Abstract
Inspection of ageing, reinforced concrete structures is a world-wide challenge. Existing evaluation techniques in civil and structural engineering have limited penetration depth and do not ...allow to precisely ascertain the configuration of reinforcement within large concrete objects. The big challenge for critical infrastructure (bridges, dams, dry docks, nuclear bioshields etc.) is understanding the internal condition of the concrete and steel, not just the location of the reinforcement. Muon scattering tomography is a non-destructive and non-invasive technique which shows great promise for high-depth 3D concrete imaging. Here a method is presented to locate reinforcement meshes placed in a large-scale concrete object. A reinforcement mesh was simulated as two layers of 2 m long bars, forming a mesh. Two layers of the mesh were placed at several distances from each other inside a large concrete block. Previously, we have shown that using our autocorrelation technique for single meshes inside the concrete and using only one week worth of data taking, bars with a diameter of 7 mm and larger, could easily be detected for a 10 cm mesh spacing. The signal for 6 mm diameter bar exceeds the background and becomes very clear after two weeks of data taking. Here we show that we can detect the vertical positions of two mesh layers inside the concrete. This is a very important result for non-destructive evaluation of civil structures.
Influenza virus-infected cells vary widely in their expression of viral genes and only occasionally activate innate immunity. Here, we develop a new method to assess how the genetic variation in ...viral populations contributes to this heterogeneity. We do this by determining the transcriptome and full-length sequences of all viral genes in single cells infected with a nominally "pure" stock of influenza virus. Most cells are infected by virions with defects, some of which increase the frequency of innate-immune activation. These immunostimulatory defects are diverse and include mutations that perturb the function of the viral polymerase protein PB1, large internal deletions in viral genes, and failure to express the virus's interferon antagonist NS1. However, immune activation remains stochastic in cells infected by virions with these defects and occasionally is triggered even by virions that express unmutated copies of all genes. Our work shows that the diverse spectrum of defects in influenza virus populations contributes to-but does not completely explain-the heterogeneity in viral gene expression and immune activation in single infected cells.
Because influenza virus has a high mutation rate, many cells are infected by mutated virions. But so far, it has been impossible to fully characterize the sequence of the virion infecting any given cell, since conventional techniques such as flow cytometry and single-cell transcriptome sequencing (scRNA-seq) only detect if a protein or transcript is present, not its sequence. Here we develop a new approach that uses long-read PacBio sequencing to determine the sequences of virions infecting single cells. We show that viral genetic variation explains some but not all of the cell-to-cell variability in viral gene expression and innate immune induction. Overall, our study provides the first complete picture of how viral mutations affect the course of infection in single cells.
Abstract
The use of muon scattering tomography for the non-invasive
characterisation of nuclear waste is well established. We report
here on the application of a combination of feature discriminators
...and multivariate analysis techniques to locate and identify
materials in nuclear waste drums. After successful training and
optimisation of the algorithms they are then tested on a range of
material configurations to assess the system's performance and
limitations. The system is able to correctly identify uranium, iron
and lead objects on a few cm scale. The system's sensitivity
to small uranium objects is also established as
0.90
+0.07
-0.12
, with a false positive rate of
0.12
+0.12
-0.07
.
Single-molecule techniques are powerful tools that can be used to study the kinetics and mechanics of a variety of enzymes and their complexes. Force spectroscopy, for example, can be used to control ...the force applied to a single molecule and thereby facilitate the investigation of real-time nucleic acid-protein interactions. In magnetic tweezers, which offer straightforward control and compatibility with fluorescence measurements or parallel tracking modes, force-measurement typically relies on the analysis of positional fluctuations through video microscopy. Significant errors in force estimates, however, may arise from incorrect spectral analysis of the Brownian motion in the magnetic tweezers. Here we investigated physical and analytical optimization procedures that can be used to improve the range over which forces can be reliably measured. To systematically probe the limitations of magnetic tweezers spectral analysis, we have developed a magnetic tweezers simulator, whose outcome was validated with experimental data. Using this simulator, we evaluate methods to correctly perform force experiments and provide guidelines for correct force calibration under configurations that can be encountered in typical magnetic tweezers experiments.
Islet cell transplantation can cure type 1 diabetes, but allograft rejection and recurrent autoimmunity may contribute to decreasing insulin independence over time. In this study we report the ...association of allograft‐specific proliferative and cytokine profiles with clinical outcome.
Peripheral blood mononuclear cells were obtained of 20 islet recipients. Cytokine values in mixed lymphocyte cultures (MLC) were determined using stimulator cells with graft‐specific HLA class II. Qualitative and quantitative cytokine profiles were determined before and after islet transplantation, blinded from clinical outcome. Cytotoxic T Lymphocyte precursor (CTLp) assays were performed to determine HLA class I alloreactivity.
Allograft‐specific cytokine profiles were skewed toward a Th2 or regulatory (Treg) phenotype after transplantation in insulin‐independent, but not in insulin‐requiring recipients. IFNγ/IL10 ratio and MLC proliferation decreased after transplantation in insulin‐independent recipients (p = 0.006 and p = 0.01, respectively). Production of the Treg cytokine IL10 inversely correlated with proliferation in alloreactive MLC (p = 0.008) and CTLp (p = 0.005). Production of IL10 combined with low‐MLC reactivity associated significantly with insulin independence.
The significant correlation between allograft‐specific cytokine profiles and clinical outcome may reflect the induction of immune regulation in successfully transplanted recipients. Islet donor‐specific IL10 production correlates with low alloreactivity and superior islet function.
In a cohort of human islet transplant recipients, allograft‐specific cytokine profiles correlated with clinical outcome, possibly indicating induction of immune regulation and providing a tool for future monitoring.
The RNA-dependent RNA polymerase (RdRp) of the influenza A virus replicates and transcribes the viral genome segments in the nucleus of the host cell. To transcribe these viral genome segments, the ...RdRp "snatches" capped RNA oligonucleotides from nascent host cell mRNAs and aligns these primers to the ultimate or penultimate nucleotide of the segments for the initiation of viral mRNA synthesis. It has been proposed that this initiation process is not processive and that the RdRp uses a prime-realign mechanism during transcription. Here we provide
evidence for the existence of this transcriptional prime-realign mechanism but show that it functions efficiently only for primers that are short or cannot stably base pair with the template. In addition, we demonstrate that transcriptional elongation is dependent on the priming loop of the PB1 subunit of the RdRp. We propose that the prime-realign mechanism may be used to rescue abortive transcription initiation events or cope with sequence variation among primers. Overall, these observations advance our mechanistic understanding of how influenza A virus initiates transcription correctly and efficiently.
Influenza A virus causes severe disease in humans and is considered a major global health threat. The virus replicates and transcribes its genome by using an enzyme called the RNA polymerase. To ensure that the genome is amplified faithfully and abundant viral mRNAs are made for viral protein synthesis, the viral RNA polymerase must transcribe the viral genome efficiently. In this report, we characterize a structure inside the polymerase that contributes to the efficiency of viral mRNA synthesis.
The influenza A virus genome consists of eight segments of single-stranded RNA. These segments are replicated and transcribed by a viral RNA-dependent RNA polymerase (RdRp) that is made up of the ...influenza virus proteins PB1, PB2, and PA. To copy the viral RNA (vRNA) genome segments and the cRNA segments, the replicative intermediate of viral replication, the RdRp must use two promoters and two different
initiation mechanisms. On the vRNA promoter, the RdRp initiates on the 3' terminus, while on the cRNA promoter, the RdRp initiates internally and subsequently realigns the nascent vRNA product to ensure that the template is copied in full. In particular, the latter process, which is also used by other RNA viruses, is not understood. Here we provide mechanistic insight into priming and realignment during influenza virus replication and show that it is controlled by the priming loop and a helix-loop-helix motif of the PB1 subunit of the RdRp. Overall, these observations advance our understanding of how the influenza A virus initiates viral replication and amplifies the genome correctly.
Influenza A viruses cause severe disease in humans and are considered a major threat to our economy and health. The viruses replicate and transcribe their genome by using an enzyme called the RNA polymerases. To ensure that the genome is amplified faithfully and that abundant viral mRNAs are made for viral protein synthesis, the RNA polymerase must work correctly. In this report, we provide insight into the mechanism that the RNA polymerase employs to ensure that the viral genome is copied correctly.
SUMMARYNegative and ambisense RNA viruses are the causative agents of important human diseases such as influenza, measles, Lassa fever, and Ebola hemorrhagic fever. The viral genome of these RNA ...viruses consists of one or more single-stranded RNA molecules that are encapsidated by viral nucleocapsid proteins to form a ribonucleoprotein complex (RNP). This RNP acts as protection, as a scaffold for RNA folding, and as the context for viral replication and transcription by a viral RNA polymerase. However, the roles of the viral nucleoproteins extend beyond these functions during the viral infection cycle. Recent advances in structural biology techniques and analysis methods have provided new insights into the formation, function, dynamics, and evolution of negative sense virus nucleocapsid proteins, as well as the role that they play in host innate immune responses against viral infection. In this review, we discuss the various roles of nucleocapsid proteins, both in the context of RNPs and in RNA-free states, as well as the open questions that remain.
To examine the effects of a 1-year exercise intervention on sex hormone levels in postmenopausal women and whether any effects are mediated by changes in body fat composition.
We randomly assigned ...189 sedentary postmenopausal women (age 50 to 69 years, body mass index of 22 to 40 kg/m(2)) to an exercise intervention (n = 96) or a control group (n = 93). The intervention combined aerobic and strength training and comprised supervised group sessions and home-based exercises (a total of 2.5 h/wk). Between-group differences in sex hormone levels (at baseline and 4 and 12 months) were examined with generalized estimating equations.
In total, 183 women (97%) completed the study. Overall, the exercise intervention did not result in favorable effects on sex hormone levels. Among women who lost more than 2% body fat, declines in all estrogens were not significantly different between exercisers and controls. Androgen levels decreased significantly in the exercise group who lost body fat compared with their peers in the control group. Furthermore, this study confirmed that fat loss was significantly associated with declines in postmenopausal estrogen levels. Although not significant, a similar trend was observed for the androgens.
This study confirms that fat loss is associated with changes in postmenopausal sex hormone levels and suggests that exercise may be effective in inducing favorable changes in these hormones.
Damage caused by incidents with transport tanks with compressed liquified gas is amongst the most extreme that can be encountered with transport vessels. This is particularly the case with the ...Boiling Liquid Expanding Vapor Explosion (BLEVE), which may occur if such a tank is exposed to fire for a prolonged period. Therefore, the local Dutch LPG transport sector adopted a thermally insulating tank coating as a ‘standard outfit’ for their tank trailers, with the aim to delay a BLEVE for a sufficiently long period for emergency services to take appropriate measures and for people near the accident location to be evacuated. On a European scale however, no consensus has been reached on the cost-benefit of such measures.
With the current drive towards “greener” and renewable energy sources, this issue has regained attention with alternative fuels such as LNG, CNG and Hydrogen and a need was felt for (better) theoretical models and experimental data concerning the behavior of transport tanks carrying these substances.
In this paper a new tank thermal (equilibrium) model is described to predict pressure and temperature behavior of a multi layered, thermally insulated tank containing a compressed liquified gas exposed to heat. Results are compared with data of three bonfire experiments, in which 3 m3 tanks, filled for ca. 50% with LPG were exposed to fire. A good match between modelled and experimental pressure and temperature evolution in time could be obtained using a constant value for the thermal conductivity of the insulation layer. The modelling showed that the thermal insulation value is crucial for an accurate prediction of these parameters as well as the opening time for a pressure safety valve. As relevant temperatures may cover a very wide range (from cryogenic in LNG-tanks to over 1000 °C in a fire) knowledge of the thermal (and physical) behavior of the insulating layer over a large temperature range is essential.
The same holds for the behavior of the PRV when subjected to fire. Extreme temperatures may also lead to deviating behavior from what is expected based on the initial settings.
•New equilibrium model calculates pressure and temperature in a thermally coated vessel with compressed liquified gases.•Model shows importance of accurate information on thermal conductivity on full temperature range.•Model has been successfully validated against three datasets from bonfire tests with thermally coated tanks filled will LPG.•Bonfire tests show pressure safety valve behavior at high temperature deviates from initially set values.
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