Genomic epidemiology reveals multiple introductions of SARS-CoV-2 followed by community and nosocomial spread, Germany, February to May 2020.

BackgroundIn the SARS-CoV-2 pandemic, viral genomes are available at unprecedented speed, but spatio-temporal bias in genome sequence sampling precludes phylogeographical inference without additional contextual data.AimWe applied genomic epidemiology to trace SARS-CoV-2 spread on an international, national and local level, to illustrate how transmission chains can be resolved to the level of a single event and single person using integrated sequence data and spatio-temporal metadata.MethodsWe investigated 289 COVID-19 cases at a university hospital in Munich, Germany, between 29 February and 27 May 2020. Using the ARTIC protocol, we obtained near full-length viral genomes from 174 SARS-CoV-2-positive respiratory samples. Phylogenetic analyses using the Auspice software were employed in combination with anamnestic reporting of travel history, interpersonal interactions and perceived high-risk exposures among patients and healthcare workers to characterise cluster outbreaks and establish likely scenarios and timelines of transmission.ResultsWe identified multiple independent introductions in the Munich Metropolitan Region during the first weeks of the first pandemic wave, mainly by travellers returning from popular skiing areas in the Alps. In these early weeks, the rate of presumable hospital-acquired infections among patients and in particular healthcare workers was high (9.6% and 54%, respectively) and we illustrated how transmission chains can be dissected at high resolution combining virus sequences and spatio-temporal networks of human interactions.ConclusionsEarly spread of SARS-CoV-2 in Europe was catalysed by superspreading events and regional hotspots during the winter holiday season. Genomic epidemiology can be employed to trace viral spread and inform effective containment strategies.

Multicentre comparison of quantitative PCR-based assays to detect SARS-CoV-2, Germany, March 2020.

Containment strategies and clinical management of coronavirus disease (COVID-19) patients during the current pandemic depend on reliable diagnostic PCR assays for the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we compare 11 different RT-PCR test systems used in seven diagnostic laboratories in Germany in March 2020. While most assays performed well, we identified detection problems in a commonly used assay that may have resulted in false-negative test results during the first weeks of the pandemic.

Differential Pathogen-Specific Immune Reconstitution in Antiretroviral Therapy-Treated Human Immunodeficiency Virus-Infected Children.

BACKGROUND: Susceptibility to coinfections in human immunodeficiency virus (HIV)-infected patients remains increased despite antiretroviral therapy (ART). To elucidate mechanisms involved in immune reconstitution, we studied immune activation, immune exhaustion, and HIV- and copathogen-specific T-cell responses in children before and after ART. METHODS: We prospectively enrolled 25 HIV-infected children to study HIV-, cytomegalovirus (CMV)-, and tuberculosis (TB)-specific T-cell responses before and 1 year after initiation of ART using intracellular cytokine (interleukin-2, interferon-γ, tumor necrosis factor-α) staining assays after in vitro stimulation. We further measured expression of activation, immune exhaustion, and memory phenotype markers and studied proliferative responses after antigen stimulation. RESULTS: We observed differential, pathogen-specific changes after 1 year of ART in cytokine profiles of CD4 T-cell responses that were associated with shifts in memory phenotype and decreased programmed cell death 1 (PD-1) expression. The proliferative capacity of HIV- and PPD-specific responses increased after 1 year of ART. Of note, the recovery of CMV- and TB-specific responses was correlated with a decrease in PD-1 expression (r = 0.83, P = .008 and r = 0.81, P = .0007, respectively). CONCLUSIONS: Reconstitution of immune responses on ART is associated with alterations in T-cell phenotype, function, and PD-1 expression that are distinct for HIV, TB, and CMV. The PD-1 pathway represents a potential target for immunotherapy in HIV-infected patients on ART with insufficient immune reconstitution.

Acute sedation-associated complications in GI endoscopy (ProSed 2 Study): results from the prospective multicentre electronic registry of sedation-associated complications.

OBJECTIVES: Sedation has been established for GI endoscopic procedures in most countries, but it is also associated with an added risk of complications. Reported complication rates are variable due to different study methodologies and often limited sample size. DESIGNS: Acute sedation-associated complications were prospectively recorded in an electronic endoscopy documentation in 39 study centres between December 2011 and August 2014 (median inclusion period 24 months). The sedation regimen was decided by each study centre. RESULTS: A total of 368 206 endoscopies was recorded; 11% without sedation. Propofol was the dominant drug used (62% only, 22.5% in combination with midazolam). Of the sedated patients, 38 (0.01%) suffered a major complication, and overall mortality was 0.005% (n=15); minor complications occurred in 0.3%. Multivariate analysis showed the following independent risk factors for all complications: American Society of Anesthesiologists class >2 (OR 2.29) and type and duration of endoscopy. Of the sedation regimens, propofol monosedation had the lowest rate (OR 0.75) compared with midazolam (reference) and combinations (OR 1.0-1.5). Compared with primary care hospitals, tertiary referral centres had higher complication rates (OR 1.61). Notably, compared with sedation by a two-person endoscopy team (endoscopist/assistant; 53.5% of all procedures), adding another person for sedation (nurse, physician) was associated with higher complication rates (ORs 1.40-4.46), probably due to higher complexity of procedures not evident in the multivariate analysis. CONCLUSIONS: This large multicentre registry study confirmed that severe acute sedation-related complications are rare during GI endoscopy with a very low mortality. The data are useful for planning risk factor-adapted sedation management to further prevent sedation-associated complications in selected patients. TRIAL REGISTRATION NUMBER: DRKS00007768; Pre-results.

Comparison of three continuous positive airway pressure (CPAP) interfaces in healthy Beagle dogs during medetomidine-propofol constant rate infusions.

OBJECTIVE: To compare the efficacy of three continuous positive airway pressure (CPAP) interfaces in dogs on gas exchange, lung volumes, amount of leak during CPAP and rebreathing in case of equipment failure or disconnection. STUDY DESIGN: Randomized, prospective, crossover, experimental trial. ANIMALS: Ten purpose-bred Beagle dogs. METHODS: Dogs were in dorsal recumbency during medetomidine-propofol constant rate infusions, breathing room air. Three interfaces were tested in each dog in a consecutive random order: custom-made mask (M), conical face mask (FM) and helmet (H). End-expiratory lung impedance (EELI) measured by electrical impedance tomography was assessed with no interface (baseline), with the interface only (No-CPAP for 3 minutes) and at 15 minutes of 7 cmH2O CPAP (CPAP-delivery). PaO2 was assessed at No-CPAP and CPAP-delivery, partial pressure of inspired carbon dioxide (PICO2; rebreathing assessment) at No-CPAP and the interface leak (ΔPleak) at CPAP-delivery. Mixed-effects linear regression models were used for statistical analysis (p<0.05). RESULTS: During CPAP-delivery, all interfaces increased EELI by 7% (p<0.001). Higher ΔPleak was observed with M and H (9 cmH2O) in comparison with FM (1 cmH2O) (p<0.001). At No-CPAP, less rebreathing occurred with M (0.5 kPa, 4 mmHg) than with FM (1.8 kPa, 14 mmHg) and with H (1.4 kPa, 11 mmHg), but also lower PaO2 was measured with M (9.3 kPa, 70 mmHg) than with H (11.9 kPa, 90 mmHg) and FM (10.8 kPa, 81 mmHg). CONCLUSIONS AND CLINICAL RELEVANCE: All three interfaces can be used to provide adequate CPAP in dogs. The leak during CPAP-delivery and the risk of rebreathing and hypoxaemia, when CPAP is not maintained, can be significant. Therefore, animals should always be supervised during administration of CPAP with any of the three interfaces. The performance of the custom-made M was not superior to the other interfaces.

Absolute quantification of microparticles by flow cytometry in ascites of patients with decompensated cirrhosis: a cohort study.

BACKGROUND: Microparticles (MPs) are small (<1 µm) cell membrane-derived vesicles that are formed in response to cellular activation or early stages of apoptosis. Increased plasma MP levels have been associated with liver disease severity. Here we investigated the clinical impact of ascites MPs in patients with decompensated liver cirrhosis. METHODS: Ascites and blood samples of 163 patients with cirrhosis (ascites n = 163, blood n = 31) were collected between February 2011 and December 2012. MPs were obtained from ascites and from blood by two-step ultracentrifugation and quantified by flow cytometry. Quantitative absolute MP levels were correlated with clinical and laboratory baseline parameters as well as patient outcomes. Ascites microparticles were stained with antibodies against CD66b (neutrophils) and CD3 (lymphocytes) in a subgroup of 60 matched patients. RESULTS: MPs were detected in all ascites and blood samples. Absolute ascites MP levels correlated with blood levels (r = 0.444, p = 0.011). Low ascites MP levels (<488.4 MP/µL) were associated with a poor 30-day survival probability (<488.4 MP/µL 71.1% vs. >488.4 MP/µL 94.7%, log rank p = 0.001) and such patients had a higher relative amount of ascites microparticles derived from neutrophils and lymphocytes. Low levels of ascites MPs, high MELD score and antibiotic treatment were independent risk factors for death within 30 days. CONCLUSIONS: Ascites MP levels predict short-term survival along with the liver function in patients with decompensated cirrhosis. Further studies which evaluate ascites MPs as disease specific biomarker with a validation cohort and which investigate its underlying mechanisms are needed. Neutrophils and lymphocytes contributed more frequently to the release of microparticles in patients with low ascites levels, possibly indicating an immune activation in this cohort.

Early response activation in repetition priming: an LRP study.

According to recent interpretations of repetition priming, response codes are automatically bound to a stimulus and retrieved during successive presentations of the stimulus, hence, affecting its current processing. Despite a solid corpus of behavioural evidence in line with this interpretation, electrophysiological studies have reported contrasting results regarding the nature and the timing of response code retrieval. The present experiment aims to establish at which stage of information processing decision and action codes are retrieved in repetition priming. To this end, the lateralized readiness potential (LRP) was analysed for primed faces to monitor motor cortex activity related to response preparation. Congruent and incongruent responses were obtained by having identical or reversed tasks between study and test. Primed stimuli presented LRP activations with opposite polarities for the two congruency conditions in the time-window 250-300 ms, indicating response-related motor cortex activity resulting from the retrieval of correct and incorrect decision/action codes for congruent and incongruent trials, respectively. This result indicates that decision and action codes bound to a primed stimulus are retrieved at early stages of stimulus processing and that these codes are transmitted to the motor cortex.

Pleural Effusion Caused by an Unusual Mass in the Right Hemithorax.

CASE PRESENTATION: An 80-year-old woman presented with complaints of weakness and dizziness. She had a medical history of subacute cerebral ischemia, vertigo, hypertension, and thalassemia minor. The patient was born and raised in Turkey and has lived in Switzerland for 50 years. Her sister died of a mesothelioma caused by asbestos exposure at the age of 60 years but had lived in Turkey until her death. The patient had neither a history of TB nor B symptoms. She has never smoked.

An Intra-Company Analysis of Inherent Particles in Biologicals Shapes the Protein Particle Mitigation Strategy Across Development Stages.

To better understand protein aggregation and inherent particle formation in the biologics pipeline at Novartis, a cross-functional team collected and analyzed historical protein particle issues. Inherent particle occurrences from the past 10 years were systematically captured in a protein particle database. Where the root cause was identified, a number of product attributes (such as development stage, process step, or protein format) were trended. Several key themes were revealed: 1) there was a higher propensity for inherent particle formation with non-mAbs than with mAbs; 2) the majority of particles were detected following manufacturing at scale, and were not predicted by the small-scale studies; 3) most issues were related to visible particles, followed by subvisible particles; 4) 50% of the issues were manufacturing related. These learnings became the foundation of a particle mitigation strategy across development and technical transfer, and resulted in a set of preventive actions. Overall, this study provides further insight into a recognized industry challenge and hopes to inspire the biopharmaceutical industry to transparently share their experiences with inherent particles formation.

Independence of data-driven and conceptually driven priming: the case of person recognition.

A central tenet of current memory theories is the independence of data-driven and conceptually driven priming. Although these types of repetition priming have different functional properties, it has remained unclear whether they are independent and additive or interact with each other. Therefore, we orthogonally combined these priming types in two experiments requiring occupation judgments for celebrities, represented by their faces or names. In both experiments, the sum of the effects of separate data-driven priming and conceptually driven priming equaled the effect of the combination of these priming types. These findings demonstrate for the first time the independence of data-driven and conceptually driven priming and so confirm a central tenet of current memory theories. This independence may thus be an important element for the development of future overarching memory theories.

A double-blind placebo-controlled trial to study therapeutic effects of probiotic Escherichia coli Nissle 1917 in subgroups of patients with irritable bowel syndrome.

PURPOSE: To study the therapeutic effects of probiotic Escherichia coli Nissle 1917 (EcN) in irritable bowel syndrome (IBS) and identify subgroups benefiting most. BACKGROUND: Some trials investigating therapeutic effects in irritable bowel syndrome have shown benefits in IBS subgroups only. Probiotic treatment seems to be promising. METHODS: Patients with irritable bowel syndrome (120; Rome II) were recruited to a prospective double-blind study and randomized to either EcN (n = 60) or placebo (n = 60) given for 12 weeks. Objectives were to describe efficacy and safety of EcN in different groups of irritable bowel syndrome. Outcome was assessed by 'Integrative Medicine Patient Satisfaction Scale'. RESULTS: Altogether, the responder rate was higher in the EcN than in the placebo group. However, only after 10 and 11 weeks, the differences were significant (Δ 20.0% points [95% CI 2.6; 37.4], p = 0.01 and Δ 18.3% points [95% CI 1.0; 35.7], p = 0.02, respectively). The best response was observed in the subgroup of patients with gastroenteritis or antibiotics prior to irritable bowel syndrome onset (Δ 45.7% points, p = 0.029). No significant differences were observed in any other subgroup. Both treatment groups showed similar adverse events and tolerance. CONCLUSIONS: Probiotic EcN shows effects in irritable bowel syndrome, especially in patients with altered enteric microflora, e.g. after gastroenterocolitis or administration of antibiotics.

Toll-like receptor 7/8-matured RNA-transduced dendritic cells as post-remission therapy in acute myeloid leukaemia: results of a phase I trial.

OBJECTIVES: Innovative post-remission therapies are needed to eliminate residual AML cells. DC vaccination is a promising strategy to induce anti-leukaemic immune responses. METHODS: We conducted a first-in-human phase I study using TLR7/8-matured DCs transfected with RNA encoding the two AML-associated antigens WT1 and PRAME as well as CMVpp65. AML patients in CR at high risk of relapse were vaccinated 10× over 26 weeks. RESULTS: Despite heavy pretreatment, DCs of sufficient number and quality were generated from a single leukapheresis in 11/12 cases, and 10 patients were vaccinated. Administration was safe and resulted in local inflammatory responses with dense T-cell infiltration. In peripheral blood, increased antigen-specific CD8+ T cells were seen for WT1 (2/10), PRAME (4/10) and CMVpp65 (9/10). For CMVpp65, increased CD4+ T cells were detected in 4/7 patients, and an antibody response was induced in 3/7 initially seronegative patients. Median OS was not reached after 1057 days; median RFS was 1084 days. A positive correlation was observed between clinical benefit and younger age as well as mounting of antigen-specific immune responses. CONCLUSIONS: Administration of TLR7/8-matured DCs to AML patients in CR at high risk of relapse was feasible and safe and resulted in induction of antigen-specific immune responses. Clinical benefit appeared to occur more likely in patients <65 and in patients mounting an immune response. Our observations need to be validated in a larger patient cohort. We hypothesise that TLR7/8 DC vaccination strategies should be combined with hypomethylating agents or checkpoint inhibition to augment immune responses. TRIAL REGISTRATION: The study was registered at https://clinicaltrials.gov on 17 October 2012 (NCT01734304) and at https://www.clinicaltrialsregister.eu (EudraCT-Number 2010-022446-24) on 10 October 2013.

Neural manifestations of memory with and without awareness.

Neurophysiological events responsible for different types of human memory tend to occur concurrently and are therefore difficult to measure independently. To surmount this problem, we produced perceptual priming (indicated by speeded responses) in the absence of conscious remembering. At encoding, faces appeared briefly while subjects' attention was diverted to other stimuli. Faces appeared again in either an implicit or explicit memory test. Neural correlates of priming were identified as brain potentials beginning 270 ms after face onset with more negative amplitudes for repeated than for new faces. Remembered faces, in contrast, activated a different configuration of intracranial sources producing positive potentials maximal at 600-700 ms. We thus disentangled and characterized distinct neural events associated with memory with and without awareness.

Validating neural correlates of familiarity.

Familiarity is a pervasive memory phenomenon that occurs in its most basic form when someone recognizes a repeated stimulus without recollecting other aspects of the requisite prior learning episode. Theoretical controversy currently abounds with respect to both the cognitive and neural characteristics of familiarity. Here, we show that the extant data, particularly brain-potential data, are insufficient for validating putative neural correlates of familiarity, and we outline strategies for making progress on this problem. Conceptual priming is an implicit-memory phenomenon that often occurs together with familiarity; experiments that conflate the two phenomena can be misleading. Avoiding this conflation is required to understand familiarity and to determine the extent to which the neurocognitive processes that support priming also drive familiarity.

Rapid response learning of brand logo priming: Evidence that brand priming is not dominated by rapid response learning.

Repetition priming increases the accuracy and speed of responses to repeatedly processed stimuli. Repetition priming can result from two complementary sources: rapid response learning and facilitation within perceptual and conceptual networks. In conceptual classification tasks, rapid response learning dominates priming of object recognition, but it does not dominate priming of person recognition. This suggests that the relative engagement of network facilitation and rapid response learning depends on the stimulus domain. Here, we addressed the importance of the stimulus domain for rapid response learning by investigating priming in another domain, brands. In three experiments, participants performed conceptual decisions for brand logos. Strong priming was present, but it was not dominated by rapid response learning. These findings add further support to the importance of the stimulus domain for the relative importance of network facilitation and rapid response learning, and they indicate that brand priming is more similar to person recognition priming than object recognition priming, perhaps because priming of both brands and persons requires individuation.

A Newly Identified Impurity in Polysorbate 80, the Long-Chain Ketone 12-Tricosanone, Forms Visible Particles in a Biopharmaceutical Drug Product.

Visible particles linked to polysorbates (PSs) used in biopharmaceutical drug products (DPs) have been observed repeatedly in recent years as an industry-wide issue, with PS degradation and insoluble degradation products, especially fatty acids and fatty acid esters, being suspected as root cause. We have shown that the visible particles observed in a monoclonal antibody DP solution in vials after 18 months of long-term storage at 5 ± 3°C were neither linked to reduction in PS (PS80) concentration nor to any known PS degradation product, but consist of 12-tricosanone, an impurity present in the raw material PS80, not a degradation product. The occurrence of visible 12-tricosanone particles in DP correlated with the usage of specific PS80 raw material lots, where 12-tricosanone was found as impurity at elevated levels. The quantities detected in these PS80 lots directly translate into the amount found in the respective monoclonal antibody DP batches. This is the first time that a clear correlation between the occurrence of the impurity 12-tricosanone in PS80 and the occurrence of visible particles in DP batches is reported. The observation and techniques described enable the control of this ketone in PS raw materials, providing means to prevent respective visible particle formation in DP.

Determination of functional residual capacity by oxygen washin-washout: a validation study.

OBJECTIVE: To validate a new system for functional residual capacity (FRC) measurements using oxygen washin/washout in spontaneously breathing humans. The system (LUFU, Drägerwerk AG, Lübeck, Germany) consists of an unmodified EVITA 4 ventilator, a side-stream paramagnetic oxygen sensor and a dedicated software. DESIGN: Laboratory study and measurements in spontaneously breathing volunteers. SETTING: Pulmonary function laboratory of a university hospital. PARTICIPANTS: 20 healthy and 15 lung diseased volunteers. INTERVENTIONS: FRC was measured by LUFU (LUFU-FRC) and by helium dilution (He-FRC); intra-thoracic gas volume (ITGV) was determined by body plethysmography. Each measurement cycle consisted of four independent LUFU-FRC determinations (step change of FiO(2) from 0.21 to 0.5 and back and from 0.21 to 1.0 and back), two helium-dilution runs and two body box measurements. Repeatability and agreement between methods were determined by comparing different measurements of one technique and by comparing different techniques among each other. MEASUREMENTS AND RESULTS: Repeatability of LUFU-FRC was estimated by comparing washin to washout and the different FiO(2)steps. The difference of the means was 3.7% at the most. Agreement between methods resulted in the following differences (mean+/-standard deviation of differences) for healthy and lung-diseased volunteers, respectively: LUFU-FRC vs. He-FRC -0.40+/-0.50 L (0.02+/-0.95 L), LUFU-FRC vs. ITGV -0.43+/-0.54 L (-0.18+/-0.61 L) and He-FRC vs. ITGV -0.03+/-0.43 L (-0.20+/-0.98 L). CONCLUSIONS: LUFU is a non-invasive method for the determination of FRC that requires only minor additional equipment and no modification to the ventilator. It can be used in difficult conditions such as breathing patterns with variations from breath to breath. The results of this study show that LUFU is sufficiently reliable and repeatable to warrant its clinical application.

Is There a Downside of Job Accommodations? An Employee Perspective on Individual Change Processes.

By modifying the work environments, work routines, and work tasks of employees with health restrictions, organizations can effectively help them continue to perform their jobs successfully. As such, job accommodations are an effective tool to secure the continued employment of aging workers who develop disabilities across their life span. However, while accommodations tackle health-related performance problems, they might create new challenges on the part of the affected employee. Building on the organizational change and accommodations literatures, we propose a theoretical framework of negative experiences during accommodation processes and apply it to qualitative data from group interviews with 73 manufacturing workers at a German industrial company who were part of the company's job accommodation program. Although problems associated with health-related impairments were mostly solved by accommodation, affected employees with disabilities reported about interpersonal problems and conflicts similar to those that typically occur during organizational change. Lack of social support as well as poor communication and information were raised as criticisms. Furthermore, our findings indicate that discrimination, bullying, and maltreatment appear to be common during accommodation processes. To make accommodation processes more successful, we derive recommendations from the organizational change literature and apply it to the accommodation context. We also emphasize unique characteristics of the accommodation setting and translate these into practical implications.

Detection of molecular bacterascites in decompensated cirrhosis defines a risk with decreased survival.

BACKGROUND: The prognostic relevance of bacterial DNA (bactDNA) detection in ascitic fluid of patients with cirrhosis is still under debate. Using quantitative real-time PCR with broad-range primers targeting the V3 and V4 variable region of the 16S rRNA gene, we measured bactDNA concentrations in patients with and without leukocytic ascites and evaluated the impact on short-term survival. PATIENTS AND METHODS: Ascites samples from 173 patients with decompensated cirrhosis were consecutively collected between February 2011 and December 2012. BactDNA-positive ascites samples were sequenced and chromatograms were identified using RipSeq. Clinical data collection and survival analyses were carried out retrospectively and correlated with ascites bactDNA levels. RESULTS: BactDNA was detected qualitatively with a similar frequency in both nonleukocytic and leukocytic ascites [40% (57/144) and 43.5% (10/23), respectively; P=0.724]. However, the median bactDNA level was significantly higher in leukocytic ascites than in nonleukocytic ascites (1.2×10 vs. 5.7×10 copies/ml; P=0.008). Patients' survival was associated significantly with bactDNA level. The 30-day and 180-day survival was reduced if bactDNA was above the quantification limit of 520 copies/ml (84 and 63% vs. 72 and 43%, respectively; P<0.05) and worst if bactDNA was above 5000 copies/ml. The bacterial spectrum was dominated by Gram-positive strains as shown by direct sequencing. CONCLUSION: BactDNA quantification in ascitic fluid samples using culture-independent 16S rRNA gene-based methods seems to be an interesting approach to identify patients at risk of reduced survival. Our study warrants further evaluation of antibiotic treatment in patients with molecular bacterascites.

Neural correlates of intentional and incidental recognition of famous faces.

Event-related potentials (ERPs) were used to study the relationship between intentional and incidental recognition of famous faces. Intentional and incidental recognition were operationally defined as repeated presentations of targets and nontargets within a modified Sternberg task. These repetitions elicited temporally and topographically distinct ERP modulations. A repetition effect around 300 ms (ERE/N250r) and a preceding modulation did not differ between intentional and incidental recognition, whereas a following repetition effect (LRE/N400) around 500 ms showed differences between incidental and intentional recognition. These results show that during the first few hundred milliseconds intentional and incidental face recognition relate to similar processing, indicating that familiar faces are recognized even when their identification is not required.