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The reliable analysis of the cell cycle status has become increasingly relevant for scientific and clinical work, especially for the determination of tumor cell growth. One established method to characterize the proliferation activity of cells is the analysis of the Ki-67 protein. Ki-67 is expressed in the nucleus during the whole cell cycle except for the G0 phase. Several different protocols exist for the examination of the Ki-67 protein in tissue and cell culture, but most of them are defined for human cells. For the analysis of the Ki-67 protein in murine tissue and cell culture there is a variety of protocols existing which recommend different fixation and permeabilization reagents or special kits. In this study, we established a reliable protocol for Ki-67 staining in murine cells and tissue based on PFA fixation, which can be used not only for flow cytometry but also for immunofluorescence microscopy analysis. We tested our protocol successfully with three different Ki-67 anti-mouse antibodies in cell culture, regenerating liver tissue and mouse melanoma tumor to demonstrate the general applicability.

Sepsis is associated with a strong inflammatory reaction triggering a complex and prolonged immune response. Septic patients have been shown to develop sustained immunosuppression due to a reduced responsiveness of leukocytes to pathogens. Changes in cellular metabolism of leukocytes have been linked to this phenomenon and contribute to the ongoing immunological derangement. However, the underlying mechanisms of these phenomena are incompletely understood. In cell culture models, we mimicked LPS tolerance conditions to provide evidence that epigenetic modifications account for monocyte metabolic changes which cause immune paralysis in restimulated septic monocytes. In detail, we observed differential methylation of CpG sites related to metabolic activity in human PBMCs 18 h after septic challenge. The examination of changes in immune function and metabolic pathways was performed in LPS-tolerized monocytic THP-1 cells. Passaged THP-1 cells, inheriting initial LPS challenge, presented with dysregulation of cytokine expression and oxygen consumption for up to 7 days after the initial LPS treatment. Proinflammatory cytokine concentrations of TNFα and IL1β were significantly suppressed following a second LPS challenge (p<0.001) on day 7 after first LPS stimulation. However, the analysis of cellular metabolism did not reveal any noteworthy alterations between tolerant and nontolerant THP-1 monocytes. No quantitative differences in ATP and NADH synthesis or participating enzymes of energy metabolism occurred. Our data demonstrate that the function and epigenetic modifications of septic and tolerized monocytes can be examined in vitro with the help of our LPS model. Changes in CpG site methylation and monocyte function point to a correlation between epigenetic modification in metabolic pathways and reduced monocyte function under postseptic conditions.

The aim of this study was to compare two variants of a novel polycarbonate polyurethane prosthesis with polyethylene terephthalate (Dacron) prosthesis in an established sheep model of carotid patch angioplasty. Two variants of the polycarbonate polyurethane prosthesis were used: (1) Polycarbonate polyurethane equal (PCU-e) prosthesis consisted of a multilayered porous structure with equal internal and external layers; (2) Polycarbonate polyurethane with more porous inner layers (PCU-mp) prosthesis had more porous inner layers than external layers. Carotid patch angioplasty was performed in 12 sheep: in six sheep, the PCU-e variant and in the other six sheep, the PCU-mp variant was implanted. Dacron patches were implanted in the contralateral carotid artery of all sheep as a control. Half of the animals with each polycarbonate polyurethane variant were euthanized after two weeks and the other half after eight weeks. Cellular infiltration, endothelialization, and neointimal hyperplasia were examined. All grafts were patent, and no thrombus was seen in any of the harvested arteries. The pores of all the three patch materials allowed infiltration of inflammatory cells, capillaries, and connective tissue. After eight weeks, a nearly complete endothelialization was visible in all patch groups without an obvious difference between the three patch materials. The neointima was thinner in the PCU patches (PCU-e: 56 +/- 13 mu m, PCU-mp: 119 +/- 60 mu m) when compared to Dacron patches (156 +/- 64 mu m) after two weeks. After 8 weeks, a further neointimal growth was detectable, without any significant difference of neointimal thickness between the three patch materials (Dacron: 274 +/- 82 mu m, PCU-e: 324 +/- 98 mu m, PCU-mp: 235 +/- 59 mu m). With the novel polycarbonate polyurethane patch materials, we achieved promising functional and morphological results with 100% patency and nearly complete endothelialization. Our findings showed at least a non-inferiority of the novel polycarbonate polyurethane patch material compared to Dacron. There were no significant differences detected between the two polycarbonate polyurethane patch variants.

Summary The cross talk between adipose tissue and the heart has an increasing importance for cardiac function under physiological and pathological conditions. This study characterizes the role of fat body lipolysis for cardiac function in Drosophila melanogaster. Perturbation of the function of the key lipolytic enzyme, brummer (bmm), an ortholog of the mammalian ATGL (adipose triglyceride lipase) exclusively in the fly's fat body, protected the heart against starvation-induced dysfunction. We further provide evidence that this protection is caused by the preservation of glycerolipid stores, resulting in a starvation-resistant maintenance of energy supply and adequate cardiac ATP synthesis. Finally, we suggest that alterations of lipolysis are tightly coupled to lipogenic processes, participating in the preservation of lipid energy substrates during starvation. Thus, we identified the inhibition of adipose tissue lipolysis and subsequent energy preservation as a protective mechanism against cardiac dysfunction during catabolic stress., Graphical abstract, Highlights • A cross talk between fat body and the heart regulates cardiac function in Drosophila • Knockdown of fat-body brummer lipase prevents starvation-induced cardiac dysfunction • This involves preservation of lipid stores and maintenance of cardiac energy supply • Brummer-mediated preservation of fat body lipid stores involves lipolysis and lipogenesis, Molecular Physiology; Lipidomics; Metabolomics

Summary ‘Photoarsenotrophy’, the use of arsenite as an electron donor for anoxygenic photosynthesis, is thought to be an ancient form of phototrophy along with the photosynthetic oxidation of Fe(II), H2S, H2 and NO2−. Photoarsenotrophy was recently identified from Paoha Island's (Mono Lake, CA) arsenic-rich hot springs. The genomes of several photoarsenotrophs revealed a gene cluster, arxB2AB1CD, where arxA is predicted to encode for the sole arsenite oxidase. The role of arxA in photosynthetic arsenite oxidation was confirmed by disrupting the gene in a representative photoarsenotrophic bacterium, resulting in the loss of light-dependent arsenite oxidation. In situ evidence of active photoarsenotrophic microbes was supported by arxA mRNA detection for the first time, in red-pigmented microbial mats within the hot springs of Paoha Island. This work expands on the genetics for photosynthesis coupled to new electron donors and elaborates on known mechanisms for arsenic metabolism, thereby highlighting the complexities of arsenic biogeochemical cycling.

The involvement of prokaryotes in the redox reactions of arsenic occurring between its +5 [arsenate; As(V)] and +3 [arsenite; As(III)] oxidation states has been well established. Most research to date has focused upon circum-neutral pH environments (e.g., freshwater or estuarine sediments) or arsenic-rich “extreme” environments like hot springs and soda lakes. In contrast, relatively little work has been conducted in acidic environments. With this in mind we conducted experiments with sediments taken from the Herman Pit, an acid mine drainage impoundment of a former mercury (cinnabar) mine. Due to the large adsorptive capacity of the abundant Fe(III)-rich minerals, we were unable to initially detect in solution either As(V) or As(III) added to the aqueous phase of live sediment slurries or autoclaved controls, although the former consumed added electron donors (i.e., lactate, acetate, hydrogen), while the latter did not. This prompted us to conduct further experiments with diluted slurries using t...

Aims In vivo studies suggest a positive influence of fresh frozen plasma (FFP) on endothelial properties and vascular barrier function, leading to improved outcomes in animal sepsis models as well as in major abdominal surgery. However, those effects are incompletely described. It was our aim to evaluate in vitro effects of FFP on endothelial key functions and to identify underlying mechanisms. Materials and methods Human pulmonary microvascular endothelial cells (HPMECs) were prestimulated with LPS, followed by incubation with FFP. Permeability for FITC-dextran was assessed, and intercellular gap formation was visualized. NF-κB nuclear translocation and expression of pro-inflammatory, pro-adhesion, and leakage-related genes were evaluated, and monocyte adhesion to ECs was assessed. Intracellular cAMP levels as well as phosphorylation of functional proteins were analyzed. In patients undergoing major abdominal surgery, Syndecan-1 serum levels were assessed prior to and following FFP transfusion. Key findings Post-incubation of HPMVECs with FFP increased intracellular cAMP levels that had been decreased by preceding LPS stimulation. On one hand, this reduced endotoxin-mediated upregulation of IL-8, ICAM-1, VCAM-1, VEGF, and ANG-2. Impaired phosphorylation of functional proteins was restored, and intercellular cohesion and barrier function were rescued. On the other hand, NF-κB nuclear translocation as well as monocyte adhesion was markedly increased by the combination of LPS and FFP. Syndecan-1 serum levels were lower in surgery patients that were transfused with FFP compared to those that were not. Significance Our data provide evidence for a differential modulation of crucial endothelial properties by FFP, potentially mediated by elevation of intracellular cAMP levels.

In tall conifers, leaf structure can vary dramatically with height due to decreasing water potential (Ψ) and increasing light availability. This variation in leaf structure can have physiological consequences such as increased respiratory costs, reduced internal carbon dioxide conductance rates and ultimately reduced maximum photosynthetic rates (Amax). In Picea sitchensis (Bong.) Carrière, the leaf structure varies along the vertical gradient in ways that suggest compensatory changes to enhance photosynthesis, and this variation seems to be driven largely by light availability rather than by Ψ. These trends in leaf structure coupled with remarkably fast growth rates and dependence on moist environments inspire two important questions about P. sitchensis: (i) does foliar water uptake minimize the adverse effects of decreasing Ψ with height on leaf structure, and (ii) do trends in leaf structure increase photosynthetic rates despite increasing height? To answer these questions, we measured foliar water uptake capacity, predawn (Ψpd) and midday water potential and gas-exchange rates as they varied between 25- and 89-m heights in 300-year-old P. sitchensis trees in northwestern California. Our major findings for P. sitchensis include the following: (i) foliar water uptake capacity was quite high relative to published values for other woody species; (ii) foliar water uptake capacity increased between the crown base and treetop; (iii) wet season Ψpd was higher than predicted by the gravitational potential gradient, indicating foliar water uptake; and (iv) the maximum photosynthetic rate increased with height, presumably due to shifts in leaf structure between the crown base and treetop, mitigating height-related decreases in Amax. These findings suggest that together, the use of fog, dew and rain deposits on leaves and shifts in the leaf structure to conserve and possibly enhance photosynthetic capacity likely contribute to the rapid growth rates measured in this species.

Ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) is a critical yet severely inefficient enzyme that catalyses the fixation of virtually all of the carbon found on Earth. Here, we report a functional metagenomic selection that recovers physiologically active RubisCO molecules directly from uncultivated and largely unknown members of natural microbial communities. Selection is based on CO2 -dependent growth in a host strain capable of expressing environmental deoxyribonucleic acid (DNA), precluding the need for pure cultures or screening of recombinant clones for enzymatic activity. Seventeen functional RubisCO-encoded sequences were selected using DNA extracted from soil and river autotrophic enrichments, a photosynthetic biofilm and a subsurface groundwater aquifer. Notably, three related form II RubisCOs were recovered which share high sequence similarity with metagenomic scaffolds from uncultivated members of the Gallionellaceae family. One of the Gallionellaceae RubisCOs was purified and shown to possess CO2 /O2 specificity typical of form II enzymes. X-ray crystallography determined that this enzyme is a hexamer, only the second form II multimer ever solved and the first RubisCO structure obtained from an uncultivated bacterium. Functional metagenomic selection leverages natural biological diversity and billions of years of evolution inherent in environmental communities, providing a new window into the discovery of CO2 -fixing enzymes not previously characterized.

We aimed at identifying a model that dynamically predicts future need for renal replacement therapy (RRT) in intensive care unit (ICU) patients and can easily be implemented for online monitoring at the bedside. 7290 interdisciplinary ICU admissions were investigated. Patients with

Aims The incidence of secondary systemic fungal infections has sharply increased in bacterial septic patients. Antimycotics exhibit immunomodulatory properties, yet these effects are incompletely understood in secondary systemic fungal infections following bacterial sepsis. We investigated a model of systemic inflammation to determine whether antimycotics (liposomal amphotericin B (L-AMB), itraconazol (ITC), and anidulafungin (ANI)) modulate the gene and protein expression as well as the phagocytic activity of lipopolysaccharide (LPS)-stimulated human monocytes. Main methods THP-1 monocytes were incubated with L-AMB, ITC or ANI and LPS. Gene expression levels of cytokines (TNF- , IL-1 , IL-6, and IL-10) were measured after 2 h, 6 h, and 24 h. Cytokine protein levels were evaluated after 24 h and phagocytic activity was determined following co-incubation with Escherichia coli. Key findings All antimycotics differentially modulated the gene and protein expression of cytokines in sepsis-like conditions. In the presence of LPS, we identified L-AMB as immunosuppressive, whereas ITC demonstrated pro-inflammatory properties. Both compounds induced remarkably less phagocytosis. Significance Our study suggests that antimycotics routinely used in septic patients alter the immune response in sepsis-like conditions by modulating cytokine gene and protein expression levels and phagocytic activity. Future treatment strategies should consider the immune status of the host and apply antimycotics accordingly in bacterial septic patients with secondary fungal infections.

Conventional antibiotics exhibit immunomodulatory properties beneficial in the treatment of sepsis. Antibiotic-resistant Gram-positive bacteria have become a problem in sepsis therapy, giving rise to increased use of last-resort antibiotics; for example, linezolid (LIN), vancomycin (VAN) and daptomycin (DAP). As the immunomodulatory properties of these antibiotics in treating sepsis are unknown, this study examined the effect of VAN, LIN and DAP on the immune response under sepsis-like conditions in vitro. Lipopolysaccharide (LPS)-activated THP-1 monocytes were incubated with LIN, VAN or DAP. Gene expression of cytokines (TNFα, IL-1β, IL-6, IL-10) and Toll-like receptors (TLR1, 2, 4, 6, 7 and 9) was monitored and phagocytosis was determined following coincubation with E. coli. The antibiotics differentially modulated the gene expression of the investigated cytokines. While LIN and VAN upregulated the expression of all TLRs, DAP downregulated mRNA levels of TLR1, TLR2 and TLR6, which recognize pathogen-associated molecular patterns from Gram-positive bacteria. In addition, LIN inhibited, whereas VAN promoted the phagocytic activity of monocytes. Our results suggest that LIN and VAN possess pro-inflammatory properties, whereas DAP might reduce the immune response to Gram-positive bacteria in sepsis. Furthermore, VAN might be beneficial in the prevention of Gram-negative infections by increasing the phagocytosis of E. coli.

Nitrogen (N) management strategies that maintain high crop productivity with reduced water quality impacts are needed for tile-drained landscapes of the US Midwest. The objectives of this study were to determine the effect of N application rate, timing, and fall nitrapyrin addition on tile drainage nitrate losses, corn ( L.) yield, N recovery efficiency, and postharvest soil nitrate content over 3 yr in a corn-soybean [ (L.) Merr.] rotation. In addition to an unfertilized control, the following eight N treatments were applied as anhydrous ammonia in a replicated, field-scale experiment with both corn and soybean phases present each year in Illinois: fall and spring applications of 78, 156, and 234 kg N ha, fall application of 156 kg N ha + nitrapyrin, and sidedress (V5-V6) application of 156 kg N ha. Across the 3-yr study period, increases in flow-weighted NO concentrations were found with increasing N rate for fall and spring N applications, whereas N load results were variable. At the same N rate, spring vs. fall N applications reduced flow-weighted NO concentrations only in the corn-soybean-corn rotation. Fall nitrapyrin and sidedress N treatments did not decrease flo8w-weighted NO concentrations in either rotation compared with fall and spring N applications, respectively, or increase corn yield, crop N uptake, or N recovery efficiency in any year. This study indicates that compared with fall N application, spring and sidedress N applications (for corn-soybean-corn) and sidedress N applications (for soybean-corn-soybean) reduced 3-yr mean flow-weighted NO concentrations while maintaining yields.

Background Perioperative and critically ill patients are often exposed to iron (in the form of parenteral-iron administration or blood transfusion) and inflammatory stimuli, but the effects of iron loading on the inflammatory response are unclear. Recent data suggest that mitochondrial reactive oxygen species have an important role in the innate immune response and that increased mitochondrial reactive oxygen species production is a result of dysfunctional mitochondria. We tested the hypothesis that increased intracellular iron potentiates lipopolysaccharide-induced inflammation by increasing mitochondrial reactive oxygen species levels. Methods Murine macrophage cells were incubated with iron and then stimulated with lipopolysaccharide. C57BL/6 wild-type mice were intraperitoneally injected with iron and then with lipopolysaccharide. Markers of inflammation and mitochondrial superoxide production were examined. Mitochondrial homeostasis (the balance between mitochondrial biogenesis and destruction) was assessed, as were mitochondrial mass and the proportion of nonfunctional to total mitochondria. Results Iron loading of mice and cells potentiated the inflammatory response to lipopolysaccharide. Iron loading increased mitochondrial superoxide production. Treatment with MitoTEMPO, a mitochondria-specific antioxidant, blunted the proinflammatory effects of iron loading. Iron loading increased mitochondrial mass in cells treated with lipopolysaccharide and increased the proportion of nonfunctional mitochondria. Iron loading also altered mitochondrial homeostasis to favor increased production of mitochondria. Conclusions Acute iron loading potentiates the inflammatory response to lipopolysaccharide, at least in part by disrupting mitochondrial homeostasis and increasing the production of mitochondrial superoxide. Improved understanding of iron homeostasis in the context of acute inflammation may yield innovative therapeutic approaches in perioperative and critically ill patients.

Aims Toll-like receptor (TLR)9 ligand CpG-oligodeoxynucleotide (CpG-ODN) exerts preconditioning in myocardial ischemia/reperfusion. We hypothesized a postconditioning effect of CpG-ODN in a murine closed-chest model of myocardial infarction. Materials and Methods C57BL/6 (12 weeks, male, WT) mice were instrumented at the left anterior descending artery, then allowed 5d of recovery before 30 min ischemia. Treatments comprised: 1) PBS: 250 μl phosphate buffer solution intraperitoneally 5 min before reperfusion and 2) IPC (ischemic postconditioning): 3 twenty-second reperfusion and occlusion episodes at the end of ischemia 3) CpG-ODN: 1668 thioate 0.2 μmol/kg BW intraperitoneally 5 min before reperfusion. Infarct size was assessed via triphenyltetrazolium chloride (TTC) staining after 2 and 24 h reperfusion. Myocardial mRNA-expression of cytokines was measured using real-time PCR after 2 h reperfusion. Phosphatidylinositol-3 kinase (PI3K)-inhibitor wortmannin was injected intraperitoneally in WT 15 min before postconditioning and PBS in each group. Cardiac function in WT was assessed with a left-ventricular pressure-volume catheter at 24 h reperfusion. Key findings Following 30 min ischemia and 2 h reperfusion, infarct size was diminished by 90% in WT postconditioned with CpG-ODN (2.4 ± 1.55 IS/AAR%) and IPC (1.98 ± 1.03 IS/AAR%) compared to PBS mice (23.2 ± 3.97 IS/AAR%). Infarct size increased following 24 h reperfusion but the differences remained robust. Expression of TNF-α and IL-10 was increased in CpG-ODN. Wortmannin abolished the postconditioning effect of CpG-ODN and IPC. Ejection fraction and preload-recruitable stroke work were significantly greater in CpG-ODN mice. Significance CpG-ODN confers postconditioning via activation of TLR9. Cardiac function is preserved following CpG-ODN postconditioning. The PI3K -inhibitor wortmannin attenuates CpG-ODN postconditioning.

Maintaining nitrogen retention efficiency (NRE) is crucial in minimizing N losses when intensifying management of temperate grasslands. Our aim was to evaluate how grassland management practices and sward compositions affect NRE (1 − N losses/soil available N), defined as the efficiency with which soil available N is retained in an ecosystem. A three-factorial grassland management experiment was established with two fertilization treatments (without and combined N, phosphorus and potassium fertilization), two mowing frequencies (cut once and thrice per year) and three sward compositions (control, monocot- and dicot-enhanced swards). We measured N losses as leaching and nitrous oxide emissions, and soil available N as gross N mineralization rates. Fertilization increased N losses due to increased nitrification and decreased microbial N immobilization, and consequently decreased NRE. Intensive mowing partly dampened high N losses following fertilization. Sward compositions influenced NRE but not N losses: control swards that developed for decades under extensive management had the highest NRE, whereas monocot-enhanced sward had the lowest NRE. NRE was highly correlated with microbial NH 4 + immobilization and microbial biomass and only marginally correlated with plant N uptake, underlining the importance of microbial N retention in the soil-plant system. Microbial N retention is reflected in NRE but not in indices commonly used to reflect plant response. NRE was able to capture the effects of sward composition and fertilization whereas N losses were only sensitive to fertilization; thus, NRE is a better index when evaluating environmental sustainability of sward compositions and management practices of grasslands.

Several strategies appear suitable to improve iron and zinc bioavailability from fortified maize, and fortification per se will increase the intake of bioavailable iron and zinc. Corn masa flour or whole maize should be fortified with sodium iron ethylenediaminetetraacetate (NaFeEDTA), ferrous fumarate, or ferrous sulfate, and degermed corn flour should be fortified with ferrous sulfate or ferrous fumarate. The choice of zinc fortificant appears to have a limited impact on zinc bioavailability. Phytic acid is a major inhibitor of both iron and zinc absorption. Degermination at the mill will reduce phytic acid content, and degermed maize appears to be a suitable vehicle for iron and zinc fortification. Enzymatic phytate degradation may be a suitable home-based technique to enhance the bioavailability of iron and zinc from fortified maize. Bioavailability experiments with low phytic acid-containing maize varieties have suggested an improved zinc bioavailability compared to wild-type counterparts. The bioavailability of folic acid from maize porridge was reported to be slightly higher than from baked wheat bread. The bioavailability of vitamin A provided as encapsulated retinyl esters is generally high and is typically not strongly influenced by the food matrix, but has not been fully investigated in maize.

Beyond lipid lowering, statins are supposed to exert pleiotropic effects positively influencing the progression of atherosclerotic lesions. The development of such lesions is associated with increased release of angiopoietin-2 (Ang-2), an endothelial cell-specific protein growth factor stored in Weibel-Palade bodies (WPBs). The aim of our study was to examine whether statin pretreatment influences the release of Ang-2 from endothelial cells. Stimulation of HUVECs and HMVECs with PMA, thrombin or histamine resulted in significant release of Ang-2, as evidenced by ELISA. Pretreatment with simvastatin and mevastatin suppressed this release to basal level, while pravastatin had no effect. Simvastatin itself increased nitric oxide (NO, EC number 1.14.13.39) synthesis, measured by Griess reaction. Combining the statin pretreatment with the eNOS inhibitor L-NNA as well as bypassing the HMG-CoA reductase (EC number: 1.1.1.34) by adding mevalonic acid or geranyl pyrophosphate restored the exocytotic effect of PMA. Immunofluorescence microscopy showed that depletion of WPBs upon PMA stimulation ceased after pretreatment with simvastatin. This study demonstrates a potent suppressive effect of statins on the release of Ang-2 from endothelial cells. Regarding its harmful effects in the development of atherosclerotic lesions, our data provide further insight into the mechanisms of the anti-atherogenic potential of statins.

In fracture and bone defect healing, MSCs largely drive tissue regeneration. MSCs have been shown to promote angiogenesis both in vivo and in vitro. Angiogenesis is a prerequisite to large tissue reconstitution. The present study investigated how mechanical loading of MSCs influences their proangiogenic capacity. The results show a significant enhancement of angiogenesis by conditioned media from mechanically stimulated compared with unstimulated MSCs in two-dimensional tube formation and three-dimensional spheroid sprouting assays. In particular, proliferation but not migration or adhesion of endothelial cells was elevated. Promotion of angiogenesis was dependent upon fibroblast growth factor receptor 1 (FGFR1) signaling. Moreover, stimulation of tube formation was inhibited by vascular endothelial growth factor receptor (VEGFR) tyrosine kinase blocking. Screening for the expression levels of different soluble regulators of angiogenesis revealed an enrichment of matrix metalloprotease 2, transforming growth factor β1, and basic fibroblast growth factor but not of vascular endothelial growth factor in response to mechanical stimulation. In conclusion, mechanical loading of MSCs seems to result in a paracrine stimulation of angiogenesis, most likely by the regulation of a network of several angiogenic molecules. The underlying mechanism appears to be dependent on the FGFR and VEGFR signaling cascades and might be mediated by an additional cross-talk with other pathways. Disclosure of potential conflicts of interest is found at the end of this article.

Compared to other population groups in the United States, caries (tooth decay) is a disproportionately prevalent disease among Latino populations, especially among low-income and rural sub-groups and children under five years of age. Fluoride is a primary preventive for caries. While water fluoridation is a major and effective public health means for delivering fluoride on a mass scale, it does not reach many rural areas or population groups such as Latinos who eschew drinking water from municipal sources. This study examines the acceptability to such groups of salt fluoridation, an alternate means of delivering fluoride long used on a global scale. An ethnographic study in California's rural Central Valley was performed. Thirty individual interviews and 5 focus groups (N = 61) were conducted in Spanish to investigate low-income Latino migrant caregivers' experiences, views and understandings of domestic salt, oral health, caries prevention and fluoride. Audio data were transcribed, translated, coded and thematically analyzed. Table salt was readily available and frequently consumed. Both adult and child daily sodium consumption was high. Despite a general feeling that it was good, and present in dentifrices or dietary supplements, most participants had little knowledge about fluoride. Concerns were raised about cardio-vascular and other possibly deleterious effects if an increase in salt consumption occurred because fluoridated salt was viewed as having 'extra' benefits. Once informed about fluoride's safety and role in caries prevention, most participants expressed willingness to use fluoridated salt, especially if it benefitted children. Reassurance about its safety and benefits, and demonstration of its taste, were important aspects of acceptance. Taste was paramount. Participants would not consume more fluoridated salt than their current salt as that would result in unpleasant changes in food flavor and taste. While salt fluoridation is acceptable, the feasibility of producing and distributing fluoridated salt in the United States is, however, complex and challenging.

Agriculture is an important source of the greenhouse gas nitrous oxide (N 2 O) and the atmospherically important nitric oxide (NO). We evaluated the effects of different grazers and plant species composition on N 2 O and NO emissions in temperate grassland. Paddocks were grazed rotationally by either cattle or sheep. Mean N 2 O emissions were 38.7 μg N 2 O-N m −2 h −1 , mean NO emissions 2.4 μg NO-N m −2 h −1 . Cumulative NO-N emissions were larger for sheep- than for cattle-grazed paddocks. Plant species composition was insignificant compared to the effect of grazers on N oxide emissions. In a controlled application experiment, plots with cattle excreta showed larger N 2 O emissions than plots with sheep excreta, reaching peak emissions of 1921 μg N 2 O-N m −2 h −1 on cattle urine patches compared to 556 μg N 2 O-N m −2 h −1 on sheep urine patches, related to different N-inputs per excretion. Peak emissions of dung-treated plots were much smaller. The N 2 O emission factors were 0.4% for cattle urine, 0.5% for sheep urine, 0.05% for cattle dung and 0.09% for sheep dung. N oxide emissions on the paddock scale were larger for sheep- compared to cattle-grazing, despite larger emissions per cattle excretion. We attributed this to the more even spread of sheep excreta compared to cattle excreta.

Summary Arsenotrophy, growth coupled to autotrophic arsenite oxidation or arsenate respiratory reduction, occurs only in the prokaryotic domain of life. The enzymes responsible for arsenotrophy belong to distinct clades within the DMSO reductase family of molybdenum-containing oxidoreductases: specifi- cally arsenate respiratory reductase, ArrA, and arsen- ite oxidase, AioA (formerly referred to as AroA and AoxB). A new arsenite oxidase clade, ArxA, repre- sented by the haloalkaliphilic bacterium Alkalilimni- cola ehrlichii strain MLHE-1 was also identified in the photosynthetic purple sulfur bacterium Ectothiorho- dospira sp. strain PHS-1. A draft genome sequence of PHS-1 was completed and an arx operon similar to MLHE-1 was identified. Gene expression studies showed that arxA was strongly induced with arsenite. Microbial ecology investigation led to the identifica- tion of additional arxA-like sequences in Mono Lake and Hot Creek sediments, both arsenic-rich environ- ments in California. Phylogenetic analyses placed these sequences as distinct members of the ArxA clade of arsenite oxidases. ArxA-like sequences were also identified in metagenome sequences of several alkaline microbial mat environments of Yellowstone National Park hot springs. These results suggest that ArxA-type arsenite oxidases appear to be widely dis- tributed in the environment presenting an opportunity for further investigations of the contribution of Arx- dependent arsenotrophy to the arsenic biogeochemi- cal cycle.

Life is mostly composed of the elements carbon, hydrogen, nitrogen, oxygen, sulfur, and phosphorus. Although these six elements make up nucleic acids, proteins, and lipids and thus the bulk of living matter, it is theoretically possible that some other elements in the periodic table could serve the same functions. Here, we describe a bacterium, strain GFAJ-1 of the Halomonadaceae, isolated from Mono Lake, California, that is able to substitute arsenic for phosphorus to sustain its growth. Our data show evidence for arsenate in macromolecules that normally contain phosphate, most notably nucleic acids and proteins. Exchange of one of the major bio-elements may have profound evolutionary and geochemical importance.

Phosphorus fertilizer is frequently fall-applied for corn (Zea mays L.). Diammonium-(DAP) or monoammonium-phosphates (MAP) are the preferred P fertilizers and the N in MAP and DAP is assumed to be available for corn. Our objective was to evaluate the effect of application time on availability of ammoniated-phosphate N to the crop in two different environments. In a laboratory study, MAP and DAP at rates of 70 and 140 mg N kg -1 soil were incubated at either 80 or 120% field capacity (FC). Nitrification proceeded rapidly in all treatments, but nitrate levels fell to near zero by 8 wk at 120% FC while remaining above 50% after 16 wk at 80% FC. In a 3-yr study at Waseca, MN and Urbana, IL with a factorial combination of three N sources [DAP, MAP, ammonium-sulfate (AMS)], two application times (fall, spring), and two N rates (45, 90 kg N ha -1 ), N rate and application time, but not N source, affected grain yield and inorganic soil-N concentrations. Yields were 0.71 and 0.37 Mg ha -1 greater for spring than fall applications at Urbana and Waseca, respectively. Late May soil-N recovery ranged from 31 to 35 and 90 to 100% for fall and spring applications, respectively. It thus appears that only about one-third of N applied in the fall as ammoniated phosphates at typical rates is available to the next year's corn crop in Corn Belt mollisols. Colder, drier fall to spring conditions may increase this proportion, while warmer, wetter conditions would be expected to lower availability.

Although arsenic is highly toxic to most organisms, certain prokaryotes are known to grow on and respire toxic metalloids of arsenic (i.e., arsenate and arsenite). Two enzymes are known to be required for this arsenic-based metabolism: (i) the arsenate respiratory reductase (ArrA) and (ii) arsenite oxidase (AoxB). Both catalytic enzymes contain molybdopterin cofactors and form distinct phylogenetic clades (ArrA and AoxB) within the dimethyl sulfoxide (DMSO) reductase family of enzymes. Here we report on the genetic identification of a “new” type of arsenite oxidase that fills a phylogenetic gap between the ArrA and AoxB clades of arsenic metabolic enzymes. This “new” arsenite oxidase is referred to as ArxA and was identified in the genome sequence of the Mono Lake isolate Alkalilimnicola ehrlichii MLHE-1, a chemolithoautotroph that can couple arsenite oxidation to nitrate reduction. A genetic system was developed for MLHE-1 and used to show that arxA (gene locus ID mlg _ 0216 ) was required for chemoautotrophic arsenite oxidation. Transcription analysis also showed that mlg _ 0216 was only expressed under anaerobic conditions in the presence of arsenite. The mlg _ 0216 gene is referred to as arxA because of its greater homology to arrA relative to aoxB and previous reports that implicated Mlg_0216 (ArxA) of MLHE-1 in reversible arsenite oxidation and arsenate reduction in vitro . Our results and past observations support the position that ArxA is a distinct clade within the DMSO reductase family of proteins. These results raise further questions about the evolutionary relationships between arsenite oxidases (AoxB) and arsenate respiratory reductases (ArrA).

Spontaneous breathing during mechanical ventilation is gaining increasing importance during intensive care but is depressed by narcotics, such as opioids. Serotonin 1A-receptor (5-HT(1A)-R) agonists have been shown to antagonize opioid-induced ventilatory depression, but both enhancement and attenuation of nociceptive reflexes have been found with different experimental models. To clarify contradictory findings, we simultaneously determined dose-response functions of the standard 5-HT(1A)-R-agonist 8-OH-DPAT and two different opioids for spontaneous ventilation and nociception. Two hypotheses were tested: 1) 8-OH-DPAT at a dose to stimulate spontaneous breathing does not activate nociceptive reflexes. 2) 8-OH-DPAT does not diminish opioid-induced antinociception.(A) A dose-response relationship of 8-OH-DPAT, spontaneous phrenic nerve activity and a nociceptive C-fiber reflex (CFR) were established simultaneously in an in situ perfused, nonanesthetized, rat brainstem-spinal cord preparation. (B) Fentanyl was administered in situ to investigate the interaction with 8-OH-DPAT on phrenic nerve activity and nociceptive CFR. Additional experiments involved the selective 5-HT(1A)-R-antagonist WAY 100 635 to exclude effects of receptors other than 5-HT(1A)-R. (C) The effects of 8-OH-DPAT on spontaneous ventilation and nociceptive tail-flick reflex with and without morphine were verified in in vivo anesthetized rats.Low-dose 8-OH-DPAT (0.001 and 0.01 microM in situ, 0.1 microg/kg in vivo) enhanced nociceptive reflexes but did not activate spontaneous ventilation. On the contrary, high doses of 8-OH-DPAT (1 microM in situ and 10-100 microg/kg in vivo) stimulated ventilation, whereas nociceptive CFR amplitude in situ returned to baseline and tail-flick reflex was depressed in vivo. Opioid-induced ventilatory depression was antagonized by 8-OH-DPAT (1 microM in situ, and 10 microg/kg in vivo), whereas antinociception sustained. Selective 5-HT(1A)-R-antagonist WAY 100 635 (1 microM) prevented the effects of 8-OH-DPAT in situ.5-HT(1A)-R-agonist 8-OH-DPAT activates spontaneous breathing without diminishing opioid-induced antinociception in rats.

Soils with natric horizons are prevalent in South Central Illinois where Peorian age loess overlies older surfaces underlain by Illinoian till paleosols. Such soils are classified as Natraqualfs and are intermixed with Albaqualfs that do not have natric horizons. The Natraqualfs make up from a few percent to more than 35% of the landscape, with Albaqualfs making up most of the landscape. Warm-season crops of corn and soybean on Natraqualfs are drought susceptible and have reduced yields because of shallow rooting and lesser amounts of plant available stored soil moisture compared with Albaqualfs; however, cool-season crops, such as wheat, have more uniform yields between Natraqualf and Albaqualf soil areas. Na + was successfully moved to deeper soil depths within the soil when the natric horizon was treated with calcium chloride (CaCl 2 ) and leached with water. Calcium chloride was used rather than calcium sulfate because CaCl 2 solubility enhanced Ca ++ concentration during the short-term leaching period. Treated and untreated plots were cropped from 2001 to 2004; however, there was no increase in corn or soybean yield during these four growing seasons for treated versus untreated plots, although the Na + concentration was moved down in the profile from treatment with CaCl 2 and leaching.

Evaluating the effects of management practices on soil physical and chemical properties would be valuable to explain field-level variability in crop production. A 23-year-old experiment on a Muscatune soil (fine-silty, mixed, superactive, mesic, Aquic Argiudolls) in Illinois with five N rates [0 (N 0 ), 70 (N 1 ), 140 (N 2 ), 210 (N 3 ) and 280 (N 4 ) kg N ha −1 ] and two cropping systems [continuous corn ( Zea mays L.) (CC), and corn–soybean ( Glycine max (L.) Merr.) rotation (CS)] was evaluated. Specific objectives were to: (i) evaluate the effects of long-term N fertilization and cropping systems on field level changes in soil physical and chemical properties and crop yield, (ii) identify the most responsive soil physical and chemical properties to N fertilizer and crop management, and (iii) investigate the relationship between the selected soil properties and crop yield. Soil was collected in May 2004 to 30 cm depth and 20 soil physical and chemical properties were measured. The univariate analysis indicated that 14 soil properties were significantly influenced by at least one treatment effect (crops, N or crops × N). Due to multicollinearity among soil properties, principal component analysis (PCA) was used to group correlated properties, resulting in five soil properties such as soil organic carbon stock (OC stock), mean weight diameter (MWD), soil C:N ratio, exchangeable potassium (K + ) and gravimetric moisture content ( ω ). Finally, the multiple regression analysis performed between PCA derived soil properties and corn and soybean yields retained all the representative soil properties from PCA except ω as yield predictors for corn ( P R 2 = 0.39) from CC system, whereas none of the soil properties were significantly related to corn and soybean yields from CS system. The soil properties most influenced by long-term N fertilization of continuous corn were successfully identified with PCA and multiple regression. The insignificant relationship between corn and soybean yields from CS system and PCA derived soil properties might be due to the lack of response of soybean to N fertilization. This study shows the integrated use of multivariate and regression analyses in identifying yield determining soil properties by eliminating the multicollinearity among soil properties.

During sepsis, a severe systemic disorder, micronutrients often are decreased. Apoptosis is regarded as an important mechanism in the development of often significant immunosuppression in the course of the disease. This study aimed to investigate alpha-tocopherol and selenium in reference to apoptosis in patients with sepsis. 16 patients were enrolled as soon as they fulfilled the criteria of severe sepsis. 10 intensive care patients without sepsis and 11 healthy volunteers served as controls. alpha-Tocopherol, selenium and nucleosomes were measured in serum. Phosphatidylserine externalization and Bcl-2 expression were analyzed in T-cells by flow cytometry. Serum alpha-tocopherol and selenium were decreased in severe sepsis but not in non-septic critically ill patients (p < 0.05). Conversely, markers of apoptosis were increased in sepsis but not in critically ill control patients: Nucleosomes were found to be elevated 3 fold in serum (p < 0.05) and phosphatidylserine was externalized on an expanded subpopulation of T-cells (p < 0.05) while Bcl-2 was expressed at lower levels (p < 0.05). The decrease of micronutrients correlated with markers of accelerated apoptosis. Accelerated apoptosis in sepsis is associated with low alpha-tocopherol and selenium. The results support the investigation of micronutrient supplementation strategies in severe sepsis.

Conductive multifilament fibres are the fundamental core for wearable technology in the biomedical engineering fields, as conductors in sensors or in bio-sensing textiles for healthcare. This contribution presents a model, simulation and the experimental analysis of a metal-polymer hybrid fibre based microstrip resonator for high frequency characterisation. The high frequency electromagnetic field simulation (HFSS) by ANSYS® is used for the modelling and finite element based simulation. It follows the design and manufacturing of the metal-polymer hybrid fibre based microstrip resonator for analysis of the scattering parameter measurements and the quality factors to 10 GHz with a vector network analyser (VNA) by Rohde & Schwarz®. Simulations and analysis also compare solid matter wire with the metal-polymer hybrid fibre.

The influence of CYP2D6 genotype and CYP2D6 inhibitors on enantiomeric plasma levels of tramadol and O-desmethyltramadol as well as response to tramadol was investigated. One hundred and seventy-four patients received one hundred intravenous tramadol 3 mg/kg for postoperative analgesia. Blood samples drawn 30, 90, and 180 min after administration were analyzed for plasma concentrations of the enantiomers (+)-, (-)tramadol and (+)-, (-)O-desmethyltramadol by liquid chromatography-tandem mass spectrometry. Different CYP2D6 genotypes displaying zero (poor metabolizer (PM)), one (heterozygous individual (HZ)/intermediate metabolizer (IM)), two extensive metabolizer (EM), and three (ultra rapid metabolizer (UM)) active genes were compared. Concentrations of O-desmethyltramadol differed in the four genotype groups. Median (1/3 quartile) area under the concentration-time curves for (+)O-desmethyltramadol were 0 (0/11.4), 38.6 (15.9/75.3), 66.5 (17.1/118.4), and 149.7 (35.4/235.4) ng x h/ml for PMs, HZ/IMs, EMs, and UMs (P

Core level shift scanned-energy mode photoelectron diffraction using the two distinct components of the C 1s emission has been used to determine the structure of the Pt(1 1 1)c(√3 × 5)rect.–CO phase formed by 0.6 ML of adsorbed CO. The results confirm earlier assignments of these components to CO in atop and bridging sites, further confirm that the best structural model involves a 2:1 occupation ratio of these two sites, and provides quantitative structural parameter values. In particular the Pt–C chemisorption bondlengths for the atop and bridging sites are, respectively, 1.86 ± 0.02 Å and 2.02 ± 0.04 Å. These values are closely similar to those found in the 0.5 ML coverage c(4 × 2) phase, involving an atop:bridge occupation ratio of 1:1, obtained in earlier quantitative low energy electron diffraction studies. The results also indicate a clear tilt of the molecular axis of atop CO species in this compression phase, consistent with the finding of an earlier electron-stimulated desorption ion angular distribution investigation

A facultative chemoautotrophic bacterium, strain MLHE-1T, was isolated from Mono Lake, an alkaline hypersaline soda lake in California, USA. Cells of strain MLHE-1T were Gram-negative, short motile rods that grew with inorganic electron donors (arsenite, hydrogen, sulfide or thiosulfate) coupled with the reduction of nitrate to nitrite. No aerobic growth was attained with arsenite or sulfide, but hydrogen sustained both aerobic and anaerobic growth. No growth occurred when nitrite or nitrous oxide was substituted for nitrate. Heterotrophic growth was observed under aerobic and anaerobic (nitrate) conditions. Cells of strain MLHE-1T could oxidize but not grow on CO, while CH4 neither supported growth nor was it oxidized. When grown chemoautotrophically, strain MLHE-1T assimilated inorganic carbon via the Calvin–Benson–Bassham reductive pentose phosphate pathway, with the activity of ribulose 1,5-bisphosphate carboxylase (RuBisCO) functioning optimally at 0.1 M NaCl and at pH 7.3. Strain MLHE-1T grew over broad ranges of pH (7.3–10.0; optimum, 9.3), salinity (15–190 g l−1; optimum 30 g l−1) and temperature (13–40 °C; optimum, 30 °C). Phylogenetic analysis of 16S rRNA gene sequences placed strain MLHE-1T in the class Gammaproteobacteria (family Ectothiorhodospiraceae) and most closely related to Alkalispirillum mobile (98.5 %) and Alkalilimnicola halodurans (98.6 %), although none of these three haloalkaliphilic micro-organisms were capable of photoautotrophic growth and only strain MLHE-1T was able to oxidize As(III). On the basis of physiological characteristics and DNA–DNA hybridization data, it is suggested that strain MLHE-1T represents a novel species within the genus Alkalilimnicola for which the name Alkalilimnicola ehrlichii is proposed. The type strain is MLHE-1T (=DSM 17681T=ATCC BAA-1101T). Aspects of the annotated full genome of Alkalilimnicola ehrlichii are discussed in the light of its physiology.

A novel chemolithotrophic metabolism based on a mixed arsenic-sulfur species has been discovered for the anaerobic deltaproteobacterium, strain MLMS-1, a haloalkaliphile isolated from Mono Lake, California, U.S. Strain MLMS-1 is the first reported obligate arsenate-respiring chemoautotroph which grows by coupling arsenate reduction to arsenite with the oxidation of sulfide to sulfate. In that pathway the formation of a mixed arsenic-sulfur species was reported. That species was assumed to be monothioarsenite ([H2As(III)S(-II)O2](-)), formed as an intermediate by abiotic reaction of arsenite with sulfide. We now report that this species is monothioarsenate ([HAs(V)S(-II)O3](2-)) as revealed by X-ray absorption spectroscopy. Monothioarsenate forms by abiotic reaction of arsenite with zerovalent sulfur. Monothioarsenate is kinetically stable under a wide range of pH and redox conditions. However, it was metabolized rapidly by strain MLMS-1 when incubated with arsenate. Incubations using monothioarsenate confirmed that strain MLMS-1 was able to grow (μ = 0.017 h(-1)) on this substrate via a disproportionation reaction by oxidizing the thio-group-sulfur (S(-II)) to zerovalent sulfur or sulfate while concurrently reducing the central arsenic atom (As(V)) to arsenite. Monothioarsenate disproportionation could be widespread in nature beyond the already studied arsenic and sulfide rich hot springs and soda lakes where it was discovered.

Background and objective Proinflammatory cytokines as well as nitric oxide (NO) play a major role in mediating the response to lipopolysaccharide (LPS). The present study tested the hypothesis that LPS induces proinflammatory cytokines in the lung via the Toll-like receptor 4 (TLR4)/CD14 signalling cascade. Methods Control mice and TLR4-deficient (TLR4-D) mice were used to test TLR4-mediated effects of LPS. Both strains received either Escherichia coli LPS (20 mg kg-1 intraperitoneal) or saline and their lungs were collected at different time points. Pulmonary nuclear factor kappaB (NFkappaB) activation was investigated with electromobility shift assay. mRNA expression of inflammatory mediators and their corresponding receptors were detected with Ribonuclease Protection Assay. Protein expression was detected by ELISA and western blotting. Inducible NO synthase (iNOS) expression was monitored by RT-PCR and iNOS activity by conversion of l-arginine to citrulline. Immune cells were sampled by bronchoalveolar lavage (BAL) and classified. Results LPS application induced CD14-, but not TLR4 protein expression in control mice. Activation of pulmonary NFkappaB was observed within 60 min in control, but not in TLR4-D mice. Six hours of LPS administration induced a significant increase in pulmonary tumour necrosis factor alpha-, interleukin-1beta- and interleukin-6 mRNA and protein expression in control mice compared to TLR4-D mice. Furthermore, LPS induced a significantly higher increase of the iNOS expression and catalytic activity in control mice than in TLR4-D mice. BAL revealed an increase in total cell count in all LPS treated mice. Conclusion Our findings suggest that TLR4 plays a key role for regulating the expression of relevant cytokines within the lung during endotoxic shock.

A radioisotope method was devised to study bacterial respiratory reduction of arsenate in sediments. The following two arsenic-rich soda lakes in California were chosen for comparison on the basis of their different salinities: Mono Lake (∼90 g/liter) and Searles Lake (∼340 g/liter). Profiles of arsenate reduction and sulfate reduction were constructed for both lakes. Reduction of [ 73 As]arsenate occurred at all depth intervals in the cores from Mono Lake (rate constant [ k ] = 0.103 to 0.04 h −1 ) and Searles Lake ( k = 0.012 to 0.002 h −1 ), and the highest activities occurred in the top sections of each core. In contrast, [ 35 S]sulfate reduction was measurable in Mono Lake ( k = 7.6 ×10 4 to 3.2 × 10 −6 h −1 ) but not in Searles Lake. Sediment DNA was extracted, PCR amplified, and separated by denaturing gradient gel electrophoresis (DGGE) to obtain phylogenetic markers (i.e., 16S rRNA genes) and a partial functional gene for dissimilatory arsenate reduction ( arrA ). The amplified arrA gene product showed a similar trend in both lakes; the signal was strongest in surface sediments and decreased to undetectable levels deeper in the sediments. More arrA gene signal was observed in Mono Lake and was detectable at a greater depth, despite the higher arsenate reduction activity observed in Searles Lake. A partial sequence (about 900 bp) was obtained for a clone (SLAS-3) that matched the dominant DGGE band found in deeper parts of the Searles Lake sample (below 3 cm), and this clone was found to be closely related to SLAS-1, a novel extremophilic arsenate respirer previously cultivated from Searles Lake.

Transverse aortic constriction (TAC) results in a transient increase of proinflammatory cytokines, which return to baseline levels within 3 d. In contrast to cytokine baseline levels, the myocardium remains capable to respond even stronger to a new stimulus. As the molecular mechanisms for this phenomenon are unknown, we tested whether TAC modulates the innate immune system in mice and changes the inflammatory reaction to a new stimulus.Following 3 d of TAC or sham-operation procedure (SOP), LPS (20 mg/kg) or PBS (control) were administered intraperitoneal for 10 min as well as for 6 h. Hemodynamic parameters were recorded to measure the effects of TAC and LPS. After TAC/SOP alone CD14 expression was monitored and after additional 6 h of LPS/PBS the expression of CD14, TLR4 and proinflammatory cytokines were determined by western-blot, ELISA and RNase protection assay, respectively. Following TAC/SOP and 10 min of LPS/PBS, NFkappaB activation was investigated by EMSA.TAC induced cardiac hypertrophy and elevated blood pressure. LPS application led to hypotension and other symptoms of sepsis. CD14 expression increased after TAC alone and even further after additional LPS challenge. However, we did not detect changes of TLR4 expression. Also NFkappaB activation increased after LPS challenge higher in the TAC than in the SOP group. LPS-stimulation induced also higher cytokine expression in the TAC than in the SOP group.TAC modulates innate immunity by regulating the expression of CD14 and changes the myocardial tissue to respond more powerful to LPS.

Nitrogen fertilizer is a fundamental input for production of corn (Zea mays L.) that can move to ground and surface waters when over-applied. Previous research has shown that chlorophyll meter (CM) readings can indicate N stress in corn, but has not addressed whether the amount of N needed can be predicted by CM readings. Our objective was to evaluate whether CM readings can predict corn N need and yield response to N. Sixty-six N rate experiments were conducted in seven north-central states over a 4-yr period. Linear regression was used to relate absolute and relative CM readings over a range of growth stages to economically optimal N rate (EONR) and yield response to N applied at growth stage V7 or earlier. Chlorophyll meter readings at all growth stages from V5 to R5 were significantly related (P < 0.0001 in 22 of 24 models, P < 0.01 in 2 models) to EONR and yield response to N. Relationships were stronger for relative than for than absolute CM readings, and also were stronger when the corn had received no N fertilizer at planting. Coefficients of determination ranged from 0.53 to 0.76 for relative CM reading as a predictor of EONR or yield response to N, and were lower for the V5 to V9 stage than for later stages. Earlier research has indicated that measurements with this level of predictive accuracy can produce N rate recommendations that are more profitable than current N management practices. Our findings suggest that CM readings (and potentially other measures of corn color) are quantitatively related to early-season EONR and yield response to N over a wide range of environments with enough accuracy to be helpful in making management decisions.

Numerous chemical analyses of gaseous and particulate samples from laboratory flames provide a library of data on the polycyclic aromatic hydrocarbon (PAH) species found in diverse flame types burning fuels consisting of pure gaseous hydrocarbons. The diesel fuels utilized by the more complex combustion in compression ignition engines are composed of thousands of hydrocarbon species. Mass spectrometry by the laser microprobe and gas chromatography were used in a complementary manner to distinguish the PAHs originating in the fuel from those produced by engine combustion. The CxHy PAH products of premixed and diffusion flame processes, which also occur in the unsteady diesel combustion, range in mass from 128 u (two rings, x = 10 , y = 8 ) to beyond 350 u (eight rings, x = 28 , y = 14 ). Graphs of the number of hydrogen atoms y vs the number of carbon atoms x for the species found by many investigators of laboratory flames show these pyrogenic PAHs to lie on or near the staircase curve that describes the most stable, pericondensed, benzenoid PAHs. In contrast, samples of diesel fuels from the United Kingdom and the United States contain petrogenic alkyl-PAHs with high hydrogen contents. Samples of diesel particulate emissions typical of the 1990s from two different sources display the full mass range of PAHs from 128 to 350 u, including both the benzenoid PAHs and the alkyl-PAHs. Thus diesel emissions, in general, may contain petrogenic fuel components ranging up to 206 u and also the combustion-generated four- to seven-ring species in the 228 to 302 u mass range that have greater carcinogenic potency. The absence of petrogenic components larger than 206 u facilitates their detection and delineation from pyrogenic PAHs by methods of chemical analysis.

Cassiopeia A was observed using the Low-Band Antennas of the LOw Frequency ARray (LOFAR) with high spectral resolution. This allowed a search for radio recombination lines (RRLs) along the line-of-sight to this source. Five carbon-alpha RRLs were detected in absorption between 40 and 50 MHz with a signal-to-noise ratio of > 5 from two independent LOFAR datasets. The derived line velocities (v_LSR ~ -50 km/s) and integrated optical depths (~ 13 s^-1) of the RRLs in our spectra, extracted over the whole supernova remnant, are consistent within each LOFAR dataset and with those previously reported. For the first time, we are able to extract spectra against the brightest hotspot of the remnant at frequencies below 330 MHz. These spectra show significantly higher (15-80 %) integrated optical depths, indicating that there is small-scale angular structure on the order of ~1 pc in the absorbing gas distribution over the face of the remnant. We also place an upper limit of 3 x 10^-4 on the peak optical depths of hydrogen and helium RRLs. These results demonstrate that LOFAR has the desired spectral stability and sensitivity to study faint recombination lines in the decameter band., Accepted for publication in A&A Letters; 5 pages, 4 colour figures

The structure, thickness and adhesion to the substrate of the hard coatings used for protection against wear and corrosion are very important characteristics and they are strongly dependent on the internal stress induced into the coating during its growth. Development of a new class of coatings with a hardness of 2500–4500 HV0.1 and a thickness of 10–50 μm might increase significantly the application area of PVD coating in hydraulics, automotive, forming and even cutting tools. An ion assisted deposition method that combines magnetron sputtering and ion implantation (CMSII) as simultaneous processes has been used to produce nc-Ti 2 N/nc-TiN nanocomposite layers with such properties. In this paper, the cantilever method was used for in situ monitoring of the internal stress during the coating deposition. Generally, the internal stress produced by CMSII method is about half of that produced by magnetron sputtering without ion implantation. The structure of these coatings were analyzed by means of TEM. The adhesion and fracture properties of the coatings were analyzed by a mini version of a tensile test.

Increased fertilizer N uptake efficiency (FNUE) leads to more economical corn (Zea mays L.) production and lower environmental impact. Excessive N application reduces FNUE and may affect subsequent crop response through its influence on NO 3 -N carryover and the amount of readily mineralizable organic N in the soil. Our objective was to determine how prior fertilizer N application rate affects (i) grain yield and agronomic optimum N rate, (ii) contributions of fertilizer- and soil-derived N to N uptake, and (iii) FNUE. Labeled 15 NH 4 15 NO 3 was applied at 0, 67, 134, 201, or 268 kg N ha -1 to subplots within a continuous corn long-term N rate study. Estimates of FNUE were higher by the difference method (49-69%) than with the isotope ( 15 N) method (31-37%), and different trends were observed with each method as N application rate increased. The disparity between methods is consistent with a differential effect of long-term N application rate on mineralization-immobilization. Recovery of labeled N from the plant-soil system ranged from 71% at the 67 kg ha -1 N application rate to 64% at the 201 kg ha -1 application rate. Fertilizer N accounted for an increasing proportion of crop N uptake as the N rate was increased, but soil N uptake was always more extensive, accounting for 54 to 83% of total plant N. Crop uptake of labeled N during the second growing season after 15 N application ranged from 2.2 kg ha -1 with the lowest N rate to 7.8 kg ha -1 with the two highest rates.

A better understanding of how N management practices affect transformations and movement of fertilizer N may lead to more efficient N management. The objectives of this work were to determine how long-term N fertilizer history in a continuous corn (Zea mays L.) production system affects (i) movement of fertilizer N through the soil profile and (ii) cycling of fertilizer N between available and nonavailable soil forms. Nitrogen-15-labeled ammonium nitrate ( 15 NH 4 15 NO 3 ) was applied at 0, 67, 134, 201, or 268 kg N ha -1 to subplots of long-term N rate plots. Twenty to 55% of labeled N was converted into either organic or clay-fixed forms during the first growing season, with the percentage decreasing with increasing N application rate. Significantly more N was released from nonavailable forms in plots where the historical N application rate had exceeded the long-term optimum (186 kg ha -1 ) than in plots that received lower rates. Little fertilizer-derived N leached from the profile during the first growing season, but losses did occur during the off-season and subsequent growing season when N application rate was higher than the optimum. It was concluded that a history of excessive N application may decrease response of subsequent crops to fertilizer N due to greater release from nonavailable N forms, most likely as a result of increased mineralization of crop residues and recently formed soil organic N.

Searles Lake is a salt-saturated, alkaline brine unusually rich in the toxic element arsenic. Arsenic speciation changed from arsenate [As(V)] to arsenite [As(III)] with sediment depth. Incubated anoxic sediment slurries displayed dissimilatory As(V)-reductase activity that was markedly stimulated by H 2 or sulfide, whereas aerobic slurries had rapid As(III)-oxidase activity. An anaerobic, extremely haloalkaliphilic bacterium was isolated from the sediment that grew via As(V) respiration, using either lactate or sulfide as its electron donor. Hence, a full biogeochemical cycle of arsenic occurs in Searles Lake, driven in part by inorganic electron donors.

The oxidoreductase MIF is currently discussed as a new promising target of immunomodulatory therapy in patients with severe sepsis. An increasing body of evidence attributes an important role especially to intracellular MIF for regulation of endotoxin responsiveness as well as regulation of nuclear transcription factors. Up to now, measurement of MIF relied on ELISA techniques, lacking the ability to directly measure intracellular MIF and distinguish between different leukocyte subpopulations. Therefore, we developed a sensitive and robust flow cytometry-based method to reliably detect intracellular levels of MIF. This method can readily be applied in cultured cells as well as in subsets of human leukocytes in whole blood. Intracellular MIF content of whole-blood leukocyte subsets is detected simultaneously, and is individually determined for T-lymphocytes, B-lymphocytes, macrophages, and granulocytes, respectively. When tested in an ex vivo whole-blood stimulation system using PMA/Ionomycin the intracellular MIF content doubled in CD3+ T-lymphocytes and increased threefold in CD14+ macrophages. Baseline intracellular MIF levels of different leukocyte subpopulations were quantified in 22 healthy blood donors. Baseline intracellular MIF levels in T-lymphocytes are twice as high compared to those in B-lymphocytes and macrophages.

Periphyton ( Cladophora sp.) samples from a suburban stream lacking detectable dissolved As were able to reduce added As(V) to As(III) when incubated under anoxic conditions and, conversely, oxidized added As(III) to As(V) with aerobic incubation. Both types of activity were abolished in autoclaved controls, thereby demonstrating its biological nature. The reduction of As(V) was inhibited by chloramphenicol, indicating that it required the synthesis of new protein. Nitrate also inhibited As(V) reduction, primarily because it served as a preferred electron acceptor to which the periphyton community was already adapted. However, part of the inhibition was also caused by microbial reoxidation of As(III) linked to nitrate. Addition of [ 14 C]glucose to anoxic samples resulted in the production of 14 CO 2 , suggesting that the observed As(V) reduction was a respiratory process coupled to the oxidation of organic matter. The population density of As(V)-reducing bacteria within the periphyton increased with time and with the amount of As(V) added, reaching values as high as ∼10 6 cells ml −1 at the end of the incubation. This indicated that dissimilatory As(V) reduction in these populations was linked to growth. However, As(V)-respiring bacteria were found to be present, albeit at lower numbers (∼10 2 ml −1 ), in freshly sampled periphyton. These results demonstrate the presence of a bacterial population within the periphyton communities that is capable of two key arsenic redox transformations that were previously studied in As-contaminated environments, which suggests that these processes are widely distributed in nature. This assumption was reinforced by experiments with estuarine samples of Cladophora sericea in which we detected a similar capacity for anaerobic As(V) reduction and aerobic As(III) oxidation.