Genome-wide SNP detection, validation, and development of an 8K SNP array for apple

As high-throughput genetic marker screening systems are essential for a range of genetics studies and plant breeding applications, the International RosBREED SNP Consortium (IRSC) has utilized the Illumina Infinium® II system to develop a medium- to high-throughput SNP screening tool for genome-wide evaluation of allelic variation in apple (Malus×domestica) breeding germplasm. For genome-wide SNP discovery, 27 apple cultivars were chosen to represent worldwide breeding germplasm and re-sequenced at low coverage with the Illumina Genome Analyzer II. Following alignment of these sequences to the whole genome sequence of 'Golden Delicious', SNPs were identified using SoapSNP. A total of 2,113,120 SNPs were detected, corresponding to one SNP to every 288 bp of the genome. The Illumina GoldenGate® assay was then used to validate a subset of 144 SNPs with a range of characteristics, using a set of 160 apple accessions. This validation assay enabled fine-tuning of the final subset of SNPs for the Illumina Infinium® II system. The set of stringent filtering criteria developed allowed choice of a set of SNPs that not only exhibited an even distribution across the apple genome and a range of minor allele frequencies to ensure utility across germplasm, but also were located in putative exonic regions to maximize genotyping success rate. A total of 7867 apple SNPs was established for the IRSC apple 8K SNP array v1, of which 5554 were polymorphic after evaluation in segregating families and a germplasm collection. This publicly available genomics resource will provide an unprecedented resolution of SNP haplotypes, which will enable marker-locus-trait association discovery, description of the genetic architecture of quantitative traits, investigation of genetic variation (neutral and functional), and genomic selection in apple.

The genome of the domesticated apple (Malus × domestica Borkh.)

We report a high-quality draft genome sequence of the domesticated apple (Malus × domestica). We show that a relatively recent (>50 million years ago) genome-wide duplication (GWD) has resulted in the transition from nine ancestral chromosomes to 17 chromosomes in the Pyreae. Traces of older GWDs partly support the monophyly of the ancestral paleohexaploidy of eudicots. Phylogenetic reconstruction of Pyreae and the genus Malus, relative to major Rosaceae taxa, identified the progenitor of the cultivated apple as M. sieversii. Expansion of gene families reported to be involved in fruit development may explain formation of the pome, a Pyreae-specific false fruit that develops by proliferation of the basal part of the sepals, the receptacle. In apple, a subclade of MADS-box genes, normally involved in flower and fruit development, is expanded to include 15 members, as are other gene families involved in Rosaceae-specific metabolism, such as transport and assimilation of sorbitol.

Concerted HO2 elimination from alpha-Aminoalkylperoxyl free radicals : Experimental and theoretical evidence from the gas phase NH2 center dot CHCO2- + O-2 reaction

We have investigated the gas-phase reaction of the alpha-aminoacetate (glycyl) radical anion (NH2(sic)CHCO2-) with O-2 using ion trap mass spectrometry, quantum chemistry, and statistical reaction rate theory. This radical is found to undergo a remarkably rapid reaction with O-2 to form the hydroperoxyl radical (HO2(sic)) and an even-electron imine (NHCHCO2-), with experiments and master equation simulations revealing that reaction proceeds at the ion molecule collision rate. This reaction is facilitated by a low-energy concerted HO2(sic) elimination mechanism in the NH2CH(OO(sic))CO2- peroxyl radical. These findings can explain the widely observed free-radical-mediated oxidation of simple amino acids to amides plus alpha-keto acids (their imine hydrolysis products). This work also suggests that imines will be the main intermediates in the atmospheric oxidation of primary and secondary amines, including amine carbon capture solvents such as 2-aminoethanol (commonly known as monoethanolamine, or MEA), in a process that avoids the ozone-promoting conversion of (sic)NO to (sic)NO2 commonly encountered in peroxyl radical chemistry.

The effect of microscopic texture on the direct plasma surface passivation of Si solar cells

Textured silicon surfaces are widely used in manufacturing of solar cells due to increasing the light absorption probability and also the antireflection properties. However, these Si surfaces have a high density of surface defects that need to be passivated. In this study, the effect of the microscopic surface texture on the plasma surface passivation of solar cells is investigated. The movement of 105 H+ ions in the texture-modified plasma sheath is studied by Monte Carlo numerical simulation. The hydrogen ions are driven by the combined electric field of the plasma sheath and the textured surface. The ion dynamics is simulated, and the relative ion distribution over the textured substrate is presented. This distribution can be used to interpret the quality of the Si dangling bonds saturation and consequently, the direct plasma surface passivation.

The SEA-CALIPSO volcano imaging experiment at Montserrat : plans, campaigns at sea and on land, scientific results, and lessons learned

Since 1995 the eruption of the andesitic Soufrière Hills Volcano (SHV), Montserrat, has been studied in substantial detail. As an important contribution to this effort, the Seismic Experiment with Airgunsource-Caribbean Andesitic Lava Island Precision Seismo-geodetic Observatory (SEA-CALIPSO) experiment was devised to image the arc crust underlying Montserrat, and, if possible, the magma system at SHV using tomography and reflection seismology. Field operations were carried out in October–December 2007, with deployment of 238 seismometers on land supplementing seven volcano observatory stations, and with an array of 10 ocean-bottom seismometers deployed offshore. The RRS James Cook on NERC cruise JC19 towed a tuned airgun array plus a digital 48-channel streamer on encircling and radial tracks for 77 h about Montserrat during December 2007, firing 4414 airgun shots and yielding about 47 Gb of data. The main objecctives of the experiment were achieved. Preliminary analyses of these data published in 2010 generated images of heterogeneous high-velocity bodies representing the cores of volcanoes and subjacent intrusions, and shallow areas of low velocity on the flanks of the island that reflect volcaniclastic deposits and hydrothermal alteration. The resolution of this preliminary work did not extend beyond 5 km depth. An improved three-dimensional (3D) seismic velocity model was then obtained by inversion of 181 665 first-arrival travel times from a more-complete sampling of the dataset, yielding clear images to 7.5 km depth of a low-velocity volume that was interpreted as the magma chamber which feeds the current eruption, with an estimated volume 13 km3. Coupled thermal and seismic modelling revealed properties of the partly crystallized magma. Seismic reflection analyses aimed at imaging structures under southern Montserrat had limited success, and suggest subhorizontal layering interpreted as sills at a depth of between 6 and 19 km. Seismic reflection profiles collected offshore reveal deep fans of volcaniclastic debris and fault offsets, leading to new tectonic interpretations. This chapter presents the project goals and planning concepts, describes in detail the campaigns at sea and on land, summarizes the major results, and identifies the key lessons learned.

Pteridine-, thymidine-, choline- and imidazole-derived alkaloids from the Australian ascidian, Leptoclinides durus

Four new acylated pteridine alkaloids, duramidines A-D, two new acylated thymidine alkaloids, leptoclinidines A and B, two new 1-acylglyceryl-3-(O- carboxyhydroxymethylcholine) alkaloids, durabetaines A and B, three new 1,3-dimethyl-5-methylsulfanylimidazole alkaloids, leptoclinidamines D-F, and the known alkaloids leptoclinidamines B and C and 6-bromo-1H-indolo-3-yl-oxoacetic acid methyl ester were isolated from the Australian ascidian Leptoclinides durus. The duramidines are the first pteridine alkaloids, possessing a three carbon side chain esterified at C-1′ with a 4-hydroxy-2′- methoxycinnamic acid, and are either hydroxylated or sulfated at C-2′. The leptoclinidines are the first 3′-indole-3-carboxylic acid ester derivatives of thymidine to be reported in the literature. The durabetaines are the first glyceryl-3-(O-carboxyhydroxymethylcholine) alkaloids to be reported from an animal source and are also the only known derivatives from this class to be acylated with aromatic carboxylic acids. MS and NMR data analysis established the structures of the new compounds. All compounds were shown to be inactive when tested for cytotoxic activity against prostate (LNCaP) and breast (MDA-MB-231) cancer cell lines and antimicrobial activity against Pseudomonas aeruginosa and Staphylococcus aureus.

Entomopathogens for control of Hypsipyla spp

Published information on the incidence of pathogens in the field and laboratory infections of Hypsipyla spp. with entomopathogens is reviewed. In addition, some preliminary results of field collections from Ghana and Costa Rica are presented. Fungal pathogens from the Deuteromycetes have been isolated from both H. robusta Moore and H. grandella Zeller. Mermithid nematodes, Hexamermis spp., have been frequently isolated from larvae in the field and incidence of infection with these pathogens can reach significant levels. Microsporidia have been found in cadavers of larvae collected in the field but none have been identified so far. A number of pathogens of other Lepidoptera have been shown to be infectious to H. grandella , including Bacillus thuringiensis , Deuteromycete fungi and a nucleopolyhedrovirus (NPV) from Autographa californica . Hypsipyla spp. are difficult targets for microbial control, since the larvae are cryptic, occur at low density and occur sporadically. In addition, there is a low damage threshold, the plant is susceptible for a number of years and the susceptible part of the plant will rapidly outgrow any surface application. Key features of the biology of entomopathogens with relevance to the control of low density and cryptic pests are discussed. In the light of this experience, we discuss strategies to improve the possibilities of microbial control of this pest and suggest areas for research.

Silvicultural management of Hypsipyla species

Existing evidence for successful silvicultural control of Hypsipyla spp. is conflicting and to a large extent anecdotal. Levels of attack have been correlated with factors such as shade, planting density, species mixtures, site characteristics, etc. These factors have often been poorly defined and are usually interdependent. The actual mechanisms that determine whether or not Hypsipyla spp. adversely affects plants we define as host-finding, host suitability, host recovery and natural enemies. These mechanisms can be influenced by the silvicultural techniques applied to a stand. Success of silvicultural techniques can usually be attributed to more than one mechanism and it is difficult to assess which is most the important for minimising the impact of Hypsipyla as these analytical data are lacking. This highlights the need for further research on silvicultural methods for controlling Hypsipyla spp. However, several silvicultural techniques that are briefly described show promise for improving the performance of future plantations. Examples of silvicultural control are reviewed with reference to these mechanisms.

Melatonin synthesis : a possible indicator of intolerance to shiftwork

Hypothesis Melatonin synthesis, which is directly controlled by the central circadian pacemaker indicates the circadian phase better than rectal temperature. Methods: Thirty four men (16-32 years, 7 morning, 13 neither, 14 evening types) performed a constant routine (24-26-hr bedrest, < 30 lux, 18-20°C, hourly isocaloric diet). Salivary melatonin level was determined hourly and rectal temperature was continuously recorded. Results: The nadir of rectal temperature occurred 1.5 hr (P = 0.017), the onset of melatonin synthesis 3 hr earlier (P < 0.0001) in morning than in evening types. Morningness was not related to the quantitative but significantly to the temporal parameters, closer to those of melatonin than of rectal temperature. Conclusions: The melatonin onset is a more reliable indicator of the diurnal type than the nadir of rectal temperature. As morningness has been associated with intolerance to shiftwork, melatonin profiling provides a suitable basis for the establishment of directed preventive measures.

Hydrolysis of genotoxic methyl-substituted oxiranes : Experimental kinetic and semiempirical studies

The kinetics of acid-catalyzed hydrolysis of seven methylated aliphatic epoxides - R1R2C(O)CR3R4 (A: R1=R2=R3=R4=H; B: R1=R2=R3=H, R4=Me; C: R1=R2=H, R3=R4=Me; D: R1=R3=H, R2=R4=Me(trans); E: R1=R3=H, R2=R4=Me(cis); F: R1=R3=R4=Me, R2=H; G: R1=R2=R3=R4=Me) - has been studied at 36 ± 1.5°C. Compounds with two methyl groups at the same carbon atom of the oxirane ring exhibit highest rate constants (k(eff) in reciprocal molar concentration per second: 11.0 ± 1.3 for C, 10.7 ± 2.1 for F, and 8.7 ± 0.7 for G as opposed to 0.124 ± 0.003 for B, 0.305 ± 0.003 for D, and 0.635 ± 0.036 for E). Ethylene oxide (A) displays the lowest rate of hydrolysis (0.027 M-1 s-1). The results are consistent with literature data available for compounds A, B, and C. To model the reactivities we have employed quantum chemical calculations (MNDO, AM1, PM3, and MINDO/3) of the main reaction species. There is a correlation of the logarithm k(eff) with the total energy of epoxide ring opening. The best correlation coefficients (r) were obtained using the AM1 and MNDO methods (0.966 and 0.957, respectively). However, unlike MNDO, AM1 predicts approximately zero energy barriers for the oxirane ring opening of compounds B, C, E and G, which is not consistent with published kinetic data. Thus, the MNDO method provides a preferential means of modeling the acidic hydrolysis of the series of methylated oxiranes. The general ranking of mutagenicity in vitro, A &gt; B &gt; C, is in line with the concept that this sequence also gradually leaves the expoxide reactivity optimal for genotoxicity toward reactivities leading to higher biological detoxifications.

Influence of polymorphisms of the human glutathione transferases and cytochrome P450 2E1 enzyme on the metabolism and toxicity of ethylene oxide and acrylonitrile

A cohort of 59 persons with industrial handling of low levels of acrylonitrile is being studied as part of a medical surveillance programme. Previously, an extended haemoglobin adduct monitoring (N-(cyanoethyl)valine and N-(hydroxyethyl)-valine) was performed regarding the glutathione transferases hGSTM1 and hGSTT1 polymorphisms but no influence of hGSTM1 or hGSTT1 polymorphisms on specific adduct levels was found. A compilation of case reports of human accidental poisonings had pointed to significant individual differences in human acrylonitrile metabolism and toxicity. Therefore, a re-evaluation of the industrial cohort included known polymorphisms of the glutathione transferases hGSTM3 and hGSTP1 as well as of the cytochrome P450 CYP2E1. A detailed statistical analysis revealed that exposed carriers of the allelic variants of hGSTP1, hGSTP1*B/hGSTP1*C, characterized by a single nucleotide polymorphism at nucleotide 313 which results in a change from Ile to Val at codon 104, had higher levels of the acrylonitrile-specific haemoglobin adduct N-(cyanoethyl)valine compared to the carriers of the codon 113 alleles hGSTP1*A and hGSTP1*D. The single nucleotide polymorphism at codon 113 of hGSTP1 (hGSTP1*A/hGSTP1*B versus hGSTP1*C/hGSTP1*D) did not show an effect, and also no influence was seen on specific haemoglobin adduct levels of the polymorphisms of hGSTM3 or CYP2E1. The data, therefore, point to a possible influence of a human enzyme polymorphism of the GSTP1 gene at codon 104 on the detoxication of acrylonitrile which calls for experimental toxicological investigation. The study also confirmed the impact of GSTT1 polymorphism on background N-(hydroxyethyl)-valine adduct levels in haemoglobin which are caused by endogenous ethylene oxide.

The glycosphingolipid P1 is an ovarian cancer-associated carbohydrate antigen involved in migration

Background The level of plasma-derived naturally circulating anti-glycan antibodies (AGA) to P1 trisaccharide has previously been shown to significantly discriminate between ovarian cancer patients and healthy women. Here we aim to identify the Ig class that causes this discrimination, to identify on cancer cells the corresponding P1 antigen recognised by circulating anti-P1 antibodies and to shed light into the possible function of this glycosphingolipid. Method An independent Australian cohort was assessed for the presence of anti-P1 IgG and IgM class antibodies using suspension array. Monoclonal and human derived anti-glycan antibodies were verified using three independent glycan-based immunoassays and flow cytometry-based inhibition assay. The P1 antigen was detected by LC-MS/MS and flow cytometry. FACS-sorted cell lines were studied on the cellular migration by colorimetric assay and real-time measurement using xCELLigence system. Results Here we show in a second independent cohort (n=155) that the discrimination of cancer patients is mediated by the IgM class of anti-P1 antibodies (P=0.0002). The presence of corresponding antigen P1 and structurally related epitopes in fresh tissue specimens and cultured cancer cells is demonstrated. We further link the antibody and antigen (P1) by showing that human naturally circulating and affinity-purified anti-P1 IgM isolated from patients ascites can bind to naturally expressed P1 on the cell surface of ovarian cancer cells. Cell-sorted IGROV1 was used to obtain two study subpopulations (P1-high, 66.1%; and P1-low, 33.3%) and observed that cells expressing high P1-levels migrate significantly faster than those with low P1-levels. Conclusions This is the first report showing that P1 antigen, known to be expressed on erythrocytes only, is also present on ovarian cancer cells. This suggests that P1 is a novel tumour-associated carbohydrate antigen recognised by the immune system in patients and may have a role in cell migration. The clinical value of our data may be both diagnostic and prognostic; patients with low anti-P1 IgM antibodies present with a more aggressive phenotype and earlier relapse.

Human amnion epithelial cell transplantation abrogates lung fibrosis and augments repair

Rationale: Chronic lung disease characterized by loss of lung tissue,inflammation, and fibrosis represents a major global health burden. Cellular therapies that could restore pneumocytes and reduce inflammation and fibrosis would be a major advance in management. Objectives: To determine whether human amnion epithelial cells (hAECs), isolated from term placenta and having stem cell–like and antiinflammatory properties, could adopt an alveolar epithelial phenotype and repair a murine model of bleomycin-induced lung injury. Methods: Primary hAECs were cultured in small airway growth medium to determine whether the cells could adopt an alveolar epithelial phenotype. Undifferentiated primary hAECs were also injected parenterally into SCID mice after bleomycin-induced lung injury and analyzed for production of surfactant protein (SP)-A, SP-B, SP-C, and SP-D. Mouse lungs were also analyzed for inflammation and collagen deposition. Measurements and Main Results: hAECs grown in small airway growth medium developed an alveolar epithelial phenotype with lamellar body formation, production of SPs A–D, and SP-D secretion. Although hAECs injected into mice lacked SPs, hAECs recovered from mouse lungs 2 weeks posttransplantation produced SPs. hAECs remained engrafted over the 4-week test period. hAEC administration reduced inflammation in association with decreased monocyte chemoattractant protein-1, tumor necrosis factor-a, IL-1 and -6, and profibrotic transforming growth factor-b in mouse lungs. In addition,lung collagen content was significantly reduced by hAEC treatment as a possible consequence of increased degradation by matrix metalloproteinase-2 and down-regulation of the tissue inhibitors f matrix metalloproteinase-1 and 2. Conclusions: hAECs offer promise as a cellular therapy for alveolar restitution and to reduce lung inflammation and fibrosis.

Enhancing MRI of prostate cancer using PSMA-targeting iron oxide magnetic nanoparticles

Introduction Novel imaging techniques for prostate cancer (PCa) are required to improve staging and real-time assessment of therapeutic response. We performed preclinical evaluation of newly-developed, biocompatible magnetic nanoparticles (MNPs) conjugated with J591, an antibody specific for prostate specific membrane antigen (PSMA), to enhance magnetic resonance imaging (MRI) of PCa. PSMA is expressed on ∼90% of PCa, including those that are castrate-resistant, rendering it as a rational target for PCa imaging. Materials and Methods The specificity of J591 for PSMA was confirmed by flow cytometric analysis of several PCa cell lines of known PSMA status. MNPs were prepared, engineered to the appropriate size, labeled with DiR fluorophore, and their toxicity to a panel of PC cells was assessed by in vitro Alamar Blue assay. Immunohistochemistry, fluorescence microscopy and Prussian Blue staining (iron uptake) were used to evaluate PSMA specificity of J591-MNP conjugates. In vivo MRI studies (16.4T MRI system) were performed using live immunodeficient mice bearing orthotopic LNCaP xenografts and injected intravenously with J591-MNPs or MNPs alone. Results MNPs were non-toxic to PCa cells. J591-MNP conjugates showed no compromise in specificity of binding to PSMA+ cells and showed enhanced iron uptake compared with MNPs alone. In vivo, tumour targeting (significant MR image contrast) was evident in mice injected with J591-MNPs, but not MNPs alone. Resected tumours from targeted mice had an accumulation of MNPs, not seen in normal control prostate. Conclusions Application of PSMA-targeting MNPs into conventional MRI has potential to enhance PCa detection and localization in real-time, improving patient management.

Disability-adjusted life years (DALYs) for 291 diseases and injuries in 21 regions, 1990-2010: A systematic analysis for the Global Burden of Disease Study 2010

BACKGROUND Measuring disease and injury burden in populations requires a composite metric that captures both premature mortality and the prevalence and severity of ill-health. The 1990 Global Burden of Disease study proposed disability-adjusted life years (DALYs) to measure disease burden. No comprehensive update of disease burden worldwide incorporating a systematic reassessment of disease and injury-specific epidemiology has been done since the 1990 study. We aimed to calculate disease burden worldwide and for 21 regions for 1990, 2005, and 2010 with methods to enable meaningful comparisons over time. METHODS We calculated DALYs as the sum of years of life lost (YLLs) and years lived with disability (YLDs). DALYs were calculated for 291 causes, 20 age groups, both sexes, and for 187 countries, and aggregated to regional and global estimates of disease burden for three points in time with strictly comparable definitions and methods. YLLs were calculated from age-sex-country-time-specific estimates of mortality by cause, with death by standardised lost life expectancy at each age. YLDs were calculated as prevalence of 1160 disabling sequelae, by age, sex, and cause, and weighted by new disability weights for each health state. Neither YLLs nor YLDs were age-weighted or discounted. Uncertainty around cause-specific DALYs was calculated incorporating uncertainty in levels of all-cause mortality, cause-specific mortality, prevalence, and disability weights. FINDINGS Global DALYs remained stable from 1990 (2·503 billion) to 2010 (2·490 billion). Crude DALYs per 1000 decreased by 23% (472 per 1000 to 361 per 1000). An important shift has occurred in DALY composition with the contribution of deaths and disability among children (younger than 5 years of age) declining from 41% of global DALYs in 1990 to 25% in 2010. YLLs typically account for about half of disease burden in more developed regions (high-income Asia Pacific, western Europe, high-income North America, and Australasia), rising to over 80% of DALYs in sub-Saharan Africa. In 1990, 47% of DALYs worldwide were from communicable, maternal, neonatal, and nutritional disorders, 43% from non-communicable diseases, and 10% from injuries. By 2010, this had shifted to 35%, 54%, and 11%, respectively. Ischaemic heart disease was the leading cause of DALYs worldwide in 2010 (up from fourth rank in 1990, increasing by 29%), followed by lower respiratory infections (top rank in 1990; 44% decline in DALYs), stroke (fifth in 1990; 19% increase), diarrhoeal diseases (second in 1990; 51% decrease), and HIV/AIDS (33rd in 1990; 351% increase). Major depressive disorder increased from 15th to 11th rank (37% increase) and road injury from 12th to 10th rank (34% increase). Substantial heterogeneity exists in rankings of leading causes of disease burden among regions. INTERPRETATION Global disease burden has continued to shift away from communicable to non-communicable diseases and from premature death to years lived with disability. In sub-Saharan Africa, however, many communicable, maternal, neonatal, and nutritional disorders remain the dominant causes of disease burden. The rising burden from mental and behavioural disorders, musculoskeletal disorders, and diabetes will impose new challenges on health systems. Regional heterogeneity highlights the importance of understanding local burden of disease and setting goals and targets for the post-2015 agenda taking such patterns into account. Because of improved definitions, methods, and data, these results for 1990 and 2010 supersede all previously published Global Burden of Disease results.