Potential of the luminol reaction in the sensitive detection of pesticide residues by flow injection analysis.

This study presents the first analytical application of the luminol chemiluminescence (CL) reaction for the sensitive detection of carbamate residues. Some experiments have been carried out to check the influence of the presence of traces of a N-methylcarbamate (carbaryl) on the CL emission produced from the oxidation of luminol using different oxidants, showing a significant enhancing effect on the CL emission when the oxidation of luminol is produced by potassium permanganate in alkaline medium, this enhancement being proportional to the carbaryl concentration. This fact has permitted the establishment of a sensitive chemiluminescence flow-injection (CL-FIA) method for the direct determination of carbaryl. The optimization of instrumental and chemical variables influencing the CL response has been carried out by applying experimental designs. Under the optimal conditions, the CL intensity was linear for a carbaryl concentration over the range 5-100 ng/mL with a detection limit of 4.9 ng/mL. This luminol-KMnO4-based FIA-CL system in basic medium shows an easy, fast and cheap alternative detection mode for the analysis of carbaryl residues in environmental water samples.     

Selection of DNA aptamers that bind the RNA-dependent RNA polymerase of hepatitis C virus and inhibit viral RNA synthesis in vitro

The RNA-dependent RNA polymerase (NS5B) of the hepatitis C virus (HCV) plays a key role in the life cycle of the virus. In order to find inhibitors of the HCV polymerase, we screened a library of 81 nucleotide (nt)-long synthetic DNA containing 35 random nucleotides by the Systematic Evolution of Ligands by Exponential enrichment (SELEX) approach. Thirty ligands selected for their binding affinity to the NS5B were classified into four groups on the basis of their sequence homologies. Among the selected molecules, two were able to inhibit in vitro the polymerase activity of the HCV NS5B. These aptamers appeared to be specific for HCV polymerase, as no inhibition of poliovirus 3D polymerase activity was observed. The binding and inhibitory potential of one aptamer (27v) was associated with the 35 nt-long variable region. This oligonucleotide displayed an apparent dissociation constant (K(d)) in the nanomolar range. Our results showed that it was able to compete with RNA templates corresponding to the 3'-ends of the (+) and the (-) HCV RNA for binding to the polymerase. The fact that a DNA aptamer could interfere with the binding of natural templates of the enzyme could help in performing structure-function analysis of the NS5B and might constitute a basis for further structure-based drug design of this crucial enzyme of HCV replication.     

Key role for ceramides in mediating insulin resistance in human muscle cells

Elevated non-esterified fatty acids, triglyceride, diacylglycerol, and ceramide have all been associated with insulin resistance in muscle. We set out to investigate the role of intramyocellular lipid metabolites in the induction of insulin resistance in human primary myoblast cultures. Muscle cells were subjected to adenovirus-mediated expression of perilipin or incubated with fatty acids for 18 h, prior to insulin stimulation and measurement of lipid metabolites and rates of glycogen synthesis. Adenovirus-driven perilipin expression lead to significant accumulation of triacylglycerol in myoblasts, without any detectable effect on insulin sensitivity, as judged by the ability of insulin to stimulate glycogen synthesis. Similarly, incubation of cells with the monounsaturated fatty acid oleate resulted in triacylglycerol accumulation without inhibiting insulin action. By contrast, the saturated fatty acid palmitate induced insulin resistance. Palmitate treatment caused less accumulation of triacylglycerol than did oleate but also induced significant accumulation of both diacylglycerol and ceramide. Insulin resistance was also caused by cell-permeable analogues of ceramide, and palmitate-induced resistance was blocked in the presence of inhibitors of de novo ceramide synthesis. Oleate co-incubation completely prevented the insulin resistance induced by palmitate. Our data are consistent with ceramide being the agent responsible for insulin resistance caused by palmitate exposure. Furthermore, the triacylglycerol derived from oleate was able to exert a protective role in sequestering palmitate, thus preventing its conversion to ceramide.     

Incorporating anthropogenic effects into trophic ecology: predator–prey interactions in a human-dominated landscape

Apex predators perform important functions that regulate ecosystems worldwide. However, little is known about how ecosystem regulation by predators is influenced by human activities. In particular, how important are top-down effects of predators relative to direct and indirect human-mediated bottom-up and top-down processes? Combining data on species'' occurrence from camera traps and hunting records, we aimed to quantify the relative effects of top-down and bottom-up processes in shaping predator and prey distributions in a human-dominated landscape in Transylvania, Romania. By global standards this system is diverse, including apex predators (brown bear and wolf), mesopredators (red fox) and large herbivores (roe and red deer). Humans and free-ranging dogs represent additional predators in the system. Using structural equation modelling, we found that apex predators suppress lower trophic levels, especially herbivores. However, direct and indirect top-down effects of humans affected the ecosystem more strongly, influencing species at all trophic levels. Our study highlights the need to explicitly embed humans and their influences within trophic cascade theory. This will greatly expand our understanding of species interactions in human-modified landscapes, which compose the majority of the Earth''s terrestrial surface.     

Establishing Tallgrass Prairie on Grazed Permanent Pasture in the Upper Midwest

The goal of the study was to learn whether native prairie grasses and, eventually, a diverse mixture of native forbs could be incorporated in permanent pastures by means of rotational grazing by cattle. An experiment was established on a farm in northeastern Iowa on a pasture that had never been plowed but had been grazed since the 1880s. One treatment was protected from grazing to test for the presence of remnant vegetation. Andropogon gerardii, Sorghastrum nutans, Panicum virgatum, and Desmanthus illinoensis were introduced in plots first treated with glyphosate; seeds were either drilled (DR) or hand-broadcast and incorporated by controlled cattle trampling (BT). Seedling establishment and aboveground biomass were followed over 3 years. There was no evidence for remnant native plants on uplands, but seven species of native forbs and four native graminoids flowered in exclosures erected within waterways. D. illinoensis initially established up to 12 seedlings/m² but had disappeared from all but one plot by the third year. Variation in native grass establishment among replicate plots within treatments was very high, ranging initially from 0.2 to 9.9 plants/m². In August of the second year, native grasses made up only 8% of the available forage in DR plots and 1% of BT plots. One year later, however, native grasses made up 56% of the available forage in DR plots and 37% of BT plots, and these differences were significant (p = 0.05). A pilot study seeded in late winter (frost seeding) suggested that seeds spread after cattle trampling produced five times more seedlings (2.5/m²) than seeds spread before cattle trampling (0.5/m²). Frost seeding had advantages because it did not require herbicide for sod suppression or tractor access to the site. New plantings could be safely grazed in early spring and late fall, before and after most native grass growth, to offset the negative economic impact of protecting new plantings from burning during the growing season. But this practice precluded subsequent prescribed burning. I propose a strategy for incorporating native wildflowers into the pasture over time with minimum cost.     

Phosphate-solubilizing peanut associated bacteria: screening for plant growth-promoting activities

Carbon and nitrogen are supplied by a variety of sources in the desert food web; both vascular and non-vascular plants and cyanobacteria supply carbon, and cyanobacteria and plant-associated rhizosphere bacteria are sources of biological nitrogen fixation. The objective of this study was to compare the relative influence of vascular plants and biological soil crusts on desert soil nematode and protozoan abundance and community composition. In the first experiment, biological soil crusts were removed by physical trampling. Treatments with crust removed had fewer nematodes and a greater relative ratio of bacterivores to microphytophages than treatments with intact crust. However, protozoa composition was similar with or without the presence of crusts. In a second experiment, nematode community composition was characterized along a spatial gradient away from stems of grasses or shrubs. Although nematodes generally occurred in increasing abundance nearer to plant stems, some genera (such as the enrichment-type Panagrolaimus) increased disproportionately more than others (such as the stress-tolerant Acromoldavicus). We propose that the impact of biological soil crusts and desert plants on soil microfauna, as reflected in the community composition of microbivorous nematodes, is a combination of carbon input, microclimate amelioration, and altered soil hydrology.     

Replication and fine mapping of asthma-associated loci in individuals of African ancestry

Asthma originates from genetic and environmental factors with about half the risk of disease attributable to heritable causes. Genome-wide association studies, mostly in populations of European ancestry, have identified numerous asthma-associated single nucleotide polymorphisms (SNPs). Studies in populations with diverse ancestries allow both for identification of robust associations that replicate across ethnic groups and for improved resolution of associated loci due to different patterns of linkage disequilibrium between ethnic groups. Here we report on an analysis of 745 African-American subjects with asthma and 3,238 African-American control subjects from the Candidate Gene Association Resource (CARe) Consortium, including analysis of SNPs imputed using 1,000 Genomes reference panels and adjustment for local ancestry. We show strong evidence that variation near RAD50/IL13, implicated in studies of European ancestry individuals, replicates in individuals largely of African ancestry. Fine mapping in African ancestry populations also refined the variants of interest for this association. We also provide strong or nominal evidence of replication at loci near ORMDL3/GSDMB, IL1RL1/IL18R1, and 10p14, all previously associated with asthma in European or Japanese populations, but not at the PYHIN1 locus previously reported in studies of African-American samples. These results improve the understanding of asthma genetics and further demonstrate the utility of genetic studies in populations other than those of largely European ancestry.     

Deciduousness in a seasonal tropical forest in western Thailand: interannual and intraspecific variation in timing, duration and environmental cues

Seasonal tropical forests exhibit a great diversity of leaf exchange patterns. Within these forests variation in the timing and intensity of leaf exchange may occur within and among individual trees and species, as well as from year to year. Understanding what generates this diversity of phenological behaviour requires a mechanistic model that incorporates rate-limiting physiological conditions, environmental cues, and their interactions. In this study we examined long-term patterns of leaf flushing for a large proportion of the hundreds of tree species that co-occur in a seasonal tropical forest community in western Thailand. We used the data to examine community-wide variation in deciduousness and tested competing hypotheses regarding the timing and triggers of leaf flushing in seasonal tropical forests. We developed metrics to quantify the nature of deciduousness (its magnitude, timing and duration) and its variability among survey years and across a range of taxonomic levels. Tree species varied widely in the magnitude, duration, and variability of leaf loss within species and across years. The magnitude of deciduousness ranged from complete crown loss to no crown loss. Among species that lost most of their crown, the duration of deciduousness ranged from 2 to 21 weeks. The duration of deciduousness in the majority of species was considerably shorter than in neotropical forests with similar rainfall periodicity. While the timing of leaf flushing varied among species, most (∼70%) flushed during the dry season. Leaf flushing was associated with changes in photoperiod in some species and the timing of rainfall in other species. However, more than a third of species showed no clear association with either photoperiod or rainfall, despite the considerable length and depth of the dataset. Further progress in resolving the underlying internal and external mechanisms controlling leaf exchange will require targeting these species for detailed physiological and microclimatic studies.     

Influence of magnesium ions on biofilm formation by Pseudomonas fluorescens

Mg²⁺ can potentially influence bacterial adhesion directly through effects on electrostatic interactions and indirectly by affecting physiology-dependent attachment processes. However, the effects of Mg²⁺ on biofilm structure are largely unknown. In this study, Pseudomonas fluorescens was used to investigate the influence of Mg²⁺ concentration (0, 0.1 and 1.0 mM MgCl₂) on biofilm growth. Planktonic and attached cells were enumerated (based on DAPI staining) while biofilm structures were examined via confocal laser scanning microscopy and three-dimensional structures were reconstructed. Mg²⁺ concentration had no influence on growth of planktonic cells but, during biofilm formation, Mg²⁺ increased the abundance of attached cells. For attached cells, the influence of Mg²⁺ concentration changed over time, suggesting that the role of Mg²⁺ in bacterial attachment is complex and dynamic. Biofilm structures were heterogeneous and surface colonization and depth increased with increasing Mg²⁺ concentrations. Overall, for P. fluorescens, Mg²⁺ increased initial attachment and altered subsequent biofilm formation and structure.