MAPK Erk1. In many cell systems, p90Rsks are the main effectors of Erk1/2 function. We have identified p90Rsk2 as the isoform that is specifically expressed in mouse spermatocytes and have shown that it is activated during the OA-triggered meiotic G2/M progression. By using the MEK inhibitor U0126, we have demonstrated that activation of p90Rsk2 during meiotic progression requires activation of the MAPK pathway. Immunofluorescence analysis indicates that activated Erks and p90Rsk2 are tightly associated with condensed chromosomes during the G2/M transition in meiotic cells. We also found that active p90Rsk2 was able to phosphorylate histone H3 at Ser10 in vitro, but that the activation of the Erk1/p90Rsk2 pathway was not necessary for phosphorylation of H3 in vivo. Furthermore, phosphorylation of H3 was not sufficient to cause condensation of meiotic chromosomes in mouse spermatocytes. Other proteins known to associate with chromatin may represent effectors of Erk1 and p90Rsk2 during chromosome condensation. Nek2 (NIMA-related kinase 2), which associates with chromosomes, plays an active role in chromatin condensation and is stimulated by treatment of pachytene spermatocytes with okadaic acid. We show that inhibition of the MAPK pathway by preincubation of spermatocytes with U0126 suppresses Nek2 activation, and that incubation of spermatocyte cell extracts with activated p90Rsk2 causes stimulation of Nek2 kinase activity. Furthermore, we show that the Nek2 kinase domain is a substrate for p90Rsk2 phosphorylation in vitro. These data establish a connection between the Erk1/p90Rsk2 pathway, Nek2 activation and chromosome condensation during the G2/M transition of the first meiotic prophase.     

Variability in reproductive traits in Jatropha curcas L. accessions during early developmental stages under warm subtropical conditions

Variability in floral, fruit, and seed characteristics, and oil content of 15 accession of Jatropha curcas during early development were assessed during two flowering periods in south Florida subtropical climate. The two flowering periods had leaf flushing in March. Field evaluation using 18 quantitative traits showed significant variation among accessions. The number of female flowers and female : male flower ratio ranged from 1 to 15 and 1 : 8.8 to 1 : 67.8, respectively. Fruit set by natural pollination was 89 and 66% during the first (1st) and second (2nd) flowering periods, respectively. A higher number of female‐type inflorescences were observed during summer. There were significant differences in seed traits, except for number of seeds per fruit. Accession TREC 31 had the highest individual seed dry weight and 100‐seed weight (0.83 g and 79.7 g, respectively). The oil content varied from 19.30% to 35.62%. Seed dry weight had positive correlation with seed fresh weight, seed length, seed thickness, seed width, and 100‐seed weight, but negative correlation with oil content. Based on the cluster analysis using 15 morphological traits, jatropha accessions were grouped into five main clusters and accessions from different geographic regions grouped together in a cluster. Principal component analyses (PCA) revealed morphological variation. The first three components explained 73.5% of the total variation and seed dry weight, 100‐seed weight, total flowers per inflorescence, male flowers per inflorescence and fruit set can be used to distinguish accessions. The PCA also indicated that flowering traits were more influenced by seed origin while seed traits were affected by flowering spans. Although evaluations were performed in plants during the juvenile phase, accessions TREC 31 and TREC 55 had superior averages for almost all characters evaluated. These results provide a preliminary assessment of the high variability in jatropha accessions evaluated and their potential for use in breeding and genetic improvement programs.     

Effects of simultaneous changes in light, nutrients, and herbivory levels, on the structure and function of a subtropical turtlegrass meadow

The interactive effects of light, nutrients, and simulated herbivory on the structure and functioning of a subtropical turtlegrass bed were analyzed monthly from May to October 2001 in Perdido Bay, FL. For each of the three factors, two levels were evaluated in a factorial design with four replicates per treatment. The variables included: light, at ambient and 40% reduction; nutrients, at ambient and 2× ambient concentrations; and herbivory, with no herbivory and simulated effects of a density of 15 sea urchins/m². In practice, light levels turned out to be 40% of surface PAR for ambient conditions, and 16% for shaded plots. Biomass removed as herbivory represented, on average, slightly less than 20% of the above-ground biomass. Separate three-way ANOVAs found no significant three-way interactions for any of the response variables, and few two-way interactions. There were no significant nutrient effects on turtlegrass above-ground biomass, although nutrient additions produced significant decreases in epibiont biomass, and net above-ground primary production (NAPP); significant increases in below-ground biomass during the peak of the growing season. Shoot density and average number of leaves per shoot increased significantly, while the C/N ratio of the oldest leaf in the enriched plots decreased significantly. Light reduction significantly negatively affected all response variables, except below-ground biomass, shoot density and leaf length. Herbivory had isolated and inconsistent significant effects on below-ground biomass, shoot density, average number of leaves per shoot, and leaf length and width. Overall, our results indicate that nutrients are not limiting in Perdido Bay, and that nutrient additions had mostly detrimental effects. Light appeared to be the most important variable limiting seagrasses growth and abundance, and as with terrestrial plants, seagrasses seemed to respond more to light and nutrients than to herbivory. However, it is essential that additional tests of the single and interactive effects of the three key factors of light, nutrients and herbivory be done to evaluate the generality of our work, since our study is the first of its kind in seagrass meadows.