Functional characterization of 8-oxoguanine DNA glycosylase of Trypanosoma cruzi

The oxidative lesion 8-oxoguanine (8-oxoG) is removed during base excision repair by the 8-oxoguanine DNA glycosylase 1 (Ogg1). This lesion can erroneously pair with adenine, and the excision of this damaged base by Ogg1 enables the insertion of a guanine and prevents DNA mutation. In this report, we identified and characterized Ogg1 from the protozoan parasite Trypanosoma cruzi (TcOgg1), the causative agent of Chagas disease. Like most living organisms, T. cruzi is susceptible to oxidative stress, hence DNA repair is essential for its survival and improvement of infection. We verified that the TcOGG1 gene encodes an 8-oxoG DNA glycosylase by complementing an Ogg1-defective Saccharomyces cerevisiae strain. Heterologous expression of TcOGG1 reestablished the mutation frequency of the yeast mutant ogg1-/- (CD138) to wild type levels. We also demonstrate that the overexpression of TcOGG1 increases T. cruzi sensitivity to hydrogen peroxide (H2O2). Analysis of DNA lesions using quantitative PCR suggests that the increased susceptibility to H2O2 of TcOGG1-overexpressor could be a consequence of uncoupled BER in abasic sites and/or strand breaks generated after TcOgg1 removes 8-oxoG, which are not rapidly repaired by the subsequent BER enzymes. This hypothesis is supported by the observation that TcOGG1-overexpressors have reduced levels of 8-oxoG both in the nucleus and in the parasite mitochondrion. The localization of TcOgg1 was examined in parasite transfected with a TcOgg1-GFP fusion, which confirmed that this enzyme is in both organelles. Taken together, our data indicate that T. cruzi has a functional Ogg1 ortholog that participates in nuclear and mitochondrial BER. © 2012 Furtado et al

Retrospective harm benefit analysis of pre-clinical animal research for six treatment interventions

The harm benefit analysis (HBA) is the cornerstone of animal research regulation and is considered to be a key ethical safeguard for animals. The HBA involves weighing the anticipated benefits of animal research against its predicted harms to animals but there are doubts about how objective and accountable this process is.i. To explore the harms to animals involved in pre-clinical animal studies and to assess these against the benefits for humans accruing from these studies; ii. To test the feasibility of conducting this type of retrospective HBA.Data on harms were systematically extracted from a sample of pre-clinical animal studies whose clinical relevance had already been investigated by comparing systematic reviews of the animal studies with systematic reviews of human studies for the same interventions (antifibrinolytics for haemorrhage, bisphosphonates for osteoporosis, corticosteroids for brain injury, Tirilazad for stroke, antenatal corticosteroids for neonatal respiratory distress and thrombolytics for stroke). Clinical relevance was also explored in terms of current clinical practice. Harms were categorised for severity using an expert panel. The quality of the research and its impact were considered. Bateson's Cube was used to conduct the HBA.The most common assessment of animal harms by the expert panel was 'severe'. Reported use of analgesia was rare and some animals (including most neonates) endured significant procedures with no, or only light, anaesthesia reported. Some animals suffered iatrogenic harms. Many were kept alive for long periods post-experimentally but only 1% of studies reported post-operative care. A third of studies reported that some animals died prior to endpoints. All the studies were of poor quality. Having weighed the actual harms to animals against the actual clinical benefits accruing from these studies, and taking into account the quality of the research and its impact, less than 7% of the studies were permissible according to Bateson's Cube: only the moderate bisphosphonate studies appeared to minimise harms to animals whilst being associated with benefit for humans.This is the first time the accountability of the HBA has been systematically explored across a range of pre-clinical animal studies. The regulatory systems in place when these studies were conducted failed to safeguard animals from severe suffering or to ensure that only beneficial, scientifically rigorous research was conducted. Our findings indicate a pressing need to: i. review regulations, particularly those that permit animals to suffer severe harms; ii. reform the processes of prospectively assessing pre-clinical animal studies to make them fit for purpose; and iii. systematically evaluate the benefits of pre-clinical animal research to permit a more realistic assessment of its likely future benefits

Study of Bc+B_c^+ decays to the K+K−π+K^+K^-\pi^+ final state and evidence for the decay Bc+→χc0π+B_c^+\to\chi_{c0}\pi^+

A study of $B_c^+\to K^+K^-\pi^+$ decays is performed for the first time using data corresponding to an integrated luminosity of 3.0 $\mathrm{fb}^{-1}$ collected by the LHCb experiment in $pp$ collisions at centre-of-mass energies of $7$ and $8$ TeV. Evidence for the decay $B_c^+\to\chi_{c0}(\to K^+K^-)\pi^+$ is reported with a significance of 4.0 standard deviations, resulting in the measurement of $\frac{\sigma(B_c^+)}{\sigma(B^+)}\times\mathcal{B}(B_c^+\to\chi_{c0}\pi^+)$ to be $(9.8^{+3.4}_{-3.0}(\mathrm{stat})\pm 0.8(\mathrm{syst}))\times 10^{-6}$. Here $\mathcal{B}$ denotes a branching fraction while $\sigma(B_c^+)$ and $\sigma(B^+)$ are the production cross-sections for $B_c^+$ and $B^+$ mesons. An indication of $\bar b c$ weak annihilation is found for the region $m(K^-\pi^+)<1.834\mathrm{\,Ge\kern -0.1em V\!/}c^2$, with a significance of 2.4 standard deviations.Comment: All figures and tables, along with any supplementary material and additional information, are available at https://lhcbproject.web.cern.ch/lhcbproject/Publications/LHCbProjectPublic/LHCb-PAPER-2016-022.html, link to supplemental material inserted in the reference

Performance of the CMS Cathode Strip Chambers with Cosmic Rays

The Cathode Strip Chambers (CSCs) constitute the primary muon tracking device in the CMS endcaps. Their performance has been evaluated using data taken during a cosmic ray run in fall 2008. Measured noise levels are low, with the number of noisy channels well below 1%. Coordinate resolution was measured for all types of chambers, and fall in the range 47 microns to 243 microns. The efficiencies for local charged track triggers, for hit and for segments reconstruction were measured, and are above 99%. The timing resolution per layer is approximately 5 ns

(De)Constructing a Natural and Flavorful Supersymmetric Standard Model

Using the framework of deconstruction, we construct simple, weakly-coupled supersymmetric models that explain the Standard Model flavor hierarchy and produce a flavorful soft spectrum compatible with precision limits. Electroweak symmetry breaking is fully natural; the mu-term is dynamically generated with no B mu-problem and the Higgs mass is easily raised above LEP limits without reliance on large radiative corrections. These models possess the distinctive spectrum of superpartners characteristic of "effective supersymmetry": the third generation superpartners tend to be light, while the rest of the scalars are heavy.Comment: 36 pages, 4 figures ; v2: references added, expanded discussion of FCNC

Animal welfare in studies on murine tuberculosis : assessing progress over a 12-year period and the need for further improvement

There is growing concern over the welfare of animals used in research, in particular when these animals develop pathology. The present study aims to identify the main sources of animal distress and to assess the possible implementation of refinement measures in experimental infection research, using mouse models of tuberculosis (TB) as a case study. This choice is based on the historical relevance of mouse studies in understanding the disease and the present and long-standing impact of TB on a global scale. Literature published between 1997 and 2009 was analysed, focusing on the welfare impact on the animals used and the implementation of refinement measures to reduce this impact. In this 12-year period, we observed a rise in reports of ethical approval of experiments. The proportion of studies classified into the most severe category did however not change significantly over the studied period. Information on important research parameters, such as method for euthanasia or sex of the animals, were absent in a substantial number of papers. Overall, this study shows that progress has been made in the application of humane endpoints in TB research, but that a considerable potential for improvement remains.Nuno H. Franco is funded by Fundação para a Ciência e Tecnologia (SFRH/BD/38337/2007). This work is funded by FEDER funds through the Operational Competitiveness Programme - COMPETE and by national funds through FCT - Fundação para a Ciência e Tecnologia under the project FCOMP-01-0124-FEDER-022718 (PEst-C/SAU/LA0002/2011

Changes in habitat associations during range expansion: disentangling the effects of climate and residence time

The distributions of many species are not at equilibrium with their environment. This includes spreading non-native species and species undergoing range shifts in response to climate change. The habitat associations of these species may change during range expansion as less favourable climatic conditions at expanding range margins may constrain species to use only the most favourable habitats, violating the species distribution model assumption of stationarity. Alternatively, changes in habitat associations could result from density-dependent habitat selection; at range margins, population densities are initially low so species can exhibit density-independent selection of the most favourable habitats, while in the range core, where population densities are higher, species spread into less favourable habitat. We investigate if the habitat preferences of the non-native common waxbill Estrilda astrild changed as they spread in three directions (north, east and south-east) in the Iberian Peninsula. There are different degrees of climatic suitability and colonization speed across range expansion axes, allowing us to separate the effects of climate from residence time. In contrast to previous studies we find a stronger effect of residence time than climate in influencing the prevalence of common waxbills. As well as a strong additive effect of residence time, there were some changes in habitat associations, which were consistent with density-dependent habitat selection. The combination of broader habitat associations and higher prevalence in areas that have been colonised for longer means that species distribution models constructed early in the invasion process are likely to underestimate species’ potential distribution

A prediction rule to stratify mortality risk of patients with pulmonary tuberculosis

Tuberculosis imposes high human and economic tolls, including in Europe. This study was conducted to develop a severity assessment tool for stratifying mortality risk in pulmonary tuberculosis (PTB) patients. A derivation cohort of 681 PTB cases was retrospectively reviewed to generate a model based on multiple logistic regression analysis of prognostic variables with 6-month mortality as the outcome measure. A clinical scoring system was developed and tested against a validation cohort of 103 patients. Five risk features were selected for the prediction model: hypoxemic respiratory failure (OR 4.7, 95% CI 2.8-7.9), age >= 50 years (OR 2.9, 95% CI 1.7-4.8), bilateral lung involvement (OR 2.5, 95% CI 1.44.4), >= 1 significant comorbidity-HIV infection, diabetes mellitus, liver failure or cirrhosis, congestive heart failure and chronic respiratory disease-(OR 2.3, 95% CI 1.3-3.8), and hemoglobin = 6) mortality risk. The mortality associated with each group was 2.9%, 22.9% and 53.9%, respectively. The model performed equally well in the validation cohort. We provide a new, easy-to-use clinical scoring system to identify PTB patients with high-mortality risk in settings with good healthcare access, helping clinicians to decide which patients are in need of closer medical care during treatment.This work was supported by Fundacao Amelia de Mello/Jose de Mello Saude and Sociedade Portuguesa de Pneumologia (SPP). This work was developed under the scope of the project NORTE-01-0145-FEDER-000013, supported by the Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER). NSO is a FCT (Fundacao para a Ciencia e Tecnologia) investigator. MS is an Associate FCT Investigator. The fundershad no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Expression variability of co-regulated genes differentiates Saccharomyces cerevisiae strains

Background: Saccharomyces cerevisiae (Baker’s yeast) is found in diverse ecological niches and is characterized by high adaptive potential under challenging environments. In spite of recent advances on the study of yeast genome diversity, little is known about the underlying gene expression plasticity. In order to shed new light onto this biological question, we have compared transcriptome profiles of five environmental isolates, clinical and laboratorial strains at different time points of fermentation in synthetic must medium, during exponential and stationary growth phases. Results: Our data unveiled diversity in both intensity and timing of gene expression. Genes involved in glucose metabolism and in the stress response elicited during fermentation were among the most variable. This gene expression diversity increased at the onset of stationary phase (diauxic shift). Environmental isolates showed lower average transcript abundance of genes involved in the stress response, assimilation of nitrogen and vitamins, and sulphur metabolism, than other strains. Nitrogen metabolism genes showed significant variation in expression among the environmental isolates. Conclusions: Wild type yeast strains respond differentially to the stress imposed by nutrient depletion, ethanol accumulation and cell density increase, during fermentation of glucose in synthetic must medium. Our results support previous data showing that gene expression variability is a source of phenotypic diversity among closely related organisms.Fundação para a Ciência e TecnologiaThe authors wish to thank Adega Cooperativa da Bairrada, Cantanhede, Portugal, for providing the commercial strains

A membrane-inserted structural model of the yeast mitofusin Fzo1

Mitofusins are large transmembrane GTPases of the dynamin-related protein family, and are required for the tethering and fusion of mitochondrial outer membranes. Their full-length structures remain unknown, which is a limiting factor in the study of outer membrane fusion. We investigated the structure and dynamics of the yeast mitofusin Fzo1 through a hybrid computational and experimental approach, combining molecular modelling and all-atom molecular dynamics simulations in a lipid bilayer with site-directed mutagenesis and in vivo functional assays. The predicted architecture of Fzo1 improves upon the current domain annotation, with a precise description of the helical spans linked by flexible hinges, which are likely of functional significance. In vivo site-directed mutagenesis validates salient aspects of this model, notably, the long-distance contacts and residues participating in hinges. GDP is predicted to interact with Fzo1 through the G1 and G4 motifs of the GTPase domain. The model reveals structural determinants critical for protein function, including regions that may be involved in GTPase domain-dependent rearrangements