Spatiotemporal variation in the reproductive ecology of two parapatric subspecies of Oenothera cespitosa (Onagraceae)

?Premise of the study: Flowering plants that rely on pollinators for most of their reproduction may experience unpredictable and inconsistent availability of effective pollinators throughout their reproductive lifetime. We investigated the reproductive ecology of two subspecies of the tufted evening primrose, Oenothera cespitosa, which occupy geographically and edaphically distinct habitats in western North America: O. cespitosa subsp. navajoensis inhabits sandstone soils on open sites or rocky slopes in the Colorado Plateau and O. cespitosa subsp. cespitosa grows in clay soils on talus slopes and exposed rocky ridges in the western Great Plains and northern Rocky Mountains of the United States. ?Methods: Pollen augmentation and selfing experiments, floral visitor observations, and single-visit effectiveness experiments were conducted over 4 years to examine the breeding system and spatiotemporal variation in pollinator behavior, assemblage, and abundance at different populations for each subspecies. ?Key results: Both subspecies of O. cespitosa were self-incompatible and pollen-limited, suggesting that the relative abundance, effectiveness, and movement patterns of different insects as pollinators influenced the quality and quantity of seed production in these plants. Medium-sized vespertine hawkmoths (Hyles lineata, Sphinx vashti) were effective pollinators when present, as were large matinal bees (Anthophora affabilis, A. dammersi, Xylocopa tabaniformis androleuca), whereas small oligolectic Lasioglossum bees primarily functioned as pollen thieves in the evening and morning. ?Conclusions: These findings highlight the importance of variability of pollinator composition and abundance in the evolution of plant breeding systems and reproductive success at varying spatial and temporal scales.     

Frugivory on Persea lingue in temperate Chilean forests: interactions between fruit availability and habitat fragmentation across multiple spatial scales

Habitat degradation and fragmentation are expected to reduce seed dispersal rates by reducing fruit availability as well as the movement and abundance of frugivores. These deleterious impacts may also interact with each other at different spatial scales, leading to nonlinear effects of fruit abundance on seed dispersal. In this study we assessed whether the degradation and fragmentation of southern Chilean forests had the potential to restrict seed dispersal the lingue (Persea lingue) tree, a fleshy-fruited tree species. Of five frugivore bird species, the austral thrush (Turdus falcklandii) and the fire-eyed diucon (Xolmis pyrope) were the only legitimate seed dispersers as well as being the most abundant species visiting lingue trees. The results showed little or no direct effect of habitat fragmentation on seed dispersal estimates, possibly because the assemblage of frugivore birds was comprised habitat-generalist species. Instead, the number of fruits removed per focal tree exhibited an enhanced response to crop size, but only in the more connected fragments. In the fruit-richer fragment networks, there was an increased fragment-size effect on the proportion of fruits removed in comparison to fruit-poor networks in which the fragment size effect was spurious. We suggest that such nonlinear effects are widespread in fragmented forest regions, resulting from the link between the spatial scales over which frugivores sample resources and the spatial heterogeneity in fruiting resources caused by habitat fragmentation and degradation.     

Helicobacter pylori EstV: identification, cloning, and characterization of the first lipase isolated from an epsilon-proteobacterium

Bacterial lipases are attracting an enormous amount of attention due to their wide biotechnological applications and due to their roles as virulence factors in some bacteria. Helicobacter pylori is a significant and widespread pathogen which produces a lipase(s) and phospholipases that seem to play a role in mucus degradation and the release of proinflammatory and cytotoxic compounds. However, no H. pylori lipase(s) has been isolated and described previously. Therefore, a search for putative lipase-encoding genes was performed by comparing the amino acid sequences of 53 known lipolytic enzymes with the deduced proteome of H. pylori. As a result, we isolated, cloned, purified, and characterized EstV, a novel lipolytic enzyme encoded by open reading frame HP0739 of H. pylori 26695, and classified it in family V of the bacterial lipases. This enzyme has the properties of a small, cell-bound carboxylesterase (EC 3.1.1.1) that is active mostly with short-chain substrates and does not exhibit interfacial activation. EstV is stable and does not require additional cofactors, and the maximum activity occurs at 50 degrees C and pH 10. This unique enzyme is the first lipase isolated from H. pylori that has been described, and it might contribute to ulcer development, as inhibition by two antiulcer substances (beta-aescin and glycyrrhizic acid) suggests. EstV is also the first lipase from an epsilon-proteobacterium to be described. Furthermore, this enzyme is a new member of family V, probably the least-known family of bacterial lipases, and the first lipase of this family for which kinetic behavior, inhibition by natural substances, and other key biochemical features are reported.     

Histamine improves antigen uptake and cross-presentation by dendritic cells

Previous studies have shown that histamine is able to modulate the function of dendritic cells (DCs). Histamine seems to be required for the normal differentiation of DCs. Moreover, it is capable of stimulating the chemotaxis of immature DCs and of promoting the differentiation of T CD4+ cells into a Th2 profile. In this study, we analyzed whether histamine was able to modulate endocytosis and cross-presentation mediated by immature DCs. Our results show that both functions are stimulated by histamine. Endocytosis of soluble HRP and FITC-OVA and cross-presentation of soluble OVA were markedly increased by histamine. Interestingly, stimulation of endocytosis and cross-presentation appeared to be mediated through different histamine receptors. In fact, the enhancement of endocytosis was prevented by the histamine2 receptor (H2R) antagonist cimetidine, whereas the stimulation of cross-presentation was prevented by the H3R/H4R antagonist thioperamide. Of note, contrasting with the observations made with soluble Ags, we found that histamine did not increase either the uptake of OVA-attached to latex beads, or the cross-presentation of OVA immobilized on latex beads. This suggests that the ability of histamine to increase endocytosis and cross-presentation is dependent on the Ag form and/or the mechanisms through which the Ag is internalized by DCs. Our results support that histamine may favor cross-presentation of soluble allergens by DCs enabling the activation of allergen-specific T CD8+ cells, which appears to play an important role in the development of allergic responses in the airway.     

Oligothiophene molecular beacons

Oligomers of thiophene are widely studied compounds for their electronic and optoelectronic properties. Despite their strong fluorescence, their use as markers for biomolecules, especially for oligonucleotides (ONs), is still largely unexplored. Here, we describe the synthesis of a series of ON molecular beacons employing different oligothiophenes as fluorescent probes and discuss their fluorescence emissions in comparative experiments with and without dabcyl as a quencher, in their hairpin and linear conformations, and as duplexes after hybridization with a complementary target.     

The usefulness of the <Emphasis Type="Italic">gfp</Emphasis> reporter gene for monitoring <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated transformation of potato dihaploid and tetraploid genotypes

Potato is one of the main targets for genetic improvement by gene transfer. The aim of the present study was to establish a robust protocol for the genetic transformation of three dihaploid and four economically important cultivars of potato using Agrobacterium tumefaciens carrying the in vivo screenable reporter gene for green fluorescent protein (gfp) and the marker gene for neomycin phosphotransferase (nptII). Stem and leaf explants were used for transformation by Agrobacterium tumefaciens strain LBA4404 carrying the binary vector pHB2892. Kanamycin selection, visual screening of GFP by epifluorescent microscopy, PCR amplification of nptII and gfp genes, as well as RT-PCR and Southern blotting of gfp and Northern blotting of nptII, were used for transgenic plant selection, identification and analysis. Genetic transformation was optimized for the best performing genotypes with a mean number of shoots expressing gfp per explant of 13 and 2 (dihaploid line 178/10 and cv. ‘Baltica’, respectively). The nptII marker and gfp reporter genes permitted selection and excellent visual screening of transgenic tissues and plants. They also revealed the effects of antibiotic selection on organogenesis and transformation frequency, and the identification of escapes and chimeras in all potato genotypes. Silencing of the gfp transgene that may represent site-specific inactivation during cell differentiation, occurred in some transgenic shoots of tetraploid cultivars and in specific chimeric clones of the dihaploid line 178/10. The regeneration of escapes could be attributed to either the protection of non-transformed cells by neighbouring transgenic cells, or the persistence of Agrobacterium cells in plant tissues after co-cultivation.     

Time-resolved methods in biophysics. 4. Broadband pump-probe spectroscopy system with sub-20 fs temporal resolution for the study of energy transfer processes in photosynthesis.

In this paper we discuss how to push the temporal resolution limits of transient absorption spectroscopy in order to detect very fast processes (energy relaxation, energy or charge transfer, vibrational coherence) taking place in molecules of biological relevance. After reviewing the main principles of femtosecond pump-probe spectroscopy, we describe an experimental setup based on two synchronized non-collinear optical parametric amplifiers (NOPAs). Each NOPA can be independently configured to generate ultra-broadband sub-10 fs visible pulses, tunable 10-15 fs visible pulses, tunable 15-40 fs near-infrared pulses (900-1500 nm). This system enables to perform pump-probe experiments over nearly two octaves of spectrum with sub-20 fs temporal resolution. We then present an application example highlighting the capability of this instrument to track excited state dynamics in biomolecules on the sub-100 fs timescale: the study of carotenoid-bacteriochlorophyll energy transfer processes in peripheral light-harvesting complexes (LH2) from purple bacteria. We show that, by comparing excited-state dynamics of the carotenoids in organic solvents and inside the LH2 complexes, it is possible to visualize in the time domain the primary events in photosynthesis.     

A new approach to the induction and recovery of Synechococcus leopoliensis CPD-photolyase for cosmetic applications

Journal of Applied Phycology - Ultraviolet (UV) radiation generates cyclobutane pyrimidine dimer (CPD) photoproducts in the DNA of skin cells, causing photoaging and non-melanoma skin cancer....     

Dynamic Mechanisms of the Bactericidal Action of an Al2O3-TiO2-Ag Granular Material on an Escherichia coli Strain

The bactericidal activity of an Al₂O₃-TiO₂-Ag granular material against an Escherichia coli strain was confirmed by a culture-based method. In particular, 100% of microorganisms were permanently inactivated in 30 to 45 min. The present work aimed to investigate the mechanisms of the bactericidal action of this material and their dynamics on Escherichia coli using different techniques. Observations by transmission electron microscopy (TEM) at different times of disinfection revealed morphological changes in the bacteria as soon as they were put in contact with the material. Notably highlighted were cell membrane damage; cytoplasm detachment; formation of vacuoles, possibly due to DNA condensation, in association with regions exhibiting different levels of electron density; and membrane lysis. PCR and flow cytometry analyses were used to confirm and quantify the observations of cell integrity. The direct exposure of cells to silver, combined with the oxidative stress induced by the reactive oxygen species (ROS) generated, was identified to be responsible for these morphological alterations. From the first 5 min of treatment with the Al₂O₃-TiO₂-Ag material, 98% of E. coli isolates were lysed. From 30 min, cell viability decreased to reach total inactivation, although approximately 1% of permeable E. coli cells and 1% of intact cells (10⁵ genomic units · ml⁻¹) were evidenced. This study demonstrates that the bactericidal effect of the material results from a synergic action of desorbed and supported silver. Supported silver was shown to generate the ROS evidenced.