Chemical environment of unusually Ge- and Pb-rich willemite, Tres Marias mine, Mexico

The Tres Marias carbonate-hosted Zn-Ge deposit in Chihuahua, Mexico contains willemite [Zn$_{2}$SiO$_{4}$] with unusually high concentrations of minor and trace elements (e.g., Pb, Ge, As, P, V); Pb concentrations are as high as 2 wt %, and Ge may reach 4000 ppm (average 900 ppm). Electron microprobe analyses and synchrotron X-ray fluorescence maps show that Zn and Ge, as well as Zn and Pb are negatively correlated, whereas Ge and Pb are positively correlated across zoned willemite crystals. In cathodoluminescence (CL) images, those areas of willemite having high trace element concentrations have no, or low CL intensities, whereas zones low in trace elements (except for P) display bright blue CL colors. X-ray absorption fine structure (XAFS) spectroscopy was used to characterize the chemical nature of Ge and Pb in willemite. Comparisons to reference spectra of natural and artificial substances points to the presence of Ge$^{4+}$ and Pb$^{2+}$ in Tres Marias willemite. No evidence for Pb$^{4+}$ was detected. Oscillatory zonation reflects trace element incorporation into willemite from the oxidation of primary Ge-bearing sphalerite and galena (PbS) by siliceous aqueous fluids

Gait rehabilitation machines based on programmable footplates

BACKGROUND: Gait restoration is an integral part of rehabilitation of brain lesioned patients. Modern concepts favour a task-specific repetitive approach, i.e. who wants to regain walking has to walk, while tone-inhibiting and gait preparatory manoeuvres had dominated therapy before. Following the first mobilization out of the bed, the wheelchair-bound patient should have the possibility to practise complex gait cycles as soon as possible. Steps in this direction were treadmill training with partial body weight support and most recently gait machines enabling the repetitive training of even surface gait and even of stair climbing. RESULTS: With treadmill training harness-secured and partially relieved wheelchair-mobilised patients could practise up to 1000 steps per session for the first time. Controlled trials in stroke and SCI patients, however, failed to show a superior result when compared to walking exercise on the floor. Most likely explanation was the effort for the therapists, e.g. manually setting the paretic limbs during the swing phase resulting in a too little gait intensity. The next steps were gait machines, either consisting of a powered exoskeleton and a treadmill (Lokomat, AutoAmbulator) or an electromechanical solution with the harness secured patient placed on movable foot plates (Gait Trainer GT I). For the latter, a large multi-centre trial with 155 non-ambulatory stroke patients (DEGAS) revealed a superior gait ability and competence in basic activities of living in the experimental group. The HapticWalker continued the end effector concept of movable foot plates, now fully programmable and equipped with 6 DOF force sensors. This device for the first time enables training of arbitrary walking situations, hence not only the simulation of floor walking but also for example of stair climbing and perturbations. CONCLUSION: Locomotor therapy is a fascinating new tool in rehabilitation, which is in line with modern principles of motor relearning promoting a task-specific repetitive approach. Sophisticated technical developments and positive randomized controlled trials form the basis of a growing acceptance worldwide to the benefits or our patients

Norvaline is accumulated after a down-shift of oxygen in Escherichia coli W3110

<p>Abstract</p> <p>Background</p> <p>Norvaline is an unusual non-proteinogenic branched-chain amino acid which has been of interest especially during the early enzymological studies on regulatory mutants of the branched-chain amino acid pathway in <it>Serratia marcescens</it>. Only recently norvaline and other modified amino acids of the branched-chain amino acid synthesis pathway got attention again when they were found to be incorporated in minor amounts in heterologous proteins with a high leucine or methionine content. Earlier experiments have convincingly shown that norvaline and norleucine are formed from pyruvate being an alternative substrate of α-isopropylmalate synthase, however so far norvaline accumulation was not shown to occur in non-recombinant strains of <it>E. coli</it>.</p> <p>Results</p> <p>Here we show that oxygen limitation causes norvaline accumulation in <it>E. coli </it>K-12 W3110 during grow in glucose-based mineral salt medium. Norvaline accumulates immediately after a shift to oxygen limitation at high glucose concentration. On the contrary free norvaline is not accumulated in <it>E. coli </it>W3110 in aerobic cultures. The analysis of medium components, supported by transcriptomic studies proposes a purely metabolic overflow mechanism from pyruvate into the branched chain amino acid synthesis pathway, which is further supported by the significant accumulation of pyruvate after the oxygen downshift. The results indicate overflow metabolism from pyruvate as necessary and sufficient, but deregulation of the branched chain amino acid pathway may be an additional modulating parameter.</p> <p>Conclusion</p> <p>Norvaline synthesis has been so far mainly related to an imbalance of the synthesis of the branched chain amino acids under conditions were pyruvate level is high. Here we show that simply a downshift of oxygen is sufficient to cause norvaline accumulation at a high glucose concentration as a consequence of the accumulation of pyruvate and its direct chain elongation over α-ketobutyrate and α-ketovalerate.</p> <p>Although the flux to norvaline is low, millimolar concentrations are accumulated in the cultivation broth, which is far above the level which has been discussed for being relevant for misincorporation of norvaline into recombinant proteins. Therefore we believe that our finding is relevant for recombinant protein production but also may even have implications for the physiology of <it>E. coli </it>under oxygen limitation in general.</p

Determination of the Entire Stent Surface Area by a New Analytical Method

Stenting is a widely used treatment procedure for coronary artery disease around the world. Stents have a complex geometry, which makes the characterization of their corrosion difficult due to the absence of a mathematical model to calculate the entire stent surface area (ESSA). Therefore, corrosion experiments with stents are mostly based on qualitative analysis. Additionally, the quantitative analysis of corrosion is conducted with simpler samples made of stent material instead of stents, in most cases. At present, several methods are available to calculate the stent outer surface area (SOSA), whereas no model exists for the calculation of the ESSA. This paper presents a novel mathematical model for the calculation of the ESSA using the SOSA as one of the main parameters. The ESSA of seven magnesium alloy stents (MeKo Laser Material Processing GmbH, Sarstedt, Germany) were calculated using the developed model. The calculated SOSA and ESSA for all stents are 33.34%(±0.26%) and 111.86 mm (±0.85 mm), respectively. The model is validated by micro-computed tomography (micro-CT), with a difference of 12.34% (±0.46%). The value of corrosion rates calculated using the ESSA computed with the developed model will be 12.34% (±0.46%) less than that of using ESSA obtained by micro-CT

Different Evolutionary Modifications as a Guide to Rewire Two-Component Systems

Two-component systems (TCS) are short signalling pathways generally occurring in prokaryotes. They frequently regulate prokaryotic stimulus responses and thus are also of interest for engineering in biotechnology and synthetic biology. The aim of this study is to better understand and describe rewiring of TCS while investigating different evolutionary scenarios. Based on large-scale screens of TCS in different organisms, this study gives detailed data, concrete alignments, and structure analysis on three general modification scenarios, where TCS were rewired for new responses and functions: (i) exchanges in the sequence within single TCS domains, (ii) exchange of whole TCS domains; (iii) addition of new components modulating TCS function. As a result, the replacement of stimulus and promotor cassettes to rewire TCS is well defined exploiting the alignments given here. The diverged TCS examples are non-trivial and the design is challenging. Designed connector proteins may also be useful to modify TCS in selected cases

The MarR-Type Repressor MhqR Confers Quinone and Antimicrobial Resistance in Staphylococcus aureus

Aims: Quinone compounds are electron carriers and have antimicrobial and toxic properties due to their mode of actions as electrophiles and oxidants. However, the regulatory mechanism of quinone resistance is less well understood in the pathogen Staphylococcus aureus. Results: Methylhydroquinone (MHQ) caused a thiol-specific oxidative and electrophile stress response in the S. aureus transcriptome as revealed by the induction of the PerR, QsrR, CstR, CtsR, and HrcA regulons. The SACOL2531-29 operon was most strongly upregulated by MHQ and was renamed as mhqRED operon based on its homology to the Bacillus subtilis locus. Here, we characterized the MarR-type regulator MhqR (SACOL2531) as quinone-sensing repressor of the mhqRED operon, which confers quinone and antimicrobial resistance in S. aureus. The mhqRED operon responds specifically to MHQ and less pronounced to pyocyanin and ciprofloxacin, but not to reactive oxygen species (ROS), hypochlorous acid, or aldehydes. The MhqR repressor binds specifically to a 9–9 bp inverted repeat (MhqR operator) upstream of the mhqRED operon and is inactivated by MHQ in vitro, which does not involve a thiol-based mechanism. In phenotypic assays, the mhqR deletion mutant was resistant to MHQ and quinone-like antimicrobial compounds, including pyocyanin, ciprofloxacin, norfloxacin, and rifampicin. In addition, the mhqR mutant was sensitive to sublethal ROS and 24 h post-macrophage infections but acquired an improved survival under lethal ROS stress and after long-term infections. Innovation: Our results provide a link between quinone and antimicrobial resistance via the MhqR regulon of S. aureus. Conclusion: The MhqR regulon was identified as a novel resistance mechanism towards quinone-like antimicrobials and contributes to virulence of S. aureus under long-term infections

Trade-off for survival: Microbiome response to chemical exposure combines activation of intrinsic resistances and adapted metabolic activity

The environmental microbiota is increasingly exposed to chemical pollution. While the emergence of multi-resistant pathogens is recognized as a global challenge, our understanding of antimicrobial resistance (AMR) development from native microbiomes and the risks associated with chemical exposure is limited. By implementing a lichen as a bioindicator organism and model for a native microbiome, we systematically examined responses towards antimicrobials (colistin, tetracycline, glyphosate, and alkylpyrazine). Despite an unexpectedly high resilience, we identified potential evolutionary consequences of chemical exposure in terms of composition and functioning of native bacterial communities. Major shifts in bacterial composition were observed due to replacement of naturally abundant taxa; e.g. Chthoniobacterales by Pseudomonadales. A general response, which comprised activation of intrinsic resistance and parallel reduction of metabolic activity at RNA and protein levels was deciphered by a multi-omics approach. Targeted analyses of key taxa based on metagenome-assembled genomes reflected these responses but also revealed diversified strategies of their players. Chemical-specific responses were also observed, e.g., glyphosate enriched bacterial r-strategists and activated distinct ARGs. Our work demonstrates that the high resilience of the native microbiota toward antimicrobial exposure is not only explained by the presence of antibiotic resistance genes but also adapted metabolic activity as a trade-off for survival. Moreover, our results highlight the importance of native microbiomes as important but so far neglected AMR reservoirs. We expect that this phenomenon is representative for a wide range of environmental microbiota exposed to chemicals that potentially contribute to the emergence of antibiotic-resistant bacteria from natural environments

S-thioallylation of proteins by the garlic defence substance allicin and its biological effects

A single clove of edible garlic (Allium sativum L.) of about 10 g produces up to 5 mg of allicin (diallylthiosulfinate), a thiol-reactive sulfur-containing defence substance that gives injured garlic tissue its characteristic smell. Allicin induces apoptosis or necrosis in a dose-dependent manner but biocompatible doses influence cellular metabolism and signalling cascades. Oxidation of protein thiols and depletion of the glutathione pool are thought to be responsible for allicin's physiological effects. Here, we studied the effect of allicin on post-translational thiol-modification in human Jurkat T-cells using shotgun LC-MS/MS analyses. We identified 332 proteins that were modified by S-thioallylation in the Jurkat cell proteome which causes a mass shift of 72 Da on cysteines. Many S-thioallylated proteins are highly abundant proteins, including cytoskeletal proteins tubulin, actin, cofilin, filamin and plastin-2, the heat shock chaperones HSP90 and HSPA4, the glycolytic enzymes GAPDH, ALDOA, PKM as well the protein translation factor EEF2. Allicin disrupted the actin cytoskeleton in murine L929 fibroblasts. Allicin stimulated the immune response by causing Zn2+ release from proteins and increasing the Zn2+-dependent IL-1-triggered production of IL-2 in murine EL-4 T-cells. Furthermore, allicin caused inhibition of enolase activity, an enzyme considered a cancer therapy target. In conclusion, our study revealed the widespread extent of S-thioallylation in the human Jurkat cell proteome and showed effects of allicin exposure on essential cellular functions of selected targets, many of which are targets for cancer therapy

Complete genome sequence of Staphylococcus aureus 6850, a highly cytotoxic and clinically virulent Methicillin-sensitive strain with distant relatedness to prototype strains

Staphylococcus aureus is a frequent human commensal bacterium and pathogen. Here we report the complete genome sequence of strain 6850 (spa type t185; sequence type 50 [ST50]), a highly cytotoxic and clinically virulent methicillin-sensitive strain from a patient with complicated S. aureus bacteremia associated with osteomyelitis and septic arthritis