On the extent and role of the small proteome in the parasitic eukaryote Trypanosoma brucei

Background: Although technical advances in genomics and proteomics research have yielded a better understanding of the coding capacity of a genome, one major challenge remaining is the identification of all expressed proteins, especially those less than 100 amino acids in length. Such information can be particularly relevant to human pathogens, such as Trypanosoma brucei, the causative agent of African trypanosomiasis, since it will provide further insight into the parasite biology and life cycle. Results: Starting with 993 T. brucei transcripts, previously shown by RNA-Sequencing not to coincide with annotated coding sequences (CDS), homology searches revealed that 173 predicted short open reading frames in these transcripts are conserved across kinetoplastids with 13 also conserved in representative eukaryotes. Mining mass spectrometry data sets revealed 42 transcripts encoding at least one matching peptide. RNAi-induced down-regulation of these 42 transcripts revealed seven to be essential in insect-form trypanosomes with two also required for the bloodstream life cycle stage. To validate the specificity of the RNAi results, each lethal phenotype was rescued by co-expressing an RNAi-resistant construct of each corresponding CDS. These previously non-annotated essential small proteins localized to a variety of cell compartments, including the cell surface, mitochondria, nucleus and cytoplasm, inferring the diverse biological roles they are likely to play in T. brucei. We also provide evidence that one of these small proteins is required for replicating the kinetoplast (mitochondrial) DNA. Conclusions: Our studies highlight the presence and significance of small proteins in a protist and expose potential new targets to block the survival of trypanosomes in the insect vector and/or the mammalian host

Transient Radiative and Conductive Heat Transfer in Ceramic Materials Subjected to Laser Heating

In this work the problem of transient energy transport in a conductive, emitting, and scattering medium is addressed. A new approach for describing the transient response of a participating one-dimensional layer subjected to a finite laser pulse is presented. The model considers heat transport in one dimension by conduction and radiation, where the radiative field is described by the two-flux approximation. The material is assumed to be gray, homogeneous, and isotropic, with physical and optical properties dependent on temperature. Numerical results have been obtained on a layer of low optical thickness using the thermophysical properties of zirconia as a typical materia

The 2017 reversal of the Beaufort Gyre: Can dynamic thickening of a seasonal ice cover during a reversal limit summer ice melt in the Beaufort Sea?

During winter 2017 the semi‐permanent Beaufort High collapsed and the anticyclonic Beaufort Gyre reversed. The reversal drove eastward ice motion through the Western Arctic, causing sea ice to converge against Banks Island, and halted the circulation of multiyear sea ice via the gyre, preventing its replenishment in the Beaufort Sea. Prior to the reversal, an anomalously thin seasonal ice cover had formed in the Beaufort following ice‐free conditions during September 2016. With the onset of the reversal in January 2017, convergence drove uncharacteristic dynamic thickening during winter. By the end of March, despite seasonal ice comprising 97% of the ice cover, the reversal created the thickest, roughest and most voluminous regional ice cover of the CryoSat‐2 record. Within the Beaufort Sea, previous work has shown that winter ice export can precondition the region for increased summer ice melt, but that a short reversal during April 2013 contributed to a reduction in summer ice loss. Hence the deformed ice cover at the end of winter 2017 could be expected to limit summer melt. In spite of this, the Beaufort ice cover fell to its fourth lowest September area as the gyre re‐established during April and divergent ice drift broke up the pack, negating the reversal's earlier preconditioning. Our work highlights that dynamic winter thickening of a regional sea ice cover, for instance during a gyre reversal, offers the potential to limit summer ice loss, but that dynamic forcing during spring dictates whether this conditioning carries through to the melt season

Training and assessment of non-technical skills in Norwegian helicopter emergency services: a cross-sectional and longitudinal study

Background Deficient non-technical skills (NTS) among providers of critical care in helicopter emergency medical services (HEMS) is a threat to patient and operational safety. Skills can be improved through simulation-based training and assessment. A previous study indicated that physicians underwent less frequent training compared to pilots and HEMS crew members (HCM) and that all professional groups in Norwegian HEMS received limited training in how to cope with fatigue. Since then, training initiatives and a fatigue risk management project has been initiated. Our study aimed to explore if the frequency of simulation-based training and assessment of NTS in Norwegian HEMS has changed since 2011 following these measures. Methods A cross-sectional web-based survey from October through December 2016, of physicians, HCM and pilots from all civilian Norwegian HEMS-bases reporting the overall extent of simulation-based training and assessment of NTS. Results Of 214 invited, 109 responses were eligible for analysis. The frequency of simulation-based training and assessment of NTS has increased significantly for all professional groups in Norwegian HEMS, most prominently for the physicians. For all groups, the frequency of assessment is generally lower than the frequency of training. Conclusions Physicians in Norwegian HEMS seem to have adjusted to the NTS training culture of the other crew member groups. This might be a consequence of improved NTS training programs. The use of behavioural marker systems systematically in HEMS should be emphasized.publishedVersio

Retention and loss of RNA interference pathways in trypanosomatid protozoans

RNA interference (RNAi) pathways are widespread in metaozoans but the genes required show variable occurrence or activity in eukaryotic microbes, including many pathogens. While some Leishmania lack RNAi activity and Argonaute or Dicer genes, we show that Leishmania braziliensis and other species within the Leishmania subgenus Viannia elaborate active RNAi machinery. Strong attenuation of expression from a variety of reporter and endogenous genes was seen. As expected, RNAi knockdowns of the sole Argonaute gene implicated this protein in RNAi. The potential for functional genetics was established by testing RNAi knockdown lines lacking the paraflagellar rod, a key component of the parasite flagellum. This sets the stage for the systematic manipulation of gene expression through RNAi in these predominantly diploid asexual organisms, and may also allow selective RNAi-based chemotherapy. Functional evolutionary surveys of RNAi genes established that RNAi activity was lost after the separation of the Leishmania subgenus Viannia from the remaining Leishmania species, a divergence associated with profound changes in the parasite infectious cycle and virulence. The genus Leishmania therefore offers an accessible system for testing hypothesis about forces that may select for the loss of RNAi during evolution, such as invasion by viruses, changes in genome plasticity mediated by transposable elements and gene amplification (including those mediating drug resistance), and/or alterations in parasite virulence

Cytochrome oxidase subunit VI of Trypanosoma brucei is imported without a cleaved presequence and is developmentally regulated at both RNA and protein levels

Mitochondrial respiration in the African trypanosome undergoes dramatic developmental stage regulation. This requires co-ordinated control of components encoded by both the nuclear genome and the kinetoplast, the unusual mitochondrial genome of these parasites. As a model for understanding the co-ordination of these genomes, we have examined the regulation and mitochondrial import of a nuclear-encoded component of the cytochrome oxidase complex, cytochrome oxidase subunit VI (COXVI). By generating transgenic trypanosomes expressing intact or mutant forms of this protein, we demonstrate that COXVI is not imported using a conventional cleaved presequence and show that sequences at the N-terminus of the protein are necessary for correct mitochondrial sorting. Analyses of endogenous and transgenic COXVI mRNA and protein expression in parasites undergoing developmental stage differentiation demonstrates a temporal order of control involving regulation in the abundance of, first, mRNA and then protein. This represents the first dissection of the regulation and import of a nuclear-encoded protein into the cytochrome oxidase complex in these organisms, which were among the earliest eukaryotes to possess a mitochondrion

Workgroup Report: Incorporating In Vitro Alternative Methods for Developmental Neurotoxicity into International Hazard and Risk Assessment Strategies

This is the report of the first workshop on Incorporating In Vitro Alternative Methods for Developmental Neurotoxicity (DNT) Testing into International Hazard and Risk Assessment Strategies, held in Ispra, Italy, on 19–21 April 2005. The workshop was hosted by the European Centre for the Validation of Alternative Methods (ECVAM) and jointly organized by ECVAM, the European Chemical Industry Council, and the Johns Hopkins University Center for Alternatives to Animal Testing. The primary aim of the workshop was to identify and catalog potential methods that could be used to assess how data from in vitro alternative methods could help to predict and identify DNT hazards. Working groups focused on two different aspects: a) details on the science available in the field of DNT, including discussions on the models available to capture the critical DNT mechanisms and processes, and b) policy and strategy aspects to assess the integration of alternative methods in a regulatory framework. This report summarizes these discussions and details the recommendations and priorities for future work

ESPRESSO: The next European exoplanet hunter

The acronym ESPRESSO stems for Echelle SPectrograph for Rocky Exoplanets and Stable Spectroscopic Observations; this instrument will be the next VLT high resolution spectrograph. The spectrograph will be installed at the Combined-Coud\'e Laboratory of the VLT and linked to the four 8.2 m Unit Telescopes (UT) through four optical Coud\'e trains. ESPRESSO will combine efficiency and extreme spectroscopic precision. ESPRESSO is foreseen to achieve a gain of two magnitudes with respect to its predecessor HARPS, and to improve the instrumental radial-velocity precision to reach the 10 cm/s level. It can be operated either with a single UT or with up to four UTs, enabling an additional gain in the latter mode. The incoherent combination of four telescopes and the extreme precision requirements called for many innovative design solutions while ensuring the technical heritage of the successful HARPS experience. ESPRESSO will allow to explore new frontiers in most domains of astrophysics that require precision and sensitivity. The main scientific drivers are the search and characterization of rocky exoplanets in the habitable zone of quiet, nearby G to M-dwarfs and the analysis of the variability of fundamental physical constants. The project passed the final design review in May 2013 and entered the manufacturing phase. ESPRESSO will be installed at the Paranal Observatory in 2016 and its operation is planned to start by the end of the same year.Comment: 12 pages, figures included, accepted for publication in Astron. Nach