Using size-selected gold clusters on graphene oxide films to aid cryo-transmission electron tomography alignment

A three-dimensional reconstruction of a nano-scale aqueous object can be achieved by taking a series of transmission electron micrographs tilted at different angles in vitreous ice: cryo-Transmission Electron Tomography. Presented here is a novel method of fine alignment for the tilt series. Size-selected gold clusters of ~2.7 nm (Au(561 ± 14)), ~3.2 nm (Au(923 ± 22)), and ~4.3 nm (Au(2057 ± 45)) in diameter were deposited onto separate graphene oxide films overlaying holes on amorphous carbon grids. After plunge freezing and subsequent transfer to cryo-Transmission Electron Tomography, the resulting tomograms have excellent (de-)focus and alignment properties during automatic acquisition. Fine alignment is accurate when the evenly distributed 3.2 nm gold particles are used as fiducial markers, demonstrated with a reconstruction of a tobacco mosaic virus. Using a graphene oxide film means the fiducial markers are not interfering with the ice bound sample and that automated collection is consistent. The use of pre-deposited size-selected clusters means there is no aggregation and a user defined concentration. The size-selected clusters are mono-dispersed and can be produced in a wide size range including 2–5 nm in diameter. The use of size-selected clusters on a graphene oxide films represents a significant technical advance for 3D cryo-electron microscopy

Statement on the dossier for a derogation request for the US authorities concerning cold-treated strawberry plants intended for planting.

Following a request from the European Commission, the EFSA Panel on Plant Health examined the report “Evaluation of Strawberry Nursery Plant Cold Treatments on Survival of the Whitefly, Bemisia tabaci”, submitted to the European Commission by the United States Department of Agriculture (USDA). The report serves as technical justification for a derogation from requirements listed in Annex IV, Part A, Section 1, point 46 of the Council Directive 2000/29/EC. The Panel concluded that due to shortcomings in the report itself and in the experimental design of the study, this report does not demonstrate the effectiveness of the cold treatment proposed for the elimination of B. tabaci from strawberry plant consignments prepared for shipment to the EU. In addition, the Panel conducted a preliminary review of the available literature on viruses of strawberry and concluded that no B. tabaci‐transmitted viruses are currently known to infect strawberry and that no viruses of Fragaria listed in European Council Directive 2000/29/EC are known to be transmitted by B. tabaci. However, adult whiteflies of B. tabaci on strawberry consignments can carry plant viruses irrespective of whether strawberry is a host plant for these viruses and thus represent a pathway for the introduction of non‐European viruses. There is also some uncertainty concerning the extent to which known or unknown B. tabaci‐transmitted viruses could infect strawberry. Irrespective of the presence of B. tabaci, strawberry transplants from the US may represent a pathway for the introduction of any non‐European virus (and other potentially harmful organisms) of Fragaria

Scientific Opinion on the assessment of the risk of solanaceous pospiviroids for the EU territory and the identification and evaluation of risk management options.

Following a request from the EU Commission, the EFSA PLH Panel conducted a risk assessment for the EU territory of pospiviroids affecting solanaceous crops, identified and evaluated risk reduction options and evaluated the EU provisional emergency measures targeting Potato spindle tuber viroid (PSTVd). The risk assessment included PSTVd, Citrus exocortis viroid, Columnea latent viroid, Mexican papita viroid, Tomato apical stunt viroid, Tomato chlorotic dwarf viroid, Tomato planta macho viroid, Chrysanthemum stunt viroid and Pepper chat fruit viroid. Four entry pathways were identified, three involving plant propagation material, with moderate probability of entry, and one involving plant products for human consumption, with low probability of entry. The probability of establishment was considered very high. Spread was considered likely within a crop and moderately likely between crop species, with exception of spread to potato, rated as unlikely. The probability of long distance spread within vegetatively propagated crops was estimated as likely/very likely. The direct consequences were expected to be major in potato and tomato, moderate in pepper, minimal/minor in other vegetables and minimal in ornamentals. Main risk assessment uncertainties derive from limited knowledge on pospiviroids other than PSTVd, although all pospiviroids are expected to have similar biological properties. Management options to reduce risk of entry, spread and consequences were identified and evaluated. No management options can prevent establishment. Examples of successful PSTVd eradication are linked to timely and strict implementation of measures. Uncertainty exists on the effectiveness of risk reduction strategies targeting only one pathway. The EU provisional emergency measures appeared to have significantly reduced PSTVd incidence in Solanum jasminoides and Brugmansia sp., even though eradication from the EU is so far incomplete. The low PSTVd incidence in food crops did not permit to conclude whether the reduction in PSTVd prevalence in ornamentals led to a reduction in outbreaks in food crops

Statement on a study proposal prepared by the UC to support a future derogation request from the EU import requirements for wood packaging material originating in the US and used to pack and transport military ammunition. Scientific Opinion.

Following a request from the European Commission, the EFSA Panel on Plant Health was asked to deliver a statement on a study proposal prepared by the US to support a future derogation request from the Community import requirements for wood packaging material (WPM) originating in the US and used to pack and transport military ammunition. The Panel reviewed the submitted study proposal and based its conclusions on the shortcomings identified in the study proposal. It was concluded that, due to these shortcomings, the evaluation of the methodological approach described in this study proposal with regard to: 1.the determination of the likelihood that live pinewood nematode (Bursaphelenchus xylophilus) and other harmful organisms relevant for the Community are present in the WPM used by the US Department of Defense to pack and transport military ammunition (Pest survey component), 2. the assessment of the likelihood that harmful organisms may be introduced into the Community through the WPM pathway (Pathway Risk Analysis component) cannot be conducted

Scientific Opinion on a technical file submitted by the US Authorities to support a request to list a new opinion among the EU import requirements for wood of Agrilus planipennis host plants.

This document presents the scientific opinion of the Panel on Plant Health on the technical file submitted by the US Authorities to support a request to list a new option among the EU import requirements for wood of Agrilus planipennis host plants. The option under consideration is a heat treatment at 60 °C for 60 min to liminate possible infestations of the wood by the emerald ash borer (EAB). The experiments leading the US Authorities to propose this option are presented in a scientific peer reviewed publication, Myers et al. (2009). The analysis of the aggregated data published by Myers et al. (2009) and based on a Probit regression model showed that the proposed treatment cannot guarantee a control level of 99 % or higher. The analysis of the individual data either from the original measurements or from a corrected dataset, using a Probit regression model, showed that it is likely to observe one live EAB out of an infestation of 100 after the proposed heat treatment of 60 °C /60 min. To ensure a control level of 99 % the temperature of the heat treatment of 60 min should be higher than 70 °C. Results obtained with a Poisson log linear model based on individual data showed that the estimated probability that one insect or more per m2 survive the proposed heat treatment was higher than 0.6 and that there is a 0.1 probability that three insects or more per m2 survive the proposed heat treatment. Based on these results, the Panel concludes that A. planipennis is likely to survive the proposed heat treatment of 60 °C/60 min with a low uncertainty, and that the alternative option proposed in the technical file submitted by the US Authorities for wood does not guarantee the wood to be free of A. planipennis

Chromium inhibition and size-selected Au nanocluster catalysis for the solution growth of low-density ZnO nanowires

he wet chemical synthesis of nanostructures has many crucial advantages over high-temperature methods, including simplicity, low-cost, and deposition on almost arbitrary substrates. Nevertheless, the density-controlled solution growth of nanowires still remains a challenge, especially at the low densities (e.g. 1 to 10 nanowires/100\u2009\u3bcm2) required, as an example, for intracellular analyses. Here, we demonstrate the solution-growth of ZnO nanowires using a thin chromium film as a nucleation inhibitor and Au size-selected nanoclusters (SSNCs) as catalytic particles for which the density and, in contrast with previous reports, size can be accurately controlled. Our results also provide evidence that the enhanced ZnO hetero-nucleation is dominated by Au SSNCs catalysis rather than by layer adaptation. The proposed approach only uses low temperatures ( 6470\u2009\ub0C) and is therefore suitable for any substrate, including printed circuit boards (PCBs) and the plastic substrates which are routinely used for cell cultures. As a proof-of-concept we report the density-controlled synthesis of ZnO nanowires on flexible PCBs, thus opening the way to assembling compact intracellular-analysis systems, including nanowires, electronics, and microfluidics, on a single substrate

Using electron paramagnetic resonance to map N@C60 during high throughput processing

The endohedral fullerene molecule, N@C60, is a candidate for molecular spin qubits (quantum bits) and spin probes owing to its exceptional electron spin properties. Advancements in the processing of N@C60 are key to obtaining samples of high purity on a reasonable timescale. We investigate enrichment by high throughput processing (flow rate of 18 L h−1 and operating pressure of 1.5–2 MPa) using high performance liquid chromatography (HPLC) as a means of scaling N@C60 production. We use detection by electron paramagnetic resonance (EPR) spectroscopy to map N@C60 during processing, and through the reconstruction of the peak position in the chromatogram, we are able to determine the retention time and relative purity of N@C60 without the need for its isolation. Based on this, we establish a procedure for time-efficient, high throughput processing to isolate N@C60 in high purity

Potential efficacy of mitochondrial genes for animal DNA barcoding: a case study using eutherian mammals

<p>Abstract</p> <p>Background</p> <p>A well-informed choice of genetic locus is central to the efficacy of DNA barcoding. Current DNA barcoding in animals involves the use of the 5' half of the mitochondrial cytochrome oxidase 1 gene (<it>CO1</it>) to diagnose and delimit species. However, there is no compelling <it>a priori </it>reason for the exclusive focus on this region, and it has been shown that it performs poorly for certain animal groups. To explore alternative mitochondrial barcoding regions, we compared the efficacy of the universal <it>CO1 </it>barcoding region with the other mitochondrial protein-coding genes in eutherian mammals. Four criteria were used for this comparison: the number of recovered species, sequence variability within and between species, resolution to taxonomic levels above that of species, and the degree of mutational saturation.</p> <p>Results</p> <p>Based on 1,179 mitochondrial genomes of eutherians, we found that the universal <it>CO1 </it>barcoding region is a good representative of mitochondrial genes as a whole because the high species-recovery rate (> 90%) was similar to that of other mitochondrial genes, and there were no significant differences in intra- or interspecific variability among genes. However, an overlap between intra- and interspecific variability was still problematic for all mitochondrial genes. Our results also demonstrated that any choice of mitochondrial gene for DNA barcoding failed to offer significant resolution at higher taxonomic levels.</p> <p>Conclusions</p> <p>We suggest that the <it>CO1 </it>barcoding region, the universal DNA barcode, is preferred among the mitochondrial protein-coding genes as a molecular diagnostic at least for eutherian species identification. Nevertheless, DNA barcoding with this marker may still be problematic for certain eutherian taxa and our approach can be used to test potential barcoding loci for such groups.</p

Genome Wide Expression Profiling Reveals Suppression of Host Defence Responses during Colonisation by Neisseria meningitides but not N. lactamica

Both Neisseria meningitidis and the closely related bacterium Neisseria lactamica colonise human nasopharyngeal mucosal surface, but only N. meningitidis invades the bloodstream to cause potentially life-threatening meningitis and septicaemia. We have hypothesised that the two neisserial species differentially modulate host respiratory epithelial cell gene expression reflecting their disease potential. Confluent monolayers of 16HBE14 human bronchial epithelial cells were exposed to live and/or dead N. meningitidis (including capsule and pili mutants) and N. lactamica, and their transcriptomes were compared using whole genome microarrays. Changes in expression of selected genes were subsequently validated using Q-RT-PCR and ELISAs. Live N. meningitidis and N. lactamica induced genes involved in host energy production processes suggesting that both bacterial species utilise host resources. N. meningitidis infection was associated with down-regulation of host defence genes. N. lactamica, relative to N. meningitidis, initiates up-regulation of proinflammatory genes. Bacterial secreted proteins alone induced some of the changes observed. The results suggest N. meningitidis and N. lactamica differentially regulate host respiratory epithelial cell gene expression through colonisation and/or protein secretion, and that this may contribute to subsequent clinical outcomes associated with these bacteria