Controlling intracavity dual-comb soliton motion in a single-fiber laser.

Ultrafast science builds on dynamic compositions of precisely timed light pulses, and evolving groups of pulses are observed in almost every mode-locked laser. However, the underlying physics has rarely been controlled or used until now. Here, we demonstrate a general approach to control soliton motion inside a dual-comb laser and the programmable synthesis of ultrashort pulse patterns. Introducing single-pulse modulation inside an Er:fiber laser, we rapidly shift the timing between two temporally separated soliton combs. Their superposition outside the cavity yields ultrashort soliton sequences. On the basis of real-time spectral interferometry, we observe the deterministic switching of intersoliton separation arising from the interplay of attracting and repulsing forces via ultrafast nonlinearity and laser gain dynamics. Harnessing these insights, we demonstrate the high-speed all-optical synthesis of nano- to picosecond pump-probe delays and programmable free-form soliton trajectories. This concept may pave the way to a new class of all-optical delay generators for ultrafast measurements at unprecedented high tuning, cycling, and acquisition speeds.

A Photoconvertible Reporter System for Bacterial Metabolic Activity Reveals That Staphylococcus aureus Enters a Dormant-Like State to Persist within Macrophages.

Staphylococcus aureus is a leading cause of difficult-to-treat infections. The capacity of S. aureus to survive and persist within phagocytic cells is an important factor contributing to therapy failures and infection recurrence. Therefore, interfering with S. aureus intracellular persistence is key to treatment success. In this study, we used a S. aureus strain carrying the reporter mKikumeGR that enables the monitoring of the metabolic status of intracellular bacteria to achieve a better understanding of the molecular mechanisms facilitating S. aureus survival and persistence within macrophages. We found that shortly after bacteria internalization, a large fraction of macrophages harbored mainly S. aureus with high metabolic activity. This population decreased gradually over time with the concomitant increase of a macrophage subpopulation harboring S. aureus with low metabolic activity, which prevailed at later times. A dual RNA-seq analysis performed in each macrophage subpopulation showed that the host transcriptional response was similar between both subpopulations. However, intracellular S. aureus exhibited disparate gene expression profiles depending on its metabolic state. Whereas S. aureus with high metabolic activity exhibited a greater expression of genes involved in protein synthesis and proliferation, bacteria with low metabolic activity displayed a higher expression of oxidative stress response-related genes, silenced genes involved in energy-consuming processes, and exhibited a dormant-like state. Consequently, we propose that reducing metabolic activity and entering into a dormant-like state constitute a survival strategy used by S. aureus to overcome the adverse environment encountered within macrophages and to persist in the intracellular niche. IMPORTANCE The capacity of Staphylococcus aureus to survive and persist within phagocytic cells has been associated with antibiotic treatment failure and recurrent infections. Here, we investigated the molecular mechanisms leading to S. aureus persistence within macrophages using a reporter system that enables to distinguish between intracellular bacteria with high and low metabolic activity in combinstion with a dual RNA-seq approach. We found that with the progression of infection, intracellular S. aureus transitions from a high metabolic state to a low metabolic dormant-like state by turning off major energy-consuming processes while remaining viable. This process seems to be driven by the level of stress encountered in the intracellular niche. Our study indicates that effective therapies by which to treat S. aureus infections should be able to target not only high metabolic bacteria but also intracellular dormant-like S. aureus.

Ultrafast imaging of terahertz electric waveforms using quantum dots.

Microscopic electric fields govern the majority of elementary excitations in condensed matter and drive electronics at frequencies approaching the Terahertz (THz) regime. However, only few imaging schemes are able to resolve sub-wavelength fields in the THz range, such as scanning-probe techniques, electro-optic sampling, and ultrafast electron microscopy. Still, intrinsic constraints on sample geometry, acquisition speed and field strength limit their applicability. Here, we harness the quantum-confined Stark-effect to encode ultrafast electric near-fields into colloidal quantum dot luminescence. Our approach, termed Quantum-probe Field Microscopy (QFIM), combines far-field imaging of visible photons with phase-resolved sampling of electric waveforms. By capturing ultrafast movies, we spatio-temporally resolve a Terahertz resonance inside a bowtie antenna and unveil the propagation of a Terahertz waveguide excitation deeply in the sub-wavelength regime. The demonstrated QFIM approach is compatible with strong-field excitation and sub-micrometer resolution-introducing a direct route towards ultrafast field imaging of complex nanodevices in-operando.

Complexome profiling on the Chlamydomonas lpa2 mutant reveals insights into PSII biogenesis and new PSII associated proteins.

While the composition and function of the major thylakoid membrane complexes are well understood, comparatively little is known about their biogenesis. The goal of this work was to shed more light on the role of auxiliary factors in the biogenesis of photosystem II (PSII). Here we have identified the homolog of LOW PSII ACCUMULATION 2 (LPA2) in Chlamydomonas. A Chlamydomonas reinhardtii lpa2 mutant grew slower in low light, was hypersensitive to high light, and exhibited aberrant structures in thylakoid membrane stacks. Chlorophyll fluorescence (Fv/Fm) was reduced by 38%. Synthesis and stability of newly made PSII core subunits D1, D2, CP43, and CP47 were not impaired. However, complexome profiling revealed that in the mutant CP43 was reduced to ~23% and D1, D2, and CP47 to ~30% of wild type levels. Levels of PSI and the cytochrome b6f complex were unchanged, while levels of the ATP synthase were increased by ~29%. PSII supercomplexes, dimers, and monomers were reduced to ~7%, ~26%, and ~60% of wild type levels, while RC47 was increased ~6-fold and LHCII by ~27%. We propose that LPA2 catalyses a step during PSII assembly without which PSII monomers and further assemblies become unstable and prone to degradation. The LHCI antenna was more disconnected from PSI in the lpa2 mutant, presumably as an adaptive response to reduce excitation of PSI. From the co-migration profiles of 1734 membrane-associated proteins, we identified three novel putative PSII associated proteins with potential roles in regulating PSII complex dynamics, assembly, and chlorophyll breakdown.

Peripheral neuropathy due to neuroborreliosis: Insensitivity for CXCL13 as early diagnostic marker.

The case of a 69-year-old woman with peripheral neuropathy caused by Lyme neuroborreliosis (LNB) in an endemic region in Eastern Austria is reported. The patient had noticed transient numbness of her left leg. On initial examination, she had patchy sensory disturbances of the left lower leg, but ancillary examinations of nerve conduction and cerebrospinal fluid (CSF), including the B-cell chemokine CXCL13, were normal. A re-tap performed 54 days later, following clinical progression with foot drop, widespread lower leg paresthesia, and pain, revealed an increased cell count, autochthonous IgM production, synthesis of Borrelia-specific IgM, and elevated CXCL13. Neurophysiological examinations disclosed an incomplete conduction block, mixed axonal and demyelinating sensorimotor neuropathy, and subacute neurogenic damage of muscles innervated by the peroneal nerve. This case study presents rare evidence of very early diagnostic findings in peripheral neuropathy caused by LNB. These are characterized by insensitivity of CXCL13 in CSF to aid earlier diagnosis and the development of an intrathecal immune response against Borrelia at a later stage. These findings reinforce the need for a re-tap to confirm the diagnosis and facilitate appropriate treatment in this rare manifestation of LNB.

Comparative study of hyperpure chlorine dioxide with two other irrigants regarding the viability of periodontal ligament stem cells.

OBJECTIVES: Periodontal ligament stem cells (PDLSCs) have an underlined significance as their high proliferative capacity and multipotent differentiation provide an important therapeutic potential. The integrity of these cells is frequently disturbed by the routinely used irrigative compounds applied as periodontal or endodontic disinfectants (e.g., hydrogen peroxide (H2O2) and chlorhexidine (CHX)). Our objectives were (i) to monitor the cytotoxic effect of a novel dental irrigative compound, chlorine dioxide (ClO2), compared to two traditional agents (H2O2, CHX) on PDLSCs and (ii) to test whether the aging factor of PDLSC cultures determines cellular responsiveness to the chemicals tested. METHODS: Impedimetry (concentration-response study), WST-1 assays (WST = water soluble tetrazolium salt), and morphology analysis were performed to measure changes in cell viability induced by the 3 disinfectants; immunocytochemistry of stem cell markers (STRO-1, CD90, and CD105) measured the induced mesenchymal characteristics. RESULTS: Cell viability experiments demonstrated that the application of ClO2 does not lead to a significant decrease in viability of PLDSCs in concentrations used to kill microbes. On the contrary, traditional irrigants, H2O2, and CHX are highly toxic on PDLSCs. Aging of PLDSC cultures (passages 3 vs. 7) has characteristic effects on their responsiveness to these agents as the increased expression of mesenchymal stem cell markers turns to decreased. CONCLUSIONS AND CLINICAL RELEVANCE: While the active ingredients of mouthwash (H2O2, CHX) applied in endodontic or periodontitis management have a serious toxic effect on PDLSCs, the novel hyperpure ClO2 is less toxic providing an environment favoring dental structure regenerations during disinfectant interventions.

In vitro functional characterization of biosimilar therapeutic antibodies.

The key factor in successful development and marketing of biosimilar antibodies is a deep understanding of their critical quality attributes and the ability to control them. Comprehensive functional characterization is therefore at the heart of the process and is a crucial part of regulatory requirements. Establishment of a scientifically sound molecule-specific functional in vitro assay panel requires diligent planning and high flexibility in order to respond to both regulatory requirements and the ever-changing demands relevant to the different stages of the development and production process. Relevance of the chosen assays to the in vivo mechanism of action is of key importance to the stepwise evidence-based demonstration of biosimilarity. Use of a sound interdisciplinary approach and orthogonal state-of-the-art techniques is also unavoidable for gaining in-depth understanding of the biosimilar candidate. The aim of the present review is to give a snapshot on the methodic landscape as depicted by the available literature discussing the in vitro techniques used for the functional characterization of approved biosimilar therapeutic antibodies. Emerging hot topics of the field and relevant structure-function relationships are also highlighted.

The Chlamydomonas deg1c Mutant Accumulates Proteins Involved in High Light Acclimation.

Degradation of periplasmic proteins (Deg)/high temperature requirement A (HtrA) proteases are ATP-independent Ser endopeptidases that perform key aspects of protein quality control in all domains of life. Here, we characterized Chlamydomonas reinhardtii DEG1C, which together with DEG1A and DEG1B is orthologous to Arabidopsis (Arabidopsis thaliana) Deg1 in the thylakoid lumen. We show that DEG1C is localized to the stroma and the periphery of thylakoid membranes. Purified DEG1C exhibited high proteolytic activity against unfolded model substrates and its activity increased with temperature and pH. DEG1C forms monomers, trimers, and hexamers that are in dynamic equilibrium. DEG1C protein levels increased upon nitrogen, sulfur, and phosphorus starvation; under heat, oxidative, and high light stress; and when Sec-mediated protein translocation was impaired. DEG1C depletion was not associated with any obvious aberrant phenotypes under nonstress conditions, high light exposure, or heat stress. However, quantitative shotgun proteomics revealed differences in the abundance of 307 proteins between a deg1c knock-out mutant and the wild type under nonstress conditions. Among the 115 upregulated proteins are PSII biogenesis factors, FtsH proteases, and proteins normally involved in high light responses, including the carbon dioxide concentrating mechanism, photorespiration, antioxidant defense, and photoprotection. We propose that the lack of DEG1C activity leads to a physiological state of the cells resembling that induced by high light intensities and therefore triggers high light protection responses.

OX40 stimulation and PD-L1 blockade synergistically augment HBV-specific CD4 T cells in patients with HBeAg-negative infection.

BACKGROUND & AIMS: Current antiviral therapies lack the potential to eliminate persistent hepatitis B virus (HBV) infection. HBV-specific T cells are crucial for HBV control and have recently been shown to be protective in patients following discontinuation of antiviral therapy. Thus, T cell-based approaches may greatly improve the therapeutic landscape of HBV infection. We aimed to augment HBV-specific CD4 T cells from chronically infected patients by targeting different immunological pathways. METHODS: Expression of various co-stimulatory and inhibitory receptors on HBV- and influenza-specific CD4 T cells was analyzed directly ex vivo by MHC class II-tetramers. Patients infected with HBV genotype D were screened for CD4 T cell responses by IFN-γ ELISpot and intracellular cytokine staining following stimulation with overlapping peptides (OLPs) spanning the HBV-polyprotein. Stimulation with recombinant IL-7, an agonistic OX40-antibody or blockade of PD-L1 was performed in antigen-specific in vitro cultures. Cytokine secretion and expression of transcription factors were analyzed by flow cytometry. Responses targeting influenza, Epstein-Barr virus and tetanus toxoid served as controls. RESULTS: Tetramer-staining revealed that the IL-7 receptor-alpha (CD127), OX40 and PD-1 constitute possible therapeutic targets as they were all strongly expressed on HBV-specific CD4 T cells ex vivo. The HBV-specific CD4 T cell responses identified by OLP screening targeted predominantly the HBV-polymerase and core proteins. Combined OX40 stimulation and PD-L1 blockade significantly augmented IFN-γ and IL-21 producing HBV-specific CD4 T cells in vitro, suggesting active T helper type 1 cell and follicular T helper cell programs. Indeed, transcription factors T-bet and Bcl6 were strongly expressed in cytokine-producing cells. CONCLUSIONS: Combined OX40 stimulation and PD-L1 blockade augmented secretion of the helper T cell signature cytokines IFN-γ and IL-21, suggesting that immunotherapeutic approaches can improve HBV-specific CD4 T cell responses. LAY SUMMARY: CD4 T cells are important in controlling viral infections but are impaired in the context of chronic hepatitis B virus (HBV) infection. Therapeutic approaches to cure chronic HBV infection are highly likely to require an immune-stimulatory component. This study demonstrates that HBV-specific CD4 T cells can be functionally augmented by combined stimulation of the co-stimulatory molecule OX40 and blockade of the inhibitory PD-1 pathway.

Can the unemployed be trained to care for the elderly? The effects of subsidized training in elderly care.

Demographic change has increased the need for elderly care. Training unemployed workers might be one way to increase the supply of elderly care nurses. This study analyzes the effectiveness of subsidized training for unemployed individuals in the elderly care professions in Germany over 11.5 years. We find that short further training and long retraining courses significantly increase workers' long-term employment. As approximately 25% to 50% of trained nurses have permanent jobs in the care sector, we estimate that approximately 5% of all employed nurses are formerly trained unemployed workers.

Immunological cure of HBV infection.

Chronic hepatitis B virus (HBV) infection is a major global health burden and cure is rarely achieved by current antiviral therapies. Therefore, there is an urgent need for new therapeutic options. Immune modulation with the goal to restore dysfunctional HBV-specific immunity is an interesting target for new therapeutic strategies. Based on the current evidence on immunology in resolving and persistent HBV infection, we will review the growing field of immunotherapeutic approaches for treatment of chronic HBV infection. We will also discuss the challenge of a heterogeneous patient population and personalized treatment as a possible key factor of success.

Detection Methods for Autoantibodies in Suspected Autoimmune Encephalitis.

This review provides an overview on different antibody test methods that can be applied in cases of suspected paraneoplastic neurological syndromes (PNS) and anti-neuronal autoimmune encephalitis (AIE) in order to explain their diagnostic value, describe potential pitfalls and limitations, and discuss novel approaches aimed at discovering further autoantibodies. Onconeuronal antibodies are well-established biomarkers for PNS and may serve as specific tumor markers. The recommended procedure to detect onconeuronal antibodies is a combination of indirect immunohistochemistry on fixed rodent cerebellum and confirmation of the specificity by line assays. Simplification of this approach by only using line assays with recombinant proteins bears the risk to miss antibody-positive samples. Anti-neuronal surface antibodies are sensitive and specific biomarkers for AIE. Their identification requires the use of test methods that allow the recognition of conformation dependent epitopes. These commonly include cell-based assays and tissue based assays with unfixed rodent brain tissue. Tissue based assays can detect most of the currently known neuronal surface antibodies and thus enable broad screening of biological samples. A complementary testing on live neuronal cell cultures may confirm that the antibody recognizes a surface epitope. In patients with peripheral neuropathy, the screening may be expanded to teased nerve fibers to identify antibodies against the node of Ranvier. This method helps to identify a novel subgroup of peripheral autoimmune neuropathies, resulting in improved immunotherapy of these patients. Tissue based assays are useful to discover additional autoantibody targets that play a role in diverse autoimmune neurological syndromes. Antibody screening assays represent promising avenues of research to improve the diagnostic yield of current assays for antibody-associated autoimmune encephalitis.

A novel hydrogel scaffold for periodontal ligament stem cells.

Periodontal ligament stem cells (PDLSCs) possess extensive regeneration potential. However, their therapeutic application demands a scaffold with appropriate properties. HydroMatrix (HydM) is a novel injectable peptide nanofiber hydrogel developed recently for cell culture. Our aim was to test whether HydM would be a suitable scaffold for proliferation and osteogenic differentiation of PDLSCs. PDLSCs were seeded on non-coated or HydM-coated surfaces. Both real-time impedance analysis and cell viability assay documented cell growth on HydM. PDLSCs showed healthy, fibroblast-like morphology on the hydrogel. After a 3-week-long culture in osteogenic medium, mineralization was much more intense in HydM cultures compared to control. Alkaline phosphatase activity of the cells grown on the gels reached the non-coated control levels. Our data provided evidence that PDLSCs can adhere, survive, migrate, and proliferate on HydM and this gel also supports their osteogenic differentiation. We first applied impedimetry for dental stem cells cultured on a scaffold. HydM is ideal for in vitro studies of PDLSCs. It may also serve not only as a reference material but also in the future as a promising biocompatible scaffold for preclinical studies.

Infants' Processing of Prosodic Cues: Electrophysiological Evidence for Boundary Perception beyond Pause Detection.

Infants as young as six months are sensitive to prosodic phrase boundaries marked by three acoustic cues: pitch change, final lengthening, and pause. Behavioral studies suggest that a language-specific weighting of these cues develops during the first year of life; recent work on German revealed that eight-month-olds, unlike six-month-olds, are capable of perceiving a prosodic boundary on the basis of pitch change and final lengthening only. The present study uses Event-Related Potentials (ERPs) to investigate the neuro-cognitive development of prosodic cue perception in German-learning infants. In adults' ERPs, prosodic boundary perception is clearly reflected by the so-called Closure Positive Shift (CPS). To date, there is mixed evidence on whether an infant CPS exists that signals early prosodic cue perception, or whether the CPS emerges only later-the latter implying that infantile brain responses to prosodic boundaries reflect acoustic, low-level pause detection. We presented six- and eight-month-olds with stimuli containing either no boundary cues, only a pitch cue, or a combination of both pitch change and final lengthening. For both age groups, responses to the former two conditions did not differ, while brain responses to prosodic boundaries cued by pitch change and final lengthening showed a positivity that we interpret as a CPS-like infant ERP component. This hints at an early sensitivity to prosodic boundaries that cannot exclusively be based on pause detection. Instead, infants' brain responses indicate an early ability to exploit subtle, relational prosodic cues in speech perception-presumably even earlier than could be concluded from previous behavioral results.

Treatment of acute rhinitis with a nasal spray containing tramazoline and essential oils: a multicenter, uncontrolled, observational trial.

BACKGROUND: In this observational trial, data were collected on the effectiveness and tolerability/safety of a nasal spray containing tramazoline and essential oils (trade name Rhinospray(®) Plus) used for symptomatic treatment of acute rhinitis due to common cold. METHODS: The trial was performed in 300 children, adolescents and adults, who were to be treated with Rhinospray(®) Plus for up to 4 times per day for up to 10 days. Primary endpoints were the change from baseline to final visit in the mean of three single symptom scores (blocked nose, sneezing, and runny nose) and the mean improvement in two quality-of-life parameters (ability to perform normal daytime activities and quality of sleep). RESULTS: A total of 108 children, 30 adolescents and 162 adults were treated with Rhinospray(®) Plus. No patient discontinued prematurely. There was a mean reduction of 2.0 ± 0.6 (standard deviation) in nasal symptom scores from baseline to final visit; 297 of 300 of patients (99.0 %) reported an improvement. The mean value for improvement in quality-of-life parameters was 1.3 ± 0.5. Improvement in daytime activities was reported by all 300 patients (100.0 %) and in quality of sleep by 292 patients (97.4 %). Effectiveness and tolerability were rated as 'very good' or 'good' by 95.4 % and 97.4 % of patients, respectively; the investigators rated effectiveness and tolerability as 'very good' or 'good' for 97.4 % and 100.0 % of patients, respectively. No adverse events were reported. CONCLUSIONS: Community-based patients reported a relief in acute rhinitis symptoms and improvement in quality of life as a result of treatment with Rhinospray(®) Plus. Treatment was well-tolerated.

On the influence of typicality and age of acquisition on semantic processing: Diverging evidence from behavioural and ERP responses.

Various behavioural studies show that semantic typicality (TYP) and age of acquisition (AOA) of a specific word influence processing time and accuracy during the performance of lexical-semantic tasks. This study examines the influence of TYP and AOA on semantic processing at behavioural (response times and accuracy data) and electrophysiological levels using an auditory category-member-verification task. Reaction time data reveal independent TYP and AOA effects, while in the accuracy data and the event-related potentials predominantly effects of TYP can be found. The present study thus confirms previous findings and extends evidence found in the visual modality to the auditory modality. A modality-independent influence on semantic word processing is manifested. However, with regard to the influence of AOA, the diverging results raise questions on the origin of AOA effects as well as on the interpretation of offline and online data. Hence, results will be discussed against the background of recent theories on N400 correlates in semantic processing. In addition, an argument in favour of a complementary use of research techniques will be made.