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Purpose: The influence of new SARS-CoV-2 variants on the post-COVID-19 condition (PCC) remains unanswered. Therefore, we examined the prevalence and predictors of PCC-related symptoms in patients infected with the SARS-CoV-2 variants delta or omicron., Methods: We compared prevalences and risk factors of acute and PCC-related symptoms three months after primary infection (3MFU) between delta- and omicron-infected patients from the Cross-Sectoral Platform of the German National Pandemic Cohort Network. Health-related quality of life (HrQoL) was determined by the EQ-5D-5L index score and trend groups were calculated to describe changes of HrQoL between different time points., Results: We considered 758 patients for our analysis (delta: n = 341; omicron: n = 417). Compared with omicron patients, delta patients had a similar prevalence of PCC at the 3MFU (p = 0.354), whereby fatigue occurred most frequently (n = 256, 34%). HrQoL was comparable between the groups with the lowest EQ-5D-5L index score (0.75, 95% CI 0.73-0.78) at disease onset. While most patients (69%, n = 348) never showed a declined HrQoL, it deteriorated substantially in 37 patients (7%) from the acute phase to the 3MFU of which 27 were infected with omicron., Conclusion: With quality-controlled data from a multicenter cohort, we showed that PCC is an equally common challenge for patients infected with the SARS-CoV-2 variants delta and omicron at least for the German population. Developing the EQ-5D-5L index score trend groups showed that over two thirds of patients did not experience any restrictions in their HrQoL due to or after the SARS-CoV-2 infection at the 3MFU., Clinical Trail Registration: The cohort is registered at ClinicalTrials.gov since February 24, 2021 (Identifier: NCT04768998)., Competing Interests: Declarations. Competing interests: The authors declare no competing interests. Conflict of interest: The authors have no competing interests to declare that are relevant to the content of this article. Ethical approval: This study was performed in line with the principles of the Declaration of Helsinki. For the NAPKON-SUEP, a primary ethics vote was obtained at the Ethics Committee of the Department of Medicine at Goethe University Frankfurt (local ethics ID approval 20-924). All further study sites received their local ethics votes at the respective ethics commissions. The NAPKON-SUEP is registered at ClinicalTrials.gov (Identifier: NCT04768998). Approval for this study was granted by the Ethics Committee of the Department of Medicine at Goethe University Frankfurt (local ethics ID approval 2021-350). Consent to participate: Written informed consent was obtained from all individual participants included in the study., (© 2024. The Author(s).)

Purpose: To analyze the longitudinal epidemiology and antimicrobial resistance (AMR) patterns of Gram-negative bloodstream infections (BSI) in Germany., Methods: Post-hoc analysis of prospectively documented BSI due to Escherichia coli, Klebsiella spp., Enterobacter spp., Pseudomonas aeruginosa and Acinetobacter baumannii from six university hospitals between 2016 and 2020. In a subanalysis 1228 episodes of BSI (E. coli N = 914, Klebsiella spp. N = 314) were analyzed for clinical endpoints and risk factors., Results: E. coli was the most prevalent cause of BSI, with 5412 cases, followed by Klebsiella spp. (2148 cases), P. aeruginosa (789 cases), Enterobacter spp. (696 cases), and A. baumannii (31 cases). BSI incidence rates were particularly high in haematology/oncology, with E. coli BSI reaching 13.9 per 1000 admissions. Most (58%) of the BSI episodes were community-acquired. A notable finding was the moderate increase of third-generation cephalosporin resistant Enterobacterales (3GCREB) for E. coli from 13.9% in 2016 to 14.4% in 2020 and a decrease for Klebsiella spp. from 16.5% in 2016 to 11.1% in 2020 corresponding to extended-spectrum betalactamase (ESBL) phenotype. In our analysis, the 3GCREB phenotype was not associated with a higher risk of death or discharge with sequelae for E. coli and Klebsiella spp., Conclusion: Our study provides longitudinal data on Gram-negative BSI in Germany on a clinical basis for the first time. These data underscores the critical need for ongoing surveillance and more pathogen-related clinical data., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Purpose: To analyse recent epidemiological trends of bloodstream infections (BSI) caused by Enterococcus spp. In adult patients admitted to tertiary care centres in Germany., Methods: Epidemiological data from the multicentre R-NET study was analysed. Patients presenting with E. faecium or E. faecalis in blood cultures in six German tertiary care university hospitals between October 2016 and June 2020 were prospectively evaluated. In vancomycin-resistant enterococci (VRE), the presence of vanA/vanB was confirmed via molecular methods., Results: In the 4-year study period, 3001 patients with BSI due to Enterococcus spp. were identified. E. faecium was detected in 1830 patients (61%) and E. faecalis in 1229 patients (41%). Most BSI occurred in (sub-) specialties of internal medicine. The pooled incidence density of enterococcal BSI increased significantly (4.0-4.5 cases per 10,000 patient days), which was primarily driven by VRE BSI (0.5 to 1.0 cases per 10,000 patient days). In 2020, the proportion of VRE BSI was > 12% in all study sites (range, 12.8-32.2%). Molecular detection of resistance in 363 VRE isolates showed a predominance of the vanB gene (77.1%)., Conclusion: This large multicentre study highlights an increase of BSI due to E. faecium, which was primarily driven by VRE. The high rates of hospital- and ICU-acquired VRE BSI point towards an important role of prior antibiotic exposure and invasive procedures as risk factors. Due to limited treatment options and high mortality rates of VRE BSI, the increasing incidence of VRE BSI is of major concern., (© 2024. The Author(s).)

Several months after COVID-19 many individuals still report persisting symptoms, the so-called 'post-COVID-19 syndrome'. An immunological dysfunction is one of the main pathophysiological hypotheses. As sleep is central to the functioning of the immune system, we investigated whether self-reported pre-existing sleep disturbance might be an independent risk factor for the development of post-COVID-19 syndrome. A total of 11,710 participants of a cross-sectional survey (all tested positive for severe acute respiratory syndrome coronavirus-2) were classified into probable post-COVID-19 syndrome, an intermediate group, and unaffected participants at an average of 8.5 months after infection. The case definition was based on newly occurring symptoms of at least moderate severity and ≥20% reduction in health status and/or working capacity. Unadjusted and adjusted odds ratios were calculated to investigate the association between pre-existing sleep disturbances and subsequent development of post-COVID-19 syndrome while controlling for a variety of demographic, lifestyle, and health factors. Pre-existing sleep disturbances were found to be an independent predictor of subsequent probable post-COVID-19 syndrome (adjusted odds ratio 2.7, 95% confidence interval 2.27-3.24). Sleep disturbances as part of the post-COVID-19 syndrome were reported by more than half of the participants and appeared to be a new symptom and to occur independent of a mood disorder in most cases. Recognition of disturbed sleep as an important risk factor for post-COVID-19 syndrome should promote improved clinical management of sleep disorders in the context of COVID-19. Further, it may stimulate further research on the effect of improving sleep on the prognosis of COVID-19 long-term sequelae and other post-viral conditions., (© 2023 The Authors. Journal of Sleep Research published by John Wiley & Sons Ltd on behalf of European Sleep Research Society.)

Background: SARS-CoV-2 infection and associated COVID-19 disease can lead to critical illness with a risk of developing a multiple organ failure. Subsequently, this may lead to various pathological sequelae, such as secondary sclerosing cholangitis after surviving COVID-19 (SSC-COVID)., Objective: The aim is to retrospectively analyze a cohort of hospitalized patients with first-wave (February 2020-June 2020) SARS-CoV-2 infection and persisting unclear cholangiopathy to determine the incidence of SSC-COVID and its risk factors., Results: A total of 249 patients were hospitalized at the university hospital in Tübingen, Germany, with SARS-CoV-2 infection during the first wave of the pandemic. Of these, 35.3% (88/249) required intensive care treatment; 16.5% (41/249) of them died due to the complications of COVID-19; 30.8% (64/208) of surviving patients could be followed up und were retrospectively analyzed at our center. The incidence of confirmed SSC-COVID was 7.8% (5/64). All SSC-COVID patients had an ICU stay >20 days, for invasive ventilation, positioning treatment, vasopressor treatment, but possible risk factors for SSC were not significant due to the small number of patients., Conclusions: SSC-COVID is an emerging disease in post-COVID patients with a high incidence in our single-center cohort. SSC-COVID should be considered as a differential diagnosis, if unclear cholangiopathy or cholestasis persists after SARS-CoV-2 infection.

Background: Antimicrobial stewardship (AMS) programmes are established across the world to treat infections efficiently, prioritize patient safety, and reduce the emergence of antimicrobial resistance. One of the core elements of AMS programmes is guidance to support and direct physicians in making efficient, safe and optimal decisions when prescribing antibiotics. To optimize and tailor AMS, we need a better understanding of prescribing physicians' experience with AMS guidance., Objectives: To explore the prescribing physicians' user experience, needs and targeted improvements of AMS guidance in hospital settings., Methods: Semi-structured interviews were conducted with 36 prescribing physicians/AMS guidance users from hospital settings in Canada, Germany, Israel, Latvia, Norway and Sweden as a part of the international PILGRIM trial. A socioecological model was applied as an overarching conceptual framework for the study., Results: Research participants were seeking more AMS guidance than is currently available to them. The most important aspects and targets for improvement of AMS guidance were: (i) quality of guidelines; (ii) availability of infectious diseases specialists; and (iii) suitability of AMS guidance to department context., Conclusions: Achieving prudent antibiotic use not only depends on individual and collective levels of commitment to follow AMS guidance but also on the quality, availability and suitability of the guidance itself. More substantial commitment from stakeholders is needed to allocate the required resources for delivering high-quality, available and relevant AMS guidance to make sure that the prescribers' AMS needs are met., (© The Author(s) 2024. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Competing Interests: Die Autorinnen/Autoren geben an, dass kein Interessenkonflikt besteht.

Objectives: Evidence on the work-related societal impact of long-term health-related consequences following SARS-CoV-2 is emerging. We characterize the modified work ability index (mWAI) of employees 6 to 12 months after an acute infection compared to pre-infection., Methods: Analyses were based on a population-based, multi-center cross-sectional study including employees aged 18-65 years with positive SARS-CoV-2 polymerase chain reaction (tested between October 2020-April 2021 in defined geographic regions in Germany). Prevalences and results of adjusted logistic regression analyses were given., Results: In 9752 employees (mean age 45.6 years, 58% females, response 24%), n = 1217 (13.1%) participants were regarded as having low mWAI compared to pre-infection. Outpatient medical treatment, inpatient treatment, and admission to intensive care during infection were associated with mWAI <15 th percentile (P15, each odds ratio [OR] >3.0). Post-COVID symptom clusters most strongly linked to mWAI

Objectives: Staphylococcus aureus bloodstream infection (SAB) is a common and severe infection. This study aims to describe temporal trends in numbers, epidemiological characteristics, clinical manifestations, and outcomes of SAB., Methods: We performed a post-hoc analysis of three prospective SAB cohorts at the University Medical Centre Freiburg between 2006 and 2019. We validated our findings in a large German multi-centre cohort of five tertiary care centres (R-Net consortium, 2017-2019). Time-dependent trends were estimated using Poisson or beta regression models., Results: We included 1797 patients in the mono-centric and 2336 patients in the multi-centric analysis. Overall, we observed an increasing number of SAB cases over 14 years (6.4%/year and 1000 patient days, 95% CI: 5.1% to 7.7%), paralleled by an increase in the proportion of community-acquired SAB (4.9%/year [95% CI: 2.1% to 7.8%]) and a decrease in the rate of methicillin-resistant-SAB (-8.5%/year [95% CI: -11.2% to -5.6%]). All of these findings were confirmed in the multi-centre validation cohort (6.2% cases per 1000 patient cases/year [95% CI: -0.6% to 12.6%], community-acquired-SAB 8.7% [95% CI: -1.2% to 19.6%], methicillin-resistant S. aureus-SAB -18.6% [95% CI: -30.6 to -5.8%]). Moreover, we found an increasing proportion of patients with multiple risk factors for complicated/difficult-to-treat SAB (8.5%/year, 95% CI: 3.6% to 13.5%, p < 0.001), alongside an overall higher level of comorbidities (Charlson comorbidity score 0.23 points/year, 95% CI: 0.09 to 0.37, p 0.005). At the same time, the rate of deep-seated foci such as osteomyelitis or deep-seated abscesses significantly increased (6.7%, 95% CI: 3.9% to 9.6%, p < 0.001). A reduction of in-hospital mortality by 0.6% per year (95% CI: 0.08% to 1%) was observed in the subgroup of patients with infectious diseases consultations., Discussion: We found an increasing number of SAB combined with a significant increase in comorbidities and complicating factors in tertiary care centres. The resulting challenges in securing adequate SAB management in the face of high patient turnover will become an important task for physicians., (Copyright © 2023. Published by Elsevier Ltd.)

Background: Deep metabolomic, proteomic and immunologic phenotyping of patients suffering from an infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have matched a wide diversity of clinical symptoms with potential biomarkers for coronavirus disease 2019 (COVID-19). Several studies have described the role of small as well as complex molecules such as metabolites, cytokines, chemokines and lipoproteins during infection and in recovered patients. In fact, after an acute SARS-CoV-2 viral infection almost 10-20% of patients experience persistent symptoms post 12 weeks of recovery defined as long-term COVID-19 syndrome (LTCS) or long post-acute COVID-19 syndrome (PACS). Emerging evidence revealed that a dysregulated immune system and persisting inflammation could be one of the key drivers of LTCS. However, how these biomolecules altogether govern pathophysiology is largely underexplored. Thus, a clear understanding of how these parameters within an integrated fashion could predict the disease course would help to stratify LTCS patients from acute COVID-19 or recovered patients. This could even allow to elucidation of a potential mechanistic role of these biomolecules during the disease course., Methods: This study comprised subjects with acute COVID-19 (n=7; longitudinal), LTCS (n=33), Recov (n=12), and no history of positive testing (n=73). 1 H-NMR-based metabolomics with IVDr standard operating procedures verified and phenotyped all blood samples by quantifying 38 metabolites and 112 lipoprotein properties. Univariate and multivariate statistics identified NMR-based and cytokine changes., Results: Here, we report on an integrated analysis of serum/plasma by NMR spectroscopy and flow cytometry-based cytokines/chemokines quantification in LTCS patients. We identified that in LTCS patients lactate and pyruvate were significantly different from either healthy controls (HC) or acute COVID-19 patients. Subsequently, correlation analysis in LTCS group only among cytokines and amino acids revealed that histidine and glutamine were uniquely attributed mainly with pro-inflammatory cytokines. Of note, triglycerides and several lipoproteins (apolipoproteins Apo-A1 and A2) in LTCS patients demonstrate COVID-19-like alterations compared with HC. Interestingly, LTCS and acute COVID-19 samples were distinguished mostly by their phenylalanine, 3-hydroxybutyrate (3-HB) and glucose concentrations, illustrating an imbalanced energy metabolism. Most of the cytokines and chemokines were present at low levels in LTCS patients compared with HC except for IL-18 chemokine, which tended to be higher in LTCS patients., Conclusion: The identification of these persisting plasma metabolites, lipoprotein and inflammation alterations will help to better stratify LTCS patients from other diseases and could help to predict ongoing severity of LTCS patients., Competing Interests: CC and HS are employed by Bruker BioSpin GmbH but were not involved in study design and analysis of the present data. Their contribution consisted in providing age- and sex-matched healthy control data and IVDr software. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Berezhnoy, Bissinger, Liu, Cannet, Schäfer, Kienzle, Bitzer, Häberle, Göpel, Trautwein and Singh.)

Background: Coronavirus disease 2019 (COVID-19) is an infection which can affect the central nervous system. In this study, we sought to investigate associations between neuroimaging findings with clinical, demographic, blood and cerebrospinal fluid (CSF) parameters, pre-existing conditions and the severity of acute COVID-19., Materials and Methods: Retrospective multicenter data retrieval from 10 university medical centers in Germany, Switzerland and Austria between February 2020 and September 2021. We included patients with COVID-19, acute neurological symptoms and cranial imaging. We collected demographics, neurological symptoms, COVID-19 severity, results of cranial imaging, blood and CSF parameters during the hospital stay., Results: 442 patients could be included. COVID-19 severity was mild in 124 (28.1%) patients (moderate n = 134/30.3%, severe n = 43/9.7%, critical n = 141/31.9%). 220 patients (49.8%) presented with respiratory symptoms, 167 (37.8%) presented with neurological symptoms first. Acute ischemic stroke (AIS) was detected in 70 (15.8%), intracranial hemorrhage (IH) in 48 (10.9%) patients. Typical risk factors were associated with AIS; extracorporeal membrane oxygenation therapy and invasive ventilation with IH. No association was found between the severity of COVID-19 or blood/CSF parameters and the occurrence of AIS or IH., Discussion: AIS was the most common finding on cranial imaging. IH was more prevalent than expected but a less common finding than AIS. Patients with IH had a distinct clinical profile compared to patients with AIS. There was no association between AIS or IH and the severity of COVID-19. A considerable proportion of patients presented with neurological symptoms first. Laboratory parameters have limited value as a screening tool., (© 2023. The Author(s).)

Background: The rapid emergence of the Omicron variant and its large number of mutations led to its classification as a variant of concern (VOC) by the World Health Organization. Subsequently, Omicron evolved into distinct sublineages (eg, BA.1 and BA.2), which currently represent the majority of global infections. Initial studies of the neutralizing response toward BA.1 in convalescent and vaccinated individuals showed a substantial reduction., Methods: We assessed antibody (immunoglobulin G [IgG]) binding, ACE2 (angiotensin-converting enzyme 2) binding inhibition, and IgG binding dynamics for the Omicron BA.1 and BA.2 variants compared to a panel of VOCs/variants of interest, in a large cohort (N = 352) of convalescent, vaccinated, and infected and subsequently vaccinated individuals., Results: While Omicron was capable of efficiently binding to ACE2, antibodies elicited by infection or immunization showed reduced binding capacities and ACE2 binding inhibition compared to wild type. Whereas BA.1 exhibited less IgG binding compared to BA.2, BA.2 showed reduced inhibition of ACE2 binding. Among vaccinated samples, antibody binding to Omicron only improved after administration of a third dose., Conclusions: Omicron BA.1 and BA.2 can still efficiently bind to ACE2, while vaccine/infection-derived antibodies can bind to Omicron. The extent of the mutations within both variants prevents a strong inhibitory binding response. As a result, both Omicron variants are able to evade control by preexisting antibodies., Competing Interests: Potential conflicts of interest. N. S. M. was a speaker at previous Luminex user meetings. The NMI is involved in applied research projects as a fee for services with the Luminex Corporation. M. Bi. reports payment or honoraria from MSD Sharp & Dohme GmbH for symposia; and also reports participation on advisory boards for Roche Pharma AG, Incyte Biosciences Germany GmbH, Bayer Vital GmbH, Bristol-Myers Squibb GmbH & Co KgaA, and MSD Sharp & Dohme GmbH. C. E. reports support for the present manuscript from MWK Sonderfördermaßnahme Kinderstudie (Kap. 1499 TG 93). B. L. reports receiving funding for the present manuscript from NaFOUniMedCovid19 (FKZ: 01KX2021) supported by the German Federal Ministry of Education and Research; and reports a leadership or fiduciary role for the German Center for Infection Research (TI BBD, DZIF), Transplant Cohort, and Steering Committee TBNet. A. Z. reports state technology funding for device infrastructure (7-4332.62-NMI/55), outside the conduct of this study. N. S.-M. reports support for the present manuscript from LAND BW (MULTICOV-AB and LAND BW, Automation in SARS-CoV-2) and payment or honoraria from Luminex Corporation for being a speaker at previous user meetings (the NMI is also involved in applied research projects as a fee for services with the Luminex Corporation). All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2022. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Objective: The aim of this study was to determine the risk of post-acute sequelae of COVID-19 associated with the continuous spectrum of BMI., Methods: Epidemiology of Long COVID (EPILOC) is a population-based study conducted in Baden-Württemberg (Germany), including subjects aged 18 to 65 years who tested positive for SARS-CoV-2 between October 2020 and April 2021. Eligible subjects answered a standardized questionnaire, including sociodemographic characteristics, lifestyle factors, and the presence of specific symptoms. Participants assessed their current general health recovery and working capacity compared with the pre-infection situation and provided their body height and weight. Generalized additive models were used to assess the association of BMI with general health recovered, working capacity recovered, and prevalence of fatigue, cognitive impairment, and chest symptoms., Results: The analyses included 11,296 individuals (41% male), with a mean age of 44.0 (SD 13.7) years. Best general health recovery was observed at BMI of 22.1 (95% CI: 21.0-27.0) kg/m 2 in men and BMI of 21.6 (95% CI: 20.3-23.1) kg/m 2 in women. In addition, we found that increasing BMI was consistently associated with post-COVID fatigue, neurocognitive impairment, and chest symptoms., Conclusions: High BMI contributes to impaired recovery after SARS-CoV-2 infection; however, a low BMI is associated with impaired recovery as well., (© 2022 The Authors. Obesity published by Wiley Periodicals LLC on behalf of The Obesity Society.)

The aim of this observational retrospective study is to improve early risk stratification of hospitalized Covid-19 patients by predicting in-hospital mortality, transfer to intensive care unit (ICU) and mechanical ventilation from electronic health record data of the first 24 h after admission. Our machine learning model predicts in-hospital mortality (AUC = 0.918), transfer to ICU (AUC = 0.821) and the need for mechanical ventilation (AUC = 0.654) from a few laboratory data of the first 24 h after admission. Models based on dichotomous features indicating whether a laboratory value exceeds or falls below a threshold perform nearly as good as models based on numerical features. We devise completely data-driven and interpretable machine-learning models for the prediction of in-hospital mortality, transfer to ICU and mechanical ventilation for hospitalized Covid-19 patients within 24 h after admission. Numerical values of. CRP and blood sugar and dichotomous indicators for increased partial thromboplastin time (PTT) and glutamic oxaloacetic transaminase (GOT) are amongst the best predictors., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

Strategic and standardised approaches to analysis and reporting of surveillance data are essential to inform antimicrobial resistance (AMR) mitigation measures, including antibiotic policies. Targeted guidance on linking full-scale AMR and antimicrobial consumption (AMC)/antimicrobial residues (AR) surveillance data from the human, animal, and environmental sectors is currently needed. This paper describes the initiative whereby a multidisciplinary panel of experts (56 from 20 countries-52 high income, 4 upper middle or lower income), representing all three sectors, elaborated proposals for structuring and reporting full-scale AMR and AMC/AR surveillance data across the three sectors. An evidence-supported, modified Delphi approach was adopted to reach consensus among the experts for dissemination frequency, language, and overall structure of reporting; core elements and metrics for AMC/AR data; core elements and metrics for AMR data. The recommendations can support multisectoral national and regional plans on antimicrobials policy to reduce resistance rates applying a One Health approach., Competing Interests: JA received grants/contracts from the Spanish Ministry of Agriculture, Food and Fisheries and EU Horizon 2020. Work of BPG was funded by the German Center for Infection research Clinical Research Unit (DZIF-CRU) at Tübingen. RD participated on Data Safety Monitoring Board of the ASTARTÉ study (for which no payments were received). WG received grants/contracts from the Horizon Europe grant (supported by UKRI) on AMR and pathogen evolution in costal environments and the UK Natural Environment Research Council grants on AMR Knowledge Exchange NE/V019279/1 and AMR evolution NE/W006251/1; WG received consultation fee for EU DG Sante AMR policy evaluation and recommendations. Work of AK was funded in the context of the project One Health EJP, which has received funding from the European Union's Horizon 2020 research and innovation programme under Grant Agreement No. 773830. EP received grants/contracts from the Austrian Ministry of Health for the National Surveillance Network of healthcare-associated infections (ANISS); EP received honorarium for participation as a chair of the Advisory board on AMR and MDRO Pfizer Austria (27.09.22). MB received grants/contracts from Janssen Vaccines, Novartis, CureVac, and Merck; MB received payment or honoraria for lectures, presentations, speakers bureaus, manuscript writing or educational events from Takeda (November 2019); MB participated on Data Safety Monitoring Board or Advisory Board of Sanofi, Spherecydes, Pfizer, Merck, Novartis, and Astra-Zeneca. MM received support for attending meetings and/or travel from the Global Antibiotic Research and Development Partnership (12.10.22–13.10.2022). LS received grants or contracts from the JPIAMR network grant 2020; LS received support for attending meetings and/or travel from ESCMID for the attendance of ECCMID 2022. DT is an employee of GlaxoSmithKline and holds shares in GlaxoSmithKline. TVB received consultation fees from Stonehaven Consulting; TVB received payment or honoraria for lectures, presentations, speakers bureaus, manuscript writing or educational events from Swedish Veterinary Council. Work of AV was supported by the COMBACTE-MAGNET consortium; AV serves as the ISAC president (International Society of Antimicrobial Chemotherapy) and board member of NVMM (Dutch Microbiology Society). All other authors declare no competing interests., (© 2022 The Authors.)

Several risk scores were developed during the COVID-19 pandemic to identify patients at risk for critical illness as a basic step to personalizing medicine even in pandemic circumstances. However, the generalizability of these scores with regard to different populations, clinical settings, healthcare systems, and new epidemiological circumstances is unknown. The aim of our study was to compare the predictive validity of qSOFA, CRB65, NEWS, COVID-GRAM, and 4C-Mortality score. In a monocentric retrospective cohort, consecutively hospitalized adults with COVID-19 from February 2020 to June 2021 were included; risk scores at admission were calculated. The area under the receiver operating characteristic curve and the area under the precision-recall curve were compared using DeLong's method and a bootstrapping approach. A total of 347 patients were included; 23.6% were admitted to the ICU, and 9.2% died in a hospital. NEWS and 4C-Score performed best for the outcomes ICU admission and in-hospital mortality. The easy-to-use bedside score NEWS has proven to identify patients at risk for critical illness, whereas the more complex COVID-19-specific scores 4C and COVID-GRAM were not superior. Decreasing mortality and ICU-admission rates affected the discriminatory ability of all scores. A further evaluation of risk assessment is needed in view of new and rapidly changing epidemiological evolution.

Objectives: To describe symptoms and symptom clusters of post-covid syndrome six to 12 months after acute infection, describe risk factors, and examine the association of symptom clusters with general health and working capacity., Design: Population based, cross sectional study SETTING: Adults aged 18-65 years with confirmed SARS-CoV-2 infection between October 2020 and March 2021 notified to health authorities in four geographically defined regions in southern Germany., Participants: 50 457 patients were invited to participate in the study, of whom 12 053 (24%) responded and 11 710 (58.8% (n=6881) female; mean age 44.1 years; 3.6% (412/11 602) previously admitted with covid-19; mean follow-up time 8.5 months) could be included in the analyses., Main Outcome Measures: Symptom frequencies (six to 12 months after versus before acute infection), symptom severity and clustering, risk factors, and associations with general health recovery and working capacity., Results: The symptom clusters fatigue (37.2% (4213/11 312), 95% confidence interval 36.4% to 38.1%) and neurocognitive impairment (31.3% (3561/11 361), 30.5% to 32.2%) contributed most to reduced health recovery and working capacity, but chest symptoms, anxiety/depression, headache/dizziness, and pain syndromes were also prevalent and relevant for working capacity, with some differences according to sex and age. Considering new symptoms with at least moderate impairment of daily life and ≤80% recovered general health or working capacity, the overall estimate for post-covid syndrome was 28.5% (3289/11 536, 27.7% to 29.3%) among participants or at least 6.5% (3289/50 457) in the infected adult population (assuming that all non-responders had completely recovered). The true value is likely to be between these estimates., Conclusions: Despite the limitation of a low response rate and possible selection and recall biases, this study suggests a considerable burden of self-reported post-acute symptom clusters and possible sequelae, notably fatigue and neurocognitive impairment, six to 12 months after acute SARS-CoV-2 infection, even among young and middle aged adults after mild infection, with a substantial impact on general health and working capacity., Trial Registration: German registry of clinical studies DRKS 00027012., Competing Interests: Competing interests: All authors have completed the ICMJE uniform disclosure form at https://www.icmje.org/disclosure-of-interest/ and declare: support from the Baden-Württemberg Federal State Ministry of Science and Art and the German pension fund (“Deutsche Rentenversicherung”) Baden-Württemberg for the submitted work; no financial relationships with any organisations that might have an interest in the submitted work in the previous three years; no other relationships or activities that could appear to have influenced the submitted work., (© Author(s) (or their employer(s)) 2019. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Objectives: Host genetic factors contribute to the variable severity of COVID-19. We examined genetic variants from genome-wide association studies and candidate gene association studies in a cohort of patients with COVID-19 and investigated the role of early SARS-CoV-2 strains in COVID-19 severity., Methods: This case-control study included 123 COVID-19 cases (hospitalized or ambulatory) and healthy controls from the state of Baden-Wuerttemberg, Germany. We genotyped 30 single nucleotide polymorphisms, using a custom-designed panel. Cases were also compared with the 1000 genomes project. Polygenic risk scores were constructed. SARS-CoV-2 genomes from 26 patients with COVID-19 were sequenced and compared between ambulatory and hospitalized cases, and phylogeny was reconstructed., Results: Eight variants reached nominal significance and two were significantly associated with at least one of the phenotypes "susceptibility to infection", "hospitalization", or "severity": rs73064425 in LZTFL1 (hospitalization and severity, P <0.001) and rs1024611 near CCL2 (susceptibility, including 1000 genomes project, P = 0.001). The polygenic risk score could predict hospitalization. Most (23/26, 89%) of the SARS-CoV-2 genomes were classified as B.1 lineage. No associations of SARS-CoV-2 mutations or lineages with severity were observed., Conclusion: These host genetic markers provide insights into pathogenesis and enable risk classification. Variants which reached nominal significance should be included in larger studies., Competing Interests: Conflict of interest All authors have no competing interests to declare., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Comorbidities are risk factors for development of severe coronavirus disease 2019 (COVID-19). However, the extent to which an underlying comorbidity influences the immune response to severe acute respiratory syndrome coronavirus 2 remains unknown., Objective: Our aim was to investigate the complex interrelations of comorbidities, the immune response, and patient outcome in COVID-19., Methods: We used high-throughput, high-dimensional, single-cell mapping of peripheral blood leukocytes and algorithm-guided analysis., Results: We discovered characteristic immune signatures associated not only with severe COVID-19 but also with the underlying medical condition. Different factors of the metabolic syndrome (obesity, hypertension, and diabetes) affected distinct immune populations, thereby additively increasing the immunodysregulatory effect when present in a single patient. Patients with disorders affecting the lung or heart, together with factors of metabolic syndrome, were clustered together, whereas immune disorder and chronic kidney disease displayed a distinct immune profile in COVID-19. In particular, severe acute respiratory syndrome coronavirus 2-infected patients with preexisting chronic kidney disease were characterized by the highest number of altered immune signatures of both lymphoid and myeloid immune branches. This overall major immune dysregulation could be the underlying mechanism for the estimated odds ratio of 16.3 for development of severe COVID-19 in this burdened cohort., Conclusion: The combinatorial systematic analysis of the immune signatures, comorbidities, and outcomes of patients with COVID-19 has provided the mechanistic immunologic underpinnings of comorbidity-driven patient risk and uncovered comorbidity-driven immune signatures., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

As global vaccination campaigns against SARS-CoV-2 proceed, there is particular interest in the longevity of immune protection, especially with regard to increasingly infectious virus variants. Neutralizing antibodies (Nabs) targeting the receptor binding domain (RBD) of SARS-CoV-2 are promising correlates of protective immunity and have been successfully used for prevention and therapy. As SARS-CoV-2 variants of concern (VOCs) are known to affect binding to the ACE2 receptor and by extension neutralizing activity, we developed a bead-based multiplex ACE2-RBD inhibition assay (RBDCoV-ACE2) as a highly scalable, time-, cost-, and material-saving alternative to infectious live-virus neutralization tests. By mimicking the interaction between ACE2 and the RBD, this serological multiplex assay allows the simultaneous analysis of ACE2 binding inhibition to the RBDs of all SARS-CoV-2 VOCs and variants of interest (VOIs) in a single well. Following validation against a classical virus neutralization test and comparison of performance against a commercially available assay, we analyzed 266 serum samples from 168 COVID-19 patients of varying severity. ACE2 binding inhibition was reduced for ten out of eleven variants examined compared to wild-type, especially for those displaying the E484K mutation such as VOCs beta and gamma. ACE2 binding inhibition, while highly individualistic, positively correlated with IgG levels. ACE2 binding inhibition also correlated with disease severity up to WHO grade 7, after which it reduced., (© 2022. The Author(s).)

Background: The burden of bloodstream infections remains high worldwide and cannot be confined to short-term in-hospital mortality. We aimed to develop scores to predict short-term and long-term mortality in patients with bloodstream infections., Methods: The Bloodstream Infection due to Multidrug-resistant Organisms: Multicenter Study on Risk Factors and Clinical Outcomes (BLOOMY) study is a prospective, multicentre cohort study at six German tertiary care university hospitals to develop and validate two scores assessing 14-day and 6-month mortality in patients with bloodstream infections. We excluded patients younger than 18 years or who were admitted to an ophthalmology or psychiatry ward. Microbiological, clinical, laboratory, treatment, and survival data were prospectively collected on day 0 and day 3 and then from day 7 onwards, weekly. Participants were followed up for 6 months. All patients in the derivation cohort who were alive on day 3 were included in the analysis. Predictive scores were developed using logistic regression and Cox proportional hazards models with a machine-learning approach. Validation was completed using the C statistic and predictive accuracy was assessed using sensitivity, specificity, and predictive values., Findings: Between Feb 1, 2017, and Jan 31, 2019, 2568 (61·5%) of 4179 eligible patients were recruited into the derivation cohort. The in-hospital mortality rate was 23·75% (95% CI 22·15-25·44; 610 of 2568 patients) and the 6-month mortality rate was 41·55% (39·54-43·59; 949 of 2284). The model predictors for 14-day mortality (C statistic 0·873, 95% CI 0·849-0·896) and 6-month mortality (0·807, 0·784-0·831) included age, body-mass index, platelet and leukocyte counts, C-reactive protein concentrations, malignancy (ie, comorbidity), in-hospital acquisition, and pathogen. Additional predictors were, for 14-day mortality, mental status, hypotension, and the need for mechanical ventilation on day 3 and, for 6-month mortality, focus of infection, in-hospital complications, and glomerular filtration rate at the end of treatment. The scores were validated in a cohort of 1023 patients with bloodstream infections, recruited between Oct 9, 2019, and Dec 31, 2020. The BLOOMY 14-day score showed a sensitivity of 61·32% (95% CI 51·81-70·04), a specificity of 86·36% (83·80-88·58), a positive predictive value (PPV) of 37·57% (30·70-44·99), and a negative predictive value (NPV) of 94·35% (92·42-95·80). The BLOOMY 6-month score showed a sensitivity of 69·93% (61·97-76·84), a specificity of 66·44% (61·86-70·73), a PPV of 40·82% (34·85-47·07), and a NPV of 86·97% (82·91-90·18)., Interpretation: The BLOOMY scores showed good discrimination and predictive values and could support the development of protocols to manage bloodstream infections and also help to estimate the short-term and long-term burdens of bloodstream infections., Funding: DZIF German Center for Infection Research., Translation: For the German translation of the abstract see Supplementary Materials section., Competing Interests: Declaration of interests SP received grants or contracts from DZIF and the Gesundheitsforum Baden-Württemberg; consulting fees from IDbyDNA and Illumina; and speaker honoraria from bioMerieux Germany, all outside of this study. WVK received funding from the German Federal Ministry of Education and Research (BMBF) and travel support from the European Society of Clinical Microbiology and Infectious Diseases, all outside of this study. JF was supported during this study by a grant paid to DZIF from BMBF (grant number 8032808811). Study group member NTe was supported during this study by a grant paid to DZIF from BMBF (grant number 8032808811). FH received funding during this study as an employee of the University Hospital Tübingen and received support for travel and attending meetings related to this study from DZIF. HS received funding paid to his institution from DZIF for study personnel and consumables; personal consultancy fees from Debiopharm, Entasis, MSD, Shionogi, and ThermoFisher; fees or honoraria for lectures, presentations, speakers' bureaus, manuscript writing, or educational events from Eumedica, Gilead, MSD, and Shionogi; and fees and support for attending meetings or travel from MSD, Gilead, and Shionogi. JR received funding for study personnel and support for attending regular study meetings from BMBF and DZIF. Study group member CI received grants or contracts paid to their institution by Shionogi and MSD; personal fees, consultancy fees, and payment or honoraria for lectures, presentations, speakers' bureaus, manuscript writing, or educational events from Shionogi and MSD; received consumables, study drugs, editorial assistance for manuscript development (outside of this study) from Eumedica; and was on an advisory board for MSD and Shionogi. JJV received grants from MSD, Gilead, Pfizer, Astellas Pharma, Basilea, DZIF, BMBF, German Aerospace Centre (DLR), University of Bristol, and Rigshospitalet Copenhagen; payment or honoraria for lectures, presentations, speakers' bureaus, manuscript writing, or educational events from MSD, Gilead, Pfizer, Astellas Pharma, Basilea, DZIF, University Hospital Freiburg, Congress and Communication, Academy for Infectious Medicine, University of Manchester, German Society for Infectious Diseases, Ärztekammer Nordrhein, University Hospital Aachen, Back Bay Strategies, German Society for Internal Medicine, Shionogi, Molecular Health, Netzwerk Universitätsmedizin, Janssen, and NordForsk; and consulting fees from Pfizer, Gilead, and Shionogi. MJGTV received consulting fees from MSD, SocraTec R&D, Farmak International Holding, and Gilead Sciences; payment or honoraria for lectures, presentations, speakers' bureaus, manuscript writing, or educational events from MSD, Ferring, Roche, and Pfizer; and grants or contracts from BioNTtech, Immunic Therapeutics, MSD, Seres Therapeutics, Takeda Pharmaceutical, Heel, and Roche. All other authors ans DZIF BLOOMY study group members declare no competing interests., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Background: More than 2 years into the COVID-19 pandemic, SARS-CoV-2 still impacts children's health and the management of pediatric hospitals. However, it is unclear which hygiene and infection control measures are effective and useful for pediatric hospitals. Here, we report infection control measures implemented at a tertiary care children's hospital. We evaluated frequency of SARS-CoV-2 detection in admitted patients, in-hospital transmission and infection related findings. Furthermore, we aimed to capture perspectives of health-care workers and caregivers on effectiveness and burden of infection control measures. Knowledge gained can inform management of the ongoing and future pandemics., Methods: We designed a retrospective observational study and survey at a pediatric tertiary care referral center. Local infection control measures and respective guidelines regarding COVID-19 were reviewed. Three thousand seven hundred sixteen children under 18 years were tested for SARS-CoV-2 at the University Children's Hospital Tuebingen and data on SARS-CoV-2 transmission were retrieved from internal records. Two surveys were conducted among 219 staff members and 229 caregivers., Results: Local infection control measures comprised the formation of a task force, triage, protective hygiene measures and an adaptable SARS-CoV-2 test strategy. Between January 2020 and March 2021, SARS-CoV-2 infection was detected in 37 children presenting to our hospital, 21 of these were admitted. One hospital-acquired infection occurred. About 90% of health-care staff perceived the majority of measures as effective and appropriate. However, visitor restrictions and cancellation of scheduled treatments were perceived least effective by hospital staff and as a particular burden for patients and their caregivers. Visits at the pediatric emergency department significantly decreased during the pandemic. We drafted a pandemic action plan by ranking infection control measures according to local transmission stages., Conclusions: SARS-CoV-2 infection control measures implemented in our tertiary care children's hospital were evaluated by health-care workers as mostly effective and appropriate. In particular, good communication, transparency of decision-making as well as universal masking and infection screening were assessed as successful measures of infection control management. Visitor restrictions and cancellation of routine appointments, in contrast, were perceived as a particular burden on patient care and should be avoided. An established pandemic action plan may guide children's hospitals in the future., (© 2022. The Author(s).)

Background: Since December 2019, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has posed a pandemic threat to global health. We are now in the fourth wave of this pandemic. As the pandemic developed, the requirements and therapeutic endoscopic procedures for SARS-CoV-2-positive patients underwent changes., Methods: Analysis of implications for an endoscopy unit during the first and second/third waves of the COVID-19 pandemic with a focus on COVID-19-related process changing. Addressed are number of SARS-CoV-2-positive patients and endoscopic examinations performed in patients who tested positive for SARS-CoV-2 during the various waves, adherence to scheduled examinations, rotation of staff to COVID-dedicated structures and, finally, impact of vaccination on infection rate among endoscopic staff., Results: During the first wave, 10 SARS-CoV-2-positive in-house patients underwent a total of 22 gastrointestinal (GI) endoscopic procedures. During the second and third waves, 59 GI endoscopies were performed in 38 patients. While in the first wave, GI bleeding was the main indication for endoscopy (82%), in the second and third waves the main indication for endoscopy was endoscopic insertion of deep feeding tubes (78%; p  < 0.001). During the first wave, 5 (17%) of 29 Interdisciplinary Endoscopy Unit (IEU) staff members were moved to designated COVID wards, which was not necessary during the following waves. Lack of protective clothing was critical during the first wave, but not in the later waves. Screening tests for patients and staff were widely available after the first wave, and IEU staff was vaccinated during the second wave., Conclusion: Strategies to ensure safe endoscopies with respect to preventing transmission of SARS-CoV-2 from patients to staff were effective. Organizational adjustments allowed the routine program to continue unaffected. Indications for GI endoscopies changed over time: during the first wave, GI endoscopies were performed for life-threatening indications, whereas later supportive procedures were the main indication., Competing Interests: Conflict of interest statement: The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (© The Author(s), 2022.)

Thromboembolic events are frequently reported in patients infected with the SARS-CoV-2 virus. The exact mechanisms of COVID-19-associated hypercoagulopathy, however, remain elusive. Recently, we observed that platelets (PLTs) from patients with severe COVID-19 infection express high levels of procoagulant markers, which were found to be associated with increased risk for thrombosis. In the current study, we investigated the time course as well as the mechanisms leading to procoagulant PLTs in COVID-19. Our study demonstrates the presence of PLT-reactive IgG antibodies that induce marked changes in PLTs in terms of increased inner-mitochondrial transmembrane potential (Δψ) depolarization, phosphatidylserine (PS) externalization, and P-selectin expression. The IgG-induced procoagulant PLTs and increased thrombus formation were mediated by ligation of PLT Fc-γ RIIA (FcγRIIA). In addition, contents of calcium and cyclic-adenosine-monophosphate (cAMP) in PLTs were identified to play a central role in antibody-induced procoagulant PLT formation. Most importantly, antibody-induced procoagulant events, as well as increased thrombus formation in severe COVID-19, were inhibited by Iloprost, a clinically approved therapeutic agent that increases the intracellular cAMP levels in PLTs. Our data indicate that upregulation of cAMP could be a potential therapeutic target to prevent antibody-mediated coagulopathy in COVID-19 disease., (© 2022 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Background: Corona virus disease 2019 (COVID-19) patients are at increased risk for thromboembolic events. It is unclear whether the risk for gastrointestinal (GI) bleeding is also increased., Methods: We considered 4128 COVID-19 patients enrolled in the Lean European Open Survey on SARS-CoV-2 (LEOSS) registry. The association between occurrence of GI bleeding and comorbidities as well as medication were examined. In addition, 1216 patients from COKA registry were analyzed focusing on endoscopy diagnostic findings., Results: A cumulative number of 97 patients (1.8%) with GI bleeding were identified in the LEOSS registry and COKA registry. Of 4128 patients from the LEOSS registry, 66 patients (1.6%) had a GI bleeding. The rate of GI bleeding in patients with intensive care unit (ICU) admission was 4.5%. The use of therapeutic dose of anticoagulants showed a significant association with the increased incidence of bleeding in the critical phase of disease. The Charlson comorbidity index and the COVID-19 severity index were significantly higher in the group of patients with GI bleeding than in the group of patients without GI bleeding (5.83 (SD = 2.93) vs. 3.66 (SD = 3.06), p < 0.01 and 3.26 (SD = 1.69) vs. 2.33 (SD = 1.53), p < 0.01, respectively). In the COKA registry 31 patients (2.5%) developed a GI bleeding. Of these, the source of bleeding was identified in upper GI tract in 21 patients (67.7%) with ulcer as the most frequent bleeding source (25.8%, n = 8) followed by gastroesophageal reflux (16.1%, n = 5). In three patients (9.7%) GI bleeding source was located in lower GI tract caused mainly by diverticular bleeding (6.5%, n = 2). In seven patients (22.6%) the bleeding localization remained unknown., Conclusion: Consistent with previous research, comorbidities and disease severity correlate with the incidence of GI bleeding. Also, therapeutic anticoagulation seems to be associated with a higher risk of GI bleeding. Overall, the risk of GI bleeding seems not to be increased in COVID-19 patients., (© 2021 The Authors. United European Gastroenterology Journal published by Wiley Periodicals LLC on behalf of United European Gastroenterology.)

Background and Objective: A number of studies have highlighted muscle-specific mechanisms of thermogenesis involving futile cycling of Ca 2+ driven by sarco (endo)plasmic reticulum Ca 2+ -ATPase (SERCA) and generating heat from ATP hydrolysis to be a promising strategy to counteract obesity and metabolic dysfunction. However, to the best of our knowledge, no experimental studies concerning the metabolic effects of pharmacologically targeting SERCA in human skeletal muscle cells have been reported. Thus, in the present study, we aimed to explore the effects of SERCA-activating compound, CDN1163, on energy metabolism in differentiated human skeletal muscle cells (myotubes)., Methods: In this study, we used primary myotube cultures derived from muscle biopsies of the musculus vastus lateralis and musculi interspinales from lean, healthy male donors. Energy metabolism in myotubes was studied using radioactive substrates. Oxygen consumption rate was assessed with the Seahorse XF24 bioanalyzer, whereas metabolic genes and protein expressions were determined by qPCR and immunoblotting, respectively., Results: Both acute (4 ​h) and chronic (5 days) treatment of myotubes with CDN1163 showed increased uptake and oxidation of glucose, as well as complete fatty acid oxidation in the presence of carbonyl cyanide 4-(trifluromethoxy)phenylhydrazone (FCCP). These effects were supported by measurement of oxygen consumption rate, in which the oxidative spare capacity and maximal respiration were enhanced after CDN1163-treatment. In addition, chronic treatment with CDN1163 improved cellular uptake of oleic acid (OA) and fatty acid β-oxidation. The increased OA metabolism was accompanied by enhanced mRNA-expression of carnitine palmitoyl transferase ( CPT ) 1B , pyruvate dehydrogenase kinase ( PDK ) 4, as well as increased AMP-activated protein kinase (AMPK) Thr172 phosphorylation. Moreover, following chronic CDN1163 treatment, the expression levels of stearoyl-CoA desaturase ( SCD ) 1 was decreased together with de novo lipogenesis from acetic acid and formation of diacylglycerol (DAG) from OA., Conclusion: Altogether, these results suggest that SERCA activation by CDN1163 enhances energy metabolism in human myotubes, which might be favourable in relation to disorders that are related to metabolic dysfunction such as obesity and type 2 diabetes mellitus., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Xiao-Rong Peng, Sven Göpel and Gavin O’ Mahony are employees of AstraZeneca. The other authors had no conflict of interest to disclose., (© 2021 The Authors.)

(1) Background: The aim of our study was to identify specific risk factors for fatal outcome in critically ill COVID-19 patients. (2) Methods: Our data set consisted of 840 patients enclosed in the LEOSS registry. Using lasso regression for variable selection, a multifactorial logistic regression model was fitted to the response variable survival. Specific risk factors and their odds ratios were derived. A nomogram was developed as a graphical representation of the model. (3) Results: 14 variables were identified as independent factors contributing to the risk of death for critically ill COVID-19 patients: age (OR 1.08, CI 1.06-1.10), cardiovascular disease (OR 1.64, CI 1.06-2.55), pulmonary disease (OR 1.87, CI 1.16-3.03), baseline Statin treatment (0.54, CI 0.33-0.87), oxygen saturation (unit = 1%, OR 0.94, CI 0.92-0.96), leukocytes (unit 1000/μL, OR 1.04, CI 1.01-1.07), lymphocytes (unit 100/μL, OR 0.96, CI 0.94-0.99), platelets (unit 100,000/μL, OR 0.70, CI 0.62-0.80), procalcitonin (unit ng/mL, OR 1.11, CI 1.05-1.18), kidney failure (OR 1.68, CI 1.05-2.70), congestive heart failure (OR 2.62, CI 1.11-6.21), severe liver failure (OR 4.93, CI 1.94-12.52), and a quick SOFA score of 3 (OR 1.78, CI 1.14-2.78). The nomogram graphically displays the importance of these 14 factors for mortality. (4) Conclusions: There are risk factors that are specific to the subpopulation of critically ill COVID-19 patients.

Purpose: To evaluate whether there is a change in findings of coronavirus disease 2019 patients in follow up lung ultrasound and to determine whether these findings can predict the development of severe disease., Materials and Methods: In this prospective monocentric study COVID-19 patients had standardized lung ultrasound (12 area evaluation) at day 1, 3 and 5. The primary end point was detection of pathologies and their change over time. The secondary end point was relationship between change in sonographic results and clinical outcome. Clinical outcome was assessed on development of severe disease defined as need for intensive care unit., Results: Data of 30 patients were analyzed, 26 patients with follow-up lung ultrasound. All of them showed lung pathologies with dynamic patterns. 26,7% developed severe disease tending to have an ubiquitous lung involvement in lung ultrasound. In patients with need for intensive care unit a previously developed increase in B-lines, subpleural consolidations and pleural line irregularities was more common. A statistically significant association between change in B-lines as well as change in pleural line irregularities and development of severe disease was observed (p<0,01)., Conclusion: The present study demonstrates that follow up lung ultrasound can be a powerful tool to track the evolution of disease and suggests that lung ultrasound is able to indicate an impending development of severe disease in COVID-19 patients., Competing Interests: The authors have declared that no competing interests exist.

Background: Cohorts of hospitalized COVID-19 patients have been studied in several countries since the beginning of the pandemic. So far, there is no complete survey of older patients in a German district that includes both outpatients and inpatients. In this retrospective observational cohort study, we aimed to investigate risk factors, mortality, and functional outcomes of all patients with COVID-19 aged 70 and older living in the district of Tübingen in the southwest of Germany., Methods: We retrospectively analysed all 256 patients who tested positive for SARS-CoV-2 in one of the earliest affected German districts during the first wave of the disease from February to April 2020. To ensure inclusion of all infected patients, we analysed reported data from the public health department as well as the results of a comprehensive screening intervention in all nursing homes of the district (n = 1169). Furthermore, we examined clinical data of all hospitalized patients with COVID-19 (n = 109)., Results: The all-cause mortality was 18%. Screening in nursing homes showed a point-prevalence of 4.6%. 39% of residents showed no COVID-specific symptoms according to the official definition at that time. The most important predictors of mortality were the need for inpatient treatment (odds ratio (OR): 3.95 [95%-confidence interval (CI): 2.00-7.86], p<0.001) and care needs before infection (non-hospitalized patients: OR: 3.79 [95%-CI: 1.01-14.27], p = 0.037, hospitalized patients: OR: 2.89 [95%-CI 1.21-6.92], p = 0.015). Newly emerged care needs were a relevant complication of COVID-19: 27% of previously self-sufficient patients who survived the disease were not able to return to their home environment after discharge from the hospital., Conclusion: Our findings demonstrate the importance of a differentiated view of risk groups and long-term effects within the older population. These findings should be included in the political and social debate during the ongoing pandemic to evaluate the true effect of COVID-19 on healthcare systems and individual functional status., Competing Interests: The authors have declared that no competing interests exist.

Background: In long-term care facilities (LTCFs) residents often receive inappropriate antibiotic treatment and infection prevention and control practices are frequently inadequate, thus favouring acquisition of MDR organisms. There is increasing evidence in the literature describing antimicrobial stewardship (AMS) activities in LTCFs, but practical guidance on how surveillance data should be linked with AMS activities in this setting is lacking. To bridge this gap, the JPIAMR ARCH and COMBACTE-MAGNET EPI-Net networks joined their efforts to provide practical guidance for linking surveillance data with AMS activities., Materials and Methods: Considering the three main topics [AMS leadership and accountability, antimicrobial usage (AMU) and AMS, and antimicrobial resistance (AMR) and AMS], a literature review was performed and a list of target actions was developed. Consensus on target actions was reached through a RAND-modified Delphi process involving 40 experts from 18 countries and different professional backgrounds adopting a One Health approach., Results: From the 25 documents identified, 25 target actions were retrieved and proposed for expert evaluation. The consensus process produced a practical checklist including 23 target actions, differentiating between essential and desirable targets according to clinical relevance and feasibility. Flexible proposals for AMS team composition and leadership were provided, with a strong emphasis on the need for well-defined and adequately supported roles and responsibilities. Specific antimicrobial classes, AMU metrics, pathogens and resistance patterns to be monitored are addressed. Effective reporting strategies are described., Conclusions: The proposed checklist represents a practical tool to support local AMS teams across a wide range of care delivery organization and availability of resources., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.)

Background: The outpatient setting is a key scenario for the implementation of antimicrobial stewardship (AMS) activities, considering that overconsumption of antibiotics occurs mainly outside hospitals. This publication is the result of a joint initiative by the JPIAMR ARCH and COMBACTE-MAGNET EPI-Net networks, which is aimed at formulating a set of target actions for linking surveillance data with AMS activities in the outpatient setting., Methods: A scoping review of the literature was carried out in three research areas: AMS leadership and accountability; antimicrobial usage and AMS; antimicrobial resistance and AMS. Consensus on the actions was reached through a RAND-modified Delphi process involving over 40 experts in infectious diseases, clinical microbiology, AMS, veterinary medicine or public health, from 18 low-, middle- and high-income countries., Results: Evidence was retrieved from 38 documents, and an initial 25 target actions were proposed, differentiating between essential or desirable targets according to clinical relevance, feasibility and applicability to settings and resources. In the first consultation round, preliminary agreement was reached for all targets. Further to a second review, 6 statements were re-considered and 3 were deleted, leading to a final list of 22 target actions in the form of a practical checklist., Conclusions: This White Paper is a pragmatic and flexible tool to guide the development of calibrated surveillance-based AMS interventions specific to the outpatient setting, which is characterized by substantial inter- and intra-country variability in the organization of healthcare structures, maintaining a global perspective and taking into account the feasibility of the target actions in low-resource settings., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.)

Objectives: To assess the association between country income status and national prevalence of invasive infections caused by the top-ranked bacteria on the WHO priority list: carbapenem-resistant (CR) Acinetobacter spp., Klebsiella spp. and Pseudomonas aeruginosa; third-generation cephalosporin-resistant (3GCR) Escherichia coli and Klebsiella spp.; and MRSA and vancomycin-resistant Enterococcus faecium (VR E. faecium)., Methods: Active surveillance systems providing yearly prevalence data from 2012 onwards for the selected bacteria were included. The gross national income (GNI) per capita was used as the indicator for income status of each country and was log transformed to account for non-linearity. The association between antibiotic prevalence data and GNI per capita was investigated individually for each bacterium through linear regression., Results: Surveillance data were available from 67 countries: 38 (57%) were high income, 16 (24%) upper-middle income, 11 (16%) lower-middle income and two (3%) low income countries. The regression showed significant inverse association (P<0.0001) between resistance prevalence of invasive infections and GNI per capita. The highest rate of increase per unit decrease in log GNI per capita was observed in 3GCR Klebsiella spp. (22.5%, 95% CI 18.2%-26.7%), CR Acinetobacter spp. (19.2% 95% CI 11.3%-27.1%) and 3GCR E. coli (15.3%, 95% CI 11.6%-19.1%). The rate of increase per unit decrease in log GNI per capita was lower in MRSA (9.5%, 95% CI 5.2%-13.7%)., Conclusions: The prevalence of invasive infections caused by the WHO top-ranked antibiotic-resistant bacteria is inversely associated with GNI per capita at the global level. Public health interventions designed to limit the burden of antimicrobial resistance should also consider determinants of poverty and inequality, especially in lower-middle income and low income countries., (© The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Objective: To delineate potential mechanisms for fasting hyperglucagonemia in childhood obesity by studying the associations between fasting plasma glucagon concentrations and plasma lipid parameters and fat compartments., Methods: Cross-sectional study of children and adolescents with obesity (n = 147) and lean controls (n = 43). Differences in free fatty acids (FFAs), triglycerides, insulin, and fat compartments (quantified by magnetic resonance imaging) across quartiles of fasting plasma glucagon concentration were analyzed. Differences in oral glucose tolerance test (OGTT) glucagon response was tested in high vs low FFAs, triglycerides, and insulin. Human islets of Langerhans were cultured at 5.5 mmol/L glucose and in the absence or presence of a FFA mixture with total FFA concentration of 0.5 mmol/L and glucagon secretion quantified., Results: In children with obesity, the quartile with the highest fasting glucagon had higher insulin (201 ± 174 vs 83 ± 39 pmol/L, P < .01), FFAs (383 ± 52 vs 338 ± 109 μmol/L, P = .02), triglycerides (1.5 ± 0.9 vs 1.0 ± 0.7 mmol/L, P < .01), visceral adipose tissue volume (1.9 ± 0.8 vs 1.2 ± 0.3 dm 3 , P < .001), and a higher prevalence of impaired glucose tolerance (IGT; 41% vs 8%, P = .01) than the lowest quartile. During OGTT, children with obesity and high insulin had a worse suppression of glucagon during the first 10 minutes after glucose intake. Glucagon secretion was 2.6-fold higher in islets treated with FFAs than in those not treated with FFAs., Conclusions: Hyperglucagonemia in childhood obesity is associated with hyperinsulinemia, high plasma FFAs, high plasma triglycerides, visceral adiposity, and IGT. The glucagonotropic effect of FFAs on isolated human islets provides a potential mechanism linking high fasting plasma FFAs and glucagon levels., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Objective: We aimed to estimate the proportion of postmigration HIV acquisition among HIV-positive migrants in Europe., Design: To reach HIV-positive migrants, we designed a cross-sectional study performed in HIV clinics., Methods: The study was conducted from July 2013 to July 2015 in 57 clinics (nine European countries), targeting individuals over 18 years diagnosed in the preceding 5 years and born abroad. Electronic questionnaires supplemented with clinical data were completed in any of 15 languages. Postmigration HIV acquisition was estimated through Bayesian approaches combining extensive information on migration and patients' characteristics. CD4 cell counts and HIV-RNA trajectories from seroconversion were estimated by bivariate linear mixed models fitted to natural history data. Postmigration acquisition risk factors were investigated with weighted logistic regression., Results: Of 2009 participants, 46% were MSM and a third originated from sub-Saharan Africa and Latin America & Caribbean, respectively. Median time in host countries was 8 years. Postmigration HIV acquisition was 63% (95% confidence interval: 57-67%); 72% among MSM, 58 and 51% in heterosexual men and women, respectively. Postmigration HIV acquisition was 71% for Latin America and Caribbean migrants and 45% for people from sub-Saharan Africa. Factors associated with postmigration HIV acquisition among heterosexual women and MSM were age at migration, length of stay in host country and HIV diagnosis year and among heterosexual men, length of stay in host country and HIV diagnosis year., Conclusion: A substantial proportion of HIV-positive migrants living in Europe acquired HIV postmigration. This has important implications for European public health policies.

There are concerns about central nervous system (CNS)-replication of HIV-1 in patients on boosted protease inhibitors. Purpose of this study was to compare HIV-1 viral loads (VLs) from patients treated with only boosted dual protease inhibitor (bdPI), versus combination antiretroviral therapy (cART group), containing two nucleoside analogue reverse transcriptase inhibitors (NRTI) and a third partner. All patients from a large German HIV-treatment cohort with available medication, clinical and demographic data, including results from simultaneous HIV-1 viral load (VL) assessments in cerebrospinal fluid (CSF) and blood plasma, were retrospectively evaluated as controlled cross-sectional study. CSF had been obtained from patients with variable neurological symptoms during 2005-2014. Statistical analysis comprised nonparametric tests, regression and correlation techniques accounting for undetectable quantifications. Statistical analysis comprised nonparametric tests, regression and correlation techniques accounting for undetectable quantifications. Overall, 155 patients were evaluable (bdPI: 24; cART: 131). At time of CSF-collection, both groups were comparable in age, gender, CD4-cell counts, or primary HIV-transmission risks, though bdPI patients were clinically more advanced. The proportion of patients with undetectable HIV-1 (<50 copies/ml) in CSF was lower for bdPI group (25 vs 49.6 %; p = 0.026), but similar in plasma (46 vs 41 %). Median CSF-VL was higher in bdPI group (600 vs 50 copies/ml; p = 0.027) and similar in plasma. Mean VL CSF/plasma ratio was 342.91 for bdPI- and 54.48 for cART patients (p < 0.001). Pearson's regression analysis revealed a trend for an elevated VL-ratio over time within bdPI group. HIV-1 replication was higher and more frequently detectable in CSF from bdPI patients, indicating a worse CNS penetration effectiveness of used boosted PI. Within bdPI group, measured CNS-viral replication was increasing over time, suggesting an over time impaired HIV-1 suppression in CSF.

Background: Migrants form a substantial proportion of the population affected by the human immunodeficiency virus (HIV) epidemic in Europe, yet HIV prevention for this population is hindered by poor understanding of access to care and of postmigration transmission dynamics., Objective: We present the design and methods of the advancing Migrant Access to health Services in Europe (aMASE) study, the first European cross-cultural study focused on multiple migrant populations. It aims to identify the structural, cultural, and financial barriers to HIV prevention, diagnosis, and treatment and to determine the likely country of HIV acquisition in HIV-positive migrant populations., Methods: We delivered 2 cross-sectional electronic surveys across 10 countries (Belgium, France, Germany, Greece, Italy, the Netherlands, Portugal, Spain, Switzerland, and United Kingdom). A clinic survey aimed to recruit up to 2000 HIV-positive patients from 57 HIV clinics in 9 countries. A unique study number linked anonymized questionnaire data to clinical records data (viral loads, CD4 cell counts, viral clades, etc). This questionnaire was developed by expert panel consensus and cognitively tested, and a pilot study was carried out in 2 countries. A Web-based community survey (n=1000) reached those living with HIV but not currently accessing HIV clinics, as well as HIV-negative migrants. It was developed in close collaboration with a community advisory group (CAG) made up of representatives from community organizations in 9 of the participating countries. The CAG played a key role in data collection by promoting the survey to higher-risk migrant groups (sub-Saharan Africans, Latin Americans, men who have sex with men, and people who inject drugs). The questionnaires have considerable content overlap, allowing for comparison. Questions cover ethnicity, migration, immigration status, HIV testing and treatment, health-seeking behavior, sexual risk, and drug use. The electronic questionnaires, which were available in 15 languages, allowed for complex routing, preventing respondents from answering irrelevant questions., Results: In total, we recruited 2249 participants from 57 HIV clinics as part of the clinic survey and retrieved 1637 complete responses as part of the community survey., Conclusions: The findings will provide much-needed information for improving HIV prevention interventions and access to services for migrant communities.

Introduction: Depression is a co-morbidity of clinical significance in HIV-positive patients with an estimated prevalence of more than 20%. Sex and gender-related differences in depression are well described in HIV-negative populations, demonstrating that more women are being affected. So far little is known about frequency and characteristics of depression in HIV-positive men and women., Materials and Methods: Primary objective of our prospective epidemiological study was the evaluation of the Beck score for depression in male and female patients of the Frankfurt HIV Cohort. The Beck Depression Inventory (BDI-II) is a self-report symptom inventory made up of 21 questions, each with 4 possible answers, correlating with a certain point value., Interpretation: score 14-19: mild depression; score 20-28: moderate depression; score ≥29: severe depression. Secondary objectives of the analysis were factors that might possibly influence the disposition for depression in HIV-positive patients, e.g. age, antiretroviral treatment history, co-morbidities and socioeconomic status., Results: Between January and October 2013, 348 patients were enrolled in the study, 161 women and 187 men of the Frankfurt HIV Cohort, who had a routine appointment at the HIV-Center of the University Clinic Frankfurt. The mean age of all study participants was 45 years (range 22-80). The majority of patients were on antiretroviral therapy (91%) at study entrance. The median BDI-II score in all patients was 8 (0-49); in female patients 10 (0-42), in male patients 6 (0-49), respectively (Table 1). Significant more women than men showed a score for moderate depression (p=0.006). Factors associated with a BDI-II score ≥20 in women were older age (>45 years), living alone, unemployment and the number of prior changes in antiretroviral therapy., Conclusions: Depression in people living with HIV shows sex and gender-related differences that might also influence antiretroviral treatment strategies. HIV specialists should be aware of these gender-specific aspects and consider routine screening for depression especially in female patients of older age or those with multiple therapy changes in history.

Introduction: Chip-based automated patch clamp systems are widely used in drug development and safety pharmacology, allowing for high quality, high throughput screening at standardized experimental conditions. The merits of automation generally come at the cost of large amounts of cells needed, since cells are not targeted individually, but randomly positioned onto the chip aperture from cells in suspension. While cell usage is of little concern when using standard cell lines such as CHO or HEK cells, it becomes a crucial constraint with cells of limited availability, such as primary or otherwise rare and expensive cells, like induced pluripotent stem (IPS) cell-derived cardiomyocytes or neurons., Methods: We established application protocols for CHO cells, IPS cell-derived neurons (iCell® Neurons, Cellular Dynamics International), cardiomyocytes (Cor.4U®, Axiogenesis) and pancreatic islet cells, minimizing cell usage for automated patch clamp recordings on Nanion's Patchliner. Use of 5 μl cell suspension per well for densities between 55,000 cells/ml and 400,000 cells/ml depending on cell type resulted in good cell capture., Results: We present a new cell application procedure optimized for the Patchliner achieving>80% success rates for using as little as 300 to 2000 cells per well depending on cell type. We demonstrate that this protocol works for standard cell lines, as well as for stem cell-derived neurons and cardiomyocytes, and for primary pancreatic islet cells. We present recordings for these cell types, demonstrating that high data quality is not compromised by altered cell application., Discussion: Our new cell application procedure achieves high success rates with unprecedentedly low cell numbers. Compared to other standard automated patch clamp systems we reduced the average amount of cells needed by more than 150 times. Reduced cell usage crucially improves cost efficiency for expensive cells and opens up automated patch clamp for primary cells of limited availability., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Objectives: The aim of this study was to compare 2 protocols regarding the initiation of oral nutrition in patients with mild acute pancreatitis., Methods: We randomized 143 patients to the Lipase directed (LIP) (n = 74) and the self selected PAT (n = 69) group. In the (PAT) group, the patients restarted eating through self-selection. In the LIP group, serum lipase had to normalize before eating., Results: The mean time between admission and oral nutrition was 2 days (interquartile range [IQR], 1-3) in the PAT group and 3 days (IQR, 2-4) in the LIP group (P < 0.005). Before and after the first meal, the mean Δ visual analogue scale (VAS) was +3.14 mm (±11.5 mm) in the PAT group and +2.85 mm (±16.4) in the LIP group (P = 0.597). The length of hospital stay was 7 days (median; IQR, 5-10.5) in the PAT group and 8 days (median; IQR, 5.75-12) in the LIP group (P = 0.315)., Conclusions: We were not able to demonstrate a difference in postprandial abdominal pain or in the length of hospital stay. Patients with self-selected eating, however, were able to restart eating 1 day earlier, and this difference was found to be significant. Our data suggest that normalization of serum lipase is not obligatory for enteral nutrition in mild acute pancreatitis.

Pancreatic islets have a central role in blood glucose homeostasis. In addition to insulin-producing beta-cells and glucagon-secreting alpha-cells, the islets contain somatostatin-releasing delta-cells. Somatostatin is a powerful inhibitor of insulin and glucagon secretion. It is normally secreted in response to glucose and there is evidence suggesting its release becomes perturbed in diabetes. Little is known about the control of somatostatin release. Closure of ATP-regulated K(+)-channels (K(ATP)-channels) and a depolarization-evoked increase in cytoplasmic free Ca(2+) concentration ([Ca(2+)](i)) have been proposed to be essential. Here, we report that somatostatin release evoked by high glucose (>or=10 mM) is unaffected by the K(ATP)-channel activator diazoxide and proceeds normally in K(ATP)-channel-deficient islets. Glucose-induced somatostatin secretion is instead primarily dependent on Ca(2+)-induced Ca(2+)-release (CICR). This constitutes a novel mechanism for K(ATP)-channel-independent metabolic control of pancreatic hormone secretion.

This article discusses the currently used methodologies for monitoring exocytosis as changes in cell capacitance. Details are given on composition of solutions, experimental protocols, and how the observed responses can be interpreted physiologically. The concepts are illustrated by examples from our own work on insulin-releasing pancreatic beta-cells. Finally, we consider the feasibility of applying capacitance measurements to endocrine cells in intact pancreatic islets, where the cells are electrically coupled to each other.

Capacitance measurements of exocytosis were applied to functionally identified alpha-, beta- and delta-cells in intact mouse pancreatic islets. The maximum rate of capacitance increase in beta-cells during a depolarization to 0 mV was equivalent to 14 granules s(-1), <5% of that observed in isolated beta-cells. Beta-cell secretion exhibited bell-shaped voltage dependence and peaked at +20 mV. At physiological membrane potentials (up to approximately -20 mV) the maximum rate of release was approximately 4 granules s(-1). Both exocytosis (measured by capacitance measurements) and insulin release (detected by radioimmunoassay) were strongly inhibited by the L-type Ca(2+) channel blocker nifedipine (25 microm) but only marginally (<20%) affected by the R-type Ca(2+) channel blocker SNX482 (100 nm). Exocytosis in the glucagon-producing alpha-cells peaked at +20 mV. The capacitance increases elicited by pulses to 0 mV exhibited biphasic kinetics and consisted of an initial transient (150 granules s(-1)) and a sustained late component (30 granules s(-1)). Whereas addition of the N-type Ca(2+) channel blocker omega-conotoxin GVIA (0.1 microm) inhibited glucagon secretion measured in the presence of 1 mm glucose to the same extent as an elevation of glucose to 20 mm, the L-type Ca(2+) channel blocker nifedipine (25 microm) had no effect. Thus, glucagon release during hyperglycaemic conditions depends principally on Ca(2+)-influx through N-type rather than L-type Ca(2+) channels. Exocytosis in the somatostatin-secreting delta-cells likewise exhibited two kinetically separable phases of capacitance increase and consisted of an early rapid (600 granules s(-1)) component followed by a sustained slower (60 granules s(-1)) component. We conclude that (1) capacitance measurements in intact pancreatic islets are feasible; (2) exocytosis measured in beta-cells in situ is significantly slower than that of isolated cells; and (3) the different types of islet cells exhibit distinct exocytotic features.

Glucose-stimulated insulin secretion consists of a transient first phase followed by a sustained second phase. Diabetes (type II) is associated with abnormalities in this release pattern. Here we review the evidence that biphasic insulin secretion reflects exocytosis of two functional subsets of secretory granules and the implications for diabetes.

1. The perforated patch whole-cell configuration of the patch-clamp technique was applied to superficial cells in intact pancreatic islets. Immunostaining in combination with confocal microscopy revealed that the superficial cells consisted of 35 % insulin-secreting B-cells and 65 % non-B-cells (A- and D-cells). 2. Two types of cell, with distinct electrophysiological properties, could be functionally identified. One of these generated oscillatory electrical activity when the islet was exposed to 10 mM glucose and had the electrophysiological characteristics of isolated B-cells maintained in tissue culture. 3. The Ca2+ current recorded from B-cells in situ was 80 % larger than that of isolated B-cells. It exhibited significant (70 %) inactivation during 100 ms depolarisations. The inactivation was voltage dependent and particularly prominent during depolarisations evoking the largest Ca2+ currents. 4. Voltage-dependent K+ currents were observed during depolarisations to membrane potentials above -20 mV. These currents inactivated little during a 200 ms depolarisation and were unaffected by varying the holding potential between -90 and -30 mV. 5. The maximum resting conductance in the absence of glucose, which reflects the conductance of ATP-regulated K+ (KATP) channels, amounted to approximately 4 nS. Glucose produced a concentration-dependent reduction of KATP channel conductance with half-maximal inhibition observed with 5 mM glucose. 6. Combining voltage- and current-clamp recording allowed the estimation of the gap junction conductance between different B-cells. These experiments indicated that the input conductance of the B-cell at stimulatory glucose concentrations ( approximately 1 nS) is almost entirely accounted for by coupling to neighbouring B-cells.

1. Measurements of cell capacitance were used to investigate the mechanisms by which acetylcholine (ACh) stimulates Ca2+-induced exocytosis in single insulin-secreting mouse pancreatic B-cells. 2. ACh (250 microM) increased exocytotic responses elicited by voltage-clamp depolarizations 2.3-fold. This effect was mediated by activation of muscarinic receptors and dependent on elevation of the cytoplasmic Ca2+ concentration ([Ca2+]i) attributable to mobilization of Ca2+ from intracellular stores. The latter action involved interference with the buffering of [Ca2+]i and the time constant (tau) for the recovery of [Ca2+]i following a voltage-clamp depolarization increased 5-fold. As a result, Ca2+ was present at concentrations sufficient to promote the replenishment of the readily releasable pool of granules (RRP; > 0.2 microM) for much longer periods in the presence than in the absence of the agonist. 3. The effect of Ca2+ on exocytosis was mediated by activation of CaM kinase II, but not protein kinase C, and involved both an increased size of the RRP from 40 to 140 granules and a decrease in tau for the refilling of the RRP from 31 to 19 s. 4. Collectively, the effects of ACh on the RRP and tau result in a > 10-fold stimulation of the rate at which granules are supplied for release.