The bacterial cytoskeleton modulates motility, type 3 secretion, and colonization in Salmonella.

Although there have been great advances in our understanding of the bacterial cytoskeleton, major gaps remain in our knowledge of its importance to virulence. In this study we have explored the contribution of the bacterial cytoskeleton to the ability of Salmonella to express and assemble virulence factors and cause disease. The bacterial actin-like protein MreB polymerises into helical filaments and interacts with other cytoskeletal elements including MreC to control cell-shape. As mreB appears to be an essential gene, we have constructed a viable ΔmreC depletion mutant in Salmonella. Using a broad range of independent biochemical, fluorescence and phenotypic screens we provide evidence that the Salmonella pathogenicity island-1 type three secretion system (SPI1-T3SS) and flagella systems are down-regulated in the absence of MreC. In contrast the SPI-2 T3SS appears to remain functional. The phenotypes have been further validated using a chemical genetic approach to disrupt the functionality of MreB. Although the fitness of ΔmreC is reduced in vivo, we observed that this defect does not completely abrogate the ability of Salmonella to cause disease systemically. By forcing on expression of flagella and SPI-1 T3SS in trans with the master regulators FlhDC and HilA, it is clear that the cytoskeleton is dispensable for the assembly of these structures but essential for their expression. As two-component systems are involved in sensing and adapting to environmental and cell surface signals, we have constructed and screened a panel of such mutants and identified the sensor kinase RcsC as a key phenotypic regulator in ΔmreC. Further genetic analysis revealed the importance of the Rcs two-component system in modulating the expression of these virulence factors. Collectively, these results suggest that expression of virulence genes might be directly coordinated with cytoskeletal integrity, and this regulation is mediated by the two-component system sensor kinase RcsC.

Decay of vibrations along deepwater mooring lines

Model tests for global design verification of deepwater floating structures cannot be made at reasonable scales. An overview of recent research efforts to tackle this challenge is given first, introducing the concept of line truncation techniques. In such a method the upper sections of each line are modelled in detail, capturing the wave action zone and all coupling effects with the vessel. These terminate to an approximate analytical model, that aims to simulate the remainder of the line. The rationale for this is that in deep water the transverse elastic waves of a line are likely to decay before they are reflected at the seabed. The focus of this paper is the verification of this rationale and the ongoing work, which is considering ways to produce a truncation model. Transverse dynamics of a mooring line are modelled using the equations of motion of an inextensible taut string, submerged in still water, one end fixed at the bottom the other assumed to follow the vessel response, which can be harmonic or random. Nonlinear hydrodynamic damping is included; bending and VIV effects are neglected. A dimensional analysis, supported by exact benchmark numerical solutions, has shown that it is possible to produce a universal curve for the decay of transverse vibrations along the line, which is suitable for any kind of line with any top motion. This has a significant engineering benefit, allowing for a rapid assessment of line dynamics - it is very useful in deciding whether a truncated line model is appropriate, and if so, at which point truncation might be applied. Initial efforts in developing a truncated model show that a linearized numerical solution in the frequency domain matches very closely the exact benchmark. Copyright © 2011 by ASME.

The decay of Saffman and batchelor turbulence subject to rotation, stratification or an imposed magnetic field

We consider unforced, statistically-axisymmetric turbulence evolving in the presence of a background rotation, an imposed stratification, or a uniform magnetic field. We focus on two canonical cases: Saffman turbulence, in which E(κ → 0) ∼ κ 2, and Batchelor turbulence, in which E(κ → 0) ∼ κ 4. It has recently been shown that, provided the large scales evolve in a self-similar manner, then u ⊥ 2ℓ ⊥ 2ℓ // = constant in Saffman turbulence and u ⊥ 2ℓ ⊥ 4ℓ // = constant in Batchelor turbulence (Davidson, 2009, 2010). Here the subscripts ⊥ and // indicate directions perpendicular and parallel to the axis of symmetry, and ℓ ⊥, ℓ //, and u ⊥ are suitably defined integral scales. These constraints on the integral scales allow us to make simple, testable predictions for the temporal evolution of ℓ ⊥, ℓ //, and u ⊥ in rotating, stratified and MHD turbulence.

The application of STEM and in situ controlled dehydration to bacterial systems using ESEM.

Transmission imaging with an environmental scanning electron microscope (ESEM) (Wet STEM) is a recent development in the field of electron microscopy, combining the simple preparation inherent to ESEM work with an alternate form of contrast available through a STEM detector. Because the technique is relatively new, there is little information available on how best to apply this technique and which samples it is best suited for. This work is a description of the sample preparation and microscopy employed by the authors for imaging bacteria with Wet STEM (scanning transmission electron microscopy). Three different bacterial samples will be presented in this study: first, used as a model system, is Escherichia coli for which the contrast mechanisms of STEM are demonstrated along with the visual effects of a dehydration-induced collapse. This collapse, although clearly in some sense artifactual, is thought to lead to structurally meaningful morphological information. Second, Wet STEM is applied to two distinct bacterial systems to demonstrate the novel types of information accessible by this approach: the plastic-producing Cupriavidus necator along with wild-type and ΔmreC knockout mutants of Salmonella enterica serovar Typhimurium. Cupriavidus necator is shown to exhibit clear internal differences between bacteria with and without plastic granules, while the ΔmreC mutant of S. Typhimurium has an internal morphology distinct from that of the wild type.

The application of STEM and in situ controlled dehydration to bacterial systems using ESEM

Transmission imaging with an environmental scanning electron microscope (ESEM) (Wet STEM) is a recent development in the field of electron microscopy, combining the simple preparation inherent to ESEM work with an alternate form of contrast available through a STEM detector. Because the technique is relatively new, there is little information available on how best to apply this technique and which samples it is best suited for. This work is a description of the sample preparation and microscopy employed by the authors for imaging bacteria with Wet STEM (scanning transmission electron microscopy). Three different bacterial samples will be presented in this study: first, used as a model system, is Escherichia coli for which the contrast mechanisms of STEM are demonstrated along with the visual effects of a dehydration-induced collapse. This collapse, although clearly in some sense artifactual, is thought to lead to structurally meaningful morphological information. Second, Wet STEM is applied to two distinct bacterial systems to demonstrate the novel types of information accessible by this approach: the plastic-producing Cupriavidus necator along with wild-type and δmreC knockout mutants of Salmonella enterica serovar Typhimurium. Cupriavidus necator is shown to exhibit clear internal differences between bacteria with and without plastic granules, while the δmreC mutant of S. Typhimurium has an internal morphology distinct from that of the wild type. © 2012 Wiley Periodicals, Inc.

Attenuated Salmonella Typhimurium lacking the pathogenicity island-2 type 3 secretion system grow to high bacterial numbers inside phagocytes in mice.

Intracellular replication within specialized vacuoles and cell-to-cell spread in the tissue are essential for the virulence of Salmonella enterica. By observing infection dynamics at the single-cell level in vivo, we have discovered that the Salmonella pathogenicity island 2 (SPI-2) type 3 secretory system (T3SS) is dispensable for growth to high intracellular densities. This challenges the concept that intracellular replication absolutely requires proteins delivered by SPI-2 T3SS, which has been derived largely by inference from in vitro cell experiments and from unrefined measurement of net growth in mouse organs. Furthermore, we infer from our data that the SPI-2 T3SS mediates exit from infected cells, with consequent formation of new infection foci resulting in bacterial spread in the tissues. This suggests a new role for SPI-2 in vivo as a mediator of bacterial spread in the body. In addition, we demonstrate that very similar net growth rates of attenuated salmonellae in organs can be derived from very different underlying intracellular growth dynamics.

High-resolution γ-ray spectroscopy: A versatile tool for nuclear β-decay studies at TRIUMF-ISAC

High-resolution γ-ray spectroscopy is essential to fully exploit the unique, high-quality beams available at the next generation of radioactive ion beam facilities such as the TRIUMF isotope separator and accelerator (ISAC). The 8π spectrometer, which consists of 20 Compton-suppressed HPGe detectors, has recently been reconfigured for a vigorous research programme in weak interaction and nuclear structure physics. With the addition of a variety of ancillary detectors it has become the world's most powerful device dedicated to β-decay studies. This paper provides a brief overview of the apparatus and highlights from recent experiments. © 2005 IOP Publishing Ltd.

Decay heat in fast reactors with transuranic fuels

In this work, we performed an evaluation of decay heat power of advanced, fast spectrum, lead and molten salt-cooled reactors, with flexible conversion ratio. The decay heat power was calculated using the BGCore computer code, which explicitly tracks over 1700 isotopes in the fuel throughout its burnup and subsequent decay. In the first stage, the capability of the BGCore code to accurately predict the decay heat power was verified by performing a benchmark calculation for a typical UO2 fuel in a Pressurized Water Reactor environment against the (ANSI/ANS-5.1-2005, "Decay Heat Power in Light Water Reactors," American National Standard) standard. Very good agreement (within 5%) between the two methods was obtained. Once BGCore calculation capabilities were verified, we calculated decay power for fast reactors with different coolants and conversion ratios, for which no standard procedure is currently available. Notable differences were observed for the decay power of the advanced reactor as compared with the conventional UO2 LWR. The importance of the observed differences was demonstrated by performing a simulation of a Station Blackout transient with the RELAP5 computer code for a lead-cooled fast reactor. The simulation was performed twice: using the code-default ANS-79 decay heat curve and using the curve calculated specifically for the studied core by BGCore code. The differences in the decay heat power resulted in failure to meet maximum cladding temperature limit criteria by ∼100 °C in the latter case, while in the transient simulation with the ANS-79 decay heat curve, all safety limits were satisfied. The results of this study show that the design of new reactor safety systems must be based on decay power curves specific to each individual case in order to assure the desired performance of these systems. © 2009 Elsevier B.V. All rights reserved.

Context-dependent decay of motor memories during skill acquisition.

Current models of motor learning posit that skill acquisition involves both the formation and decay of multiple motor memories that can be engaged in different contexts. Memory formation is assumed to be context dependent, so that errors most strongly update motor memories associated with the current context. In contrast, memory decay is assumed to be context independent, so that movement in any context leads to uniform decay across all contexts. We demonstrate that for both object manipulation and force-field adaptation, contrary to previous models, memory decay is highly context dependent. We show that the decay of memory associated with a given context is greatest for movements made in that context, with more distant contexts showing markedly reduced decay. Thus, both memory formation and decay are strongest for the current context. We propose that this apparently paradoxical organization provides a mechanism for optimizing performance. While memory decay tends to reduce force output, memory formation can correct for any errors that arise, allowing the motor system to regulate force output so as to both minimize errors and avoid unnecessary energy expenditure. The motor commands for any given context thus result from a balance between memory formation and decay, while memories for other contexts are preserved.

Vibration decay along deepwater moorings with complex geometry and configuration, as a tool for truncation

This paper is concerned with the difficulties in model testing deepwater structures at reasonable scales. An overview of recent research efforts to tackle this challenge is given first, introducing the concept of line truncation. Passive truncation has traditionally been the preferred method by industry; however, these techniques tend to suffer in capturing accurately line dynamic response and so reproducing peak tensions. In an attempt to improve credibility of model test data the proposed truncation procedure sets up the truncated model, based on line dynamic response rather than quasi-static system stiffness. Vibration decay of transverse elastic waves due to fluid drag forces is assessed and it is found that below a certain length criterion, the transverse vibrational characteristics for each line are inertia driven, hence with respect to these motions the truncated model can assume a linear damper whose coefficient depends on the local line properties and vibration frequency. Initially a simplified taut string model is assumed for which the line is submerged in still water, one end fixed at the bottom the other assumed to follow the vessel response, which can be harmonic or random. A dimensional analysis, supported by exact benchmark numerical solutions, has shown that it is possible to produce a general guideline for the truncation length criterion, which is suitable for any kind of line with any top motion. The focus of this paper is to extend this work to a more complex line configuration of a conventional deepwater mooring line and so enhance the generality of the truncation guideline. The paper will close with an example case study of a spread mooring system, applying this method to create an equivalent numerical model at a reduced depth that replicates exactly the static and dynamic characteristics of the full depth system. Copyright © 2012 by ASME.

Molecular characterization and expression profiles in response to bacterial infection of Chinese soft-shelled turtle interleukin-8 (IL-8), the first reptilian chemokine gene

In this study, an IL-8 homologue has been cloned and identified from a reptile, Chinese soft-shelled turtle for the first time. The full-length cDNA of turtle IL-8 was 1188 bp and contained a 312 bp open reading frame (ORF) coding for a protein of 104 amino acids. The chemokine CXC domain, which contained Glu-Leu-Arg (ELR) motif and four cysteine residues, was well conserved in turtle IL-8. The 4924 bp genomic DNA of turtle IL-8 contained four exons and three introns. Phylogenetic analysis showed that the amino acid sequence of turtle IL-8 clustered together with birds. RT-PCR analysis showed that turtle IL-8 mRNA was constitutively expressed liver, spleen, kidney, heart, blood and intestine tissues of control turtles. Real-time quantitative PCR analysis further indicated that the turtle IL-8 mRNA expression was apparent in various tissues at 8 h and up-regulated significantly during 8 h-7 d after Aeromonas hydrophila infection. The present studies will help us to understand the evolution of IL-8 molecule and the inflammatory response mechanism in reptiles. (C) 2009 Elsevier Ltd. All rights reserved.