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The aim of this study was to evaluate the safety and efficacy of a modified CAL-WR.The use of segmental colectomy in patients with endoscopically unresectable colonic lesions results in significant morbidity and mortality. CAL-WR is an alternative procedure that may reduce morbidity.This prospective multicenter study was performed in 13 Dutch hospitals between January 2017 and December 2019. Inclusion criteria were (1) colonic lesions inaccessible using current endoscopic resection techniques (judged by an expert panel), (2) non-lifting residual/recurrent adenomatous tissue after previous polypectomy or (3) an undetermined resection margin after endoscopic removal of a low-risk pathological T1 (pT1) colon carcinoma. Thirty-day morbidity, technical success rate and radicality were evaluated.Of the 118 patients included (56% male, mean age 66 years, standard deviation ± 8 years), 66 (56%) had complex lesions unsuitable for endoscopic removal, 34 (29%) had non-lifting residual/recurrent adenoma after previous polypectomy and 18 (15%) had uncertain resection margins after polypectomy of a pT1 colon carcinoma. CAL-WR was technically successful in 93% and R0 resection was achieved in 91% of patients. Minor complications (Clavien-Dindo i-ii) were noted in 7 patients (6%) and an additional oncologic segmental resection was performed in 12 cases (11%). Residual tissue at the scar was observed in 5% of patients during endoscopic follow-up.CAL-WR is an effective, organ-preserving approach that results in minor complications and circumvents the need for major surgery. CAL-WR, therefore, deserves consideration when endoscopic excision of circumscribed lesions is impossible or incomplete.

Objective: The goal of this review was to assess the relationships among aims, designs, and outcomes of integrated inpatient medical and psychiatric care units (IMPUs) and gather the evidence base on the effectiveness of these units. Methods: Using online searches of Embase, Medline, Web of Science, PsycINFO, Scopus, CINAHL, Cochrane, and Google Scholar, the authors identified and reviewed literature describing the aims and outcomes of specific IMPU designs. Results: The search yielded 55 studies, in which the authors identified 39 IMPUs that focused on patients with mood, psychotic, somatic symptom, substance use, organic, and personality disorders and a broad array of medical diagnoses. Most units were psychiatric-medical units and had medium medical and psychiatric acuity capabilities. The studies reviewed provided little information on the cost-effectiveness of various IMPU designs. Although some comparative studies indicated reductions in hospital length of stay (LOS), these studies were generally of low quality and rarely reported other intended outcomes. Conclusions: IMPUs may help shorten LOS. IMPUs should focus care on patients with complex conditions and high acuity to maximize health system value. Implementing compulsory admission facilities; qualified psychiatric, medical, and nursing staff involvement; and cross-disciplinary training may improve IMPUs’ capacity to treat high-acuity patients. Future research should relate IMPU designs to intended outcomes.

Background: Active patient involvement in treatment decisions is seen as a feature of patient-centred care that will ultimately lead to better healthcare services and patient outcomes. Although many factors have been identified that influence patient involvement in treatment decisions, little is known about the different views that patients have on which factors are most important. Objective: This study explores the views of patients with a chronic condition on factors influencing their involvement in treatment decisions. Design: Q-methodology was used to study the views of patients. Respondents were asked to rank a set of 42 statements from the least important to the most important for active patient involvement in treatment decision-making. The set of 42 statements was developed based on a literature search and a pilot in which two external researchers, 15 patients and four healthcare professionals participated. A total of 136 patients with one of three major chronic conditions were included: diabetes types 1 and 2, respiratory disease (i.e., chronic obstructive pulmonary disease and asthma) and cancer (i.e., breast cancer and prostate cancer). Data were collected in a face-to-face interview setting in the Netherlands. Results: Four distinct views on the factors influencing active patient involvement were identified among patients with a chronic condition. (1) Enabled involvement: the extent to which patients are facilitated and empowered to participate will lead to patient involvement. (2) Relationship-driven involvement: the relationship between patients and healthcare professionals drives patient involvement. (3) Disease impact-driven involvement: the severity of disease drives patient involvement. (4) Cognition-driven involvement: knowledge and information drive patient involvement. Discussion and Conclusion: From the patients' perspective, this study shows that there is no one-size-fits-all approach to involving patients more actively in their healthcare journey. Strategies aiming to enhance active patient involvement among patients with a chronic condition should consider this diversity in perspectives among these patients. Patient Contribution: Patients are the respondents as this study researches their perspective on factors influencing patient involvement. In addition, patients were involved in pilot-testing the statement set.

Older persons are vulnerable to depression SFduring the ageing process. Financial resources and social participation are expected to have an impact on depressive symptoms. This study investigated the relationship between financial support from children and depression among Chinese older persons, as well as the mediating effect of social participation in this relationship. Data from 7163 participants aged 60 and above were extracted from wave 2015 and 2018 of the China Health and Retirement Longitudinal Survey (CHARLS). A multivariate regression analysis was performed on both cross-sectional data and two-wave longitudinal data to test our hypotheses. The results revealed that financial support from children was negatively associated with depressive symptoms in both the short-term and the long-term. In addition, this relationship was partially mediated by social participation in the short-term association and fully mediated by social participation in the long-term, where financial support was positively related to social participation, and social participation was negatively associated with depressive symptoms. This study offers an in-depth insight into the relationship between financial support from children and depression among Chinese older persons. Policies and initiatives to stimulate social participation should be promoted to improve older persons’ mental health.

Heparan sulfate proteoglycans (HSPGs) mediate essential interactions throughout the extracellular matrix (ECM), providing signals that regulate cellular growth and development. Altered HSPG composition during tumorigenesis strongly aids cancer progression. Heparanase (HPSE) is the principal enzyme responsible for extracellular heparan sulfate catabolism and is markedly up-regulated in aggressive cancers. HPSE overactivity degrades HSPGs within the ECM, facilitating metastatic dissemination and releasing mitogens that drive cellular proliferation. Reducing extracellular HPSE activity reduces cancer growth, but few effective inhibitors are known, and none are clinically approved. Inspired by the natural glycosidase inhibitor cyclophellitol, we developed nanomolar mechanism-based, irreversible HPSE inhibitors that are effective within physiological environments. Application of cyclophellitol-derived HPSE inhibitors reduces cancer aggression in cellulo and significantly ameliorates murine metastasis. Mechanism-based irreversible HPSE inhibition is an unexplored anticancer strategy. We demonstrate the feasibility of such compounds to control pathological HPSE-driven malignancies.

Until the 1980s, institutional elder care was virtually unknown in China. In a few decades, China had to construct a universal social safety net and assure basic elderly care. China’s government has been facing several challenges: the eroding traditional family care, the funding to assure care services for the older population, as well as the shortage of care delivery services and nursing staff. This paper examines China’s Five-Year Policy Plans from 1994 to 2020. Our narrative review analysis focuses on six main topics revealed in these policies: care infrastructure, community involvement, home-based care, filial piety, active aging and elder industry. Based on this analysis, we identified several successive and often simultaneously strategic steps that China introduced to contend with the aging challenge. In Western countries, elder care policies have been shifting to the home care approach. China introduced home care as the elder care cornerstone and encouraged the revival of the filial piety tradition. Although China has a unique approach, the care policies for the aged population in China and Western countries are converging by emphasizing home-based care, informal care and healthy aging.

Comparative Study Haemophilus influenzae is an opportunistic pathogen adapted to the human respiratory tract. Non-typeable H. influenzae are highly heterogeneous, but few studies have analysed the genomic variability of capsulated strains. This study aims to examine the genetic diversity of 37 serotype f isolates from the Netherlands, Portugal, and Spain, and to compare all capsulated genomes available on public databases. Serotype f isolates belonged to CC124 and shared few single nucleotide polymorphisms (SNPs) (n = 10,999), but a high core genome (> 80%). Three main clades were identified by the presence of 75, 60 and 41 exclusive genes for each clade, respectively. Multi-locus sequence type analysis of all capsulated genomes revealed a reduced number of clonal complexes associated with each serotype. Pangenome analysis showed a large pool of genes (n = 6360), many of which were accessory genome (n = 5323). Phylogenetic analysis revealed that serotypes a, b, and f had greater diversity. The total number of SNPs in serotype f was significantly lower than in serotypes a, b, and e (p < 0.0001), indicating low variability within the serotype f clonal complexes. Capsulated H. influenzae are genetically homogeneous, with few lineages in each serotype. Serotype f has high genetic stability regardless of time and country of isolation. Tis study was funded by Instituto de Salud Carlos III (ISCIII) through the Projects from the Fondo de Investigaciones Sanitarias “PI16/00977” to SM, and CIBER de Enfermedades Respiratorias (CIBERES–CB06/06/0037), co-funded by the European Regional Development Fund/European Social Fund (ERDF/ESF, “Investing in your future”), and CERCA Programme/Generalitat de Catalunya for institutional support. Bioinformatic analysis was supported by an Amazon Web Services (AWS) research grant to SM. AC was supported by FPU grant “FPU16/02202” (Formación de Profesorado Universitario, Ministerio de Educación, Spain), and SM was supported by Miguel Servet contract “CP19/00096” (ISCIII). info:eu-repo/semantics/publishedVersion

Complement C5 inhibitor eculizumab has a great impact on the treatment of patients with paroxysmal nocturnal hemoglobinuria (PNH). However, this treatment success has a major drawback: a substantially increased susceptibility for life-threatening Neisseria meningitidis infections. Therefore, N meningitidis vaccination is strongly advised before initiating complement C5–blocking therapy. In this study, we show that the multicomponent N meningitidis serogroup B (4CMenB) vaccination of PNH patients treated with eculizumab results in a significant increase in anti–N meningitidis serogroup B (MenB) plasma immunoglobulin G (IgG) levels. Anti-MenB IgG was able to bind to the bacterial surface and initiate complement activation; however, inhibition of the membrane attack complex formation completely blocked whole blood–mediated killing of MenB. This would suggest that, despite 4CMenB vaccination, PNH patients taking C5 inhibitors are not sufficiently protected against MenB infection, which is in line with the fact that vaccinated PNH patients still experience meningococcal infections.

Contains fulltext : 282502.pdf (Publisher’s version ) (Open Access) The Streptococcus pneumoniae capsule is regarded as indispensable in bacteremia. We report an infant with a ventricular septal defect and infective endocarditis caused by nontypeable S. pneumoniae. In-depth investigation confirmed a deficient capsule yet favored pneumococcal fitness for causing infective endocarditis, rather than a host immune disorder, as the cause of infective endocarditis in this case.

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Background Nasopharyngeal colonisation with clinically relevant bacterial pathogens is a risk factor for severe infections, such as pneumonia and bacteraemia. In this study, we investigated the determinants of nasopharyngeal carriage in febrile patients in rural Burkina Faso. Methods From March 2016 to June 2017, we recruited 924 paediatric and adult patients presenting with fever, hypothermia or suspicion of severe infection to the Centre Medical avec Antenne Chirurgicale Saint Camille de Nanoro, Burkina Faso. We recorded a broad range of clinical data, collected nasopharyngeal swabs and tested them for the presence of Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus and Klebsiella pneumoniae by quantitative polymerase chain reaction. Using logistic regression, we investigated the determinants of carriage and aimed to find correlations with clinical outcome. Results Nasopharyngeal colonisation with S. pneumoniae, H. influenzae and M. catarrhalis was highly prevalent and strongly dependent on age and season. Females were less likely to be colonised with S. pneumoniae (OR 0.71, p = 0.022, 95% CI 0.53–0.95) and M. catarrhalis (OR 0.73, p = 0.044, 95% CI 0.54–0.99) than males. Colonisation rates were highest in the age groups K. pneumoniae prevalence was low and not significantly correlated with age or season. For S. pneumoniae and H. influenzae, we found a positive association between nasopharyngeal carriage and clinical pneumonia [OR 1.75, p = 0.008, 95% CI 1.16–2.63 (S. pneumoniae) and OR 1.90, p = 0.004, 95% CI 1.23–2.92 (H. influenzae)]. S. aureus carriage was correlated with mortality (OR 4.01, p Conclusions Age, sex and season are important determinants of nasopharyngeal colonisation with S. pneumoniae, H. influenzae and M. catarrhalis in patients with fever in Burkina Faso. S. pneumoniae and H. influenzae carriage is associated with clinical pneumonia and S. aureus carriage is associated with mortality in patients with fever. These findings may help to understand the dynamics of colonisation and the associated transmission of these pathogens. Furthermore, understanding the determinants of nasopharyngeal colonisation and the association with disease could potentially improve the diagnosis of febrile patients.

Contains fulltext : 282357.pdf (Publisher’s version ) (Open Access) The nasopharyngeal commensal Streptococcus pneumoniae can become invasive and cause metastatic infection. This requires the pneumococcus to have the ability to adapt, grow, and reside in diverse host environments. Therefore, we studied whether the likelihood of severe disease manifestations was related to pneumococcal growth kinetics. For 383 S. pneumoniae blood isolates and 25 experimental mutants, we observed highly reproducible growth curves in nutrient-rich medium. The derived growth features were lag time, maximum growth rate, maximum density, and stationary-phase time before lysis. First, the pathogenicity of each growth feature was probed by comparing isolates from patients with and without marked preexisting comorbidity. Then, growth features were related to the propensity of causing severe manifestations of invasive pneumococcal disease (IPD). A high maximum bacterial density was the most pronounced pathogenic growth feature, which was also an independent predictor of 30-day mortality (P = 0.03). Serotypes with an epidemiologically higher propensity for causing meningitis displayed a relatively high maximum density (P

Whereas proteolytic cleavage is crucial for peptide presentation by classical major histocompatibility complex (MHC) proteins to T cells, glycolipids presented by CD1 molecules are typically presented in an unmodified form. However, the mycobacterial lipid antigen mannosyl-β1-phosphomycoketide (MPM) may be processed through hydrolysis in antigen presenting cells, forming mannose and phosphomycoketide (PM). To further test the hypothesis that some lipid antigens are processed, and to generate antigens that lead to defined epitopes for future tuberculosis vaccines or diagnostic tests, we aimed to create hydrolysis-resistant MPM variants that retain their antigenicity. Here, we designed and tested three different, versatile synthetic strategies to chemically stabilize MPM analogs. Crystallographic studies of CD1c complexes with these three new MPM analogs showed anchoring of the lipid tail and phosphate group that is highly comparable to nature-identical MPM, with considerable conformational flexibility for the mannose head group. MPM-3, a difluoromethylene-modified version of MPM that is resistant to hydrolysis showed altered recognition by cells, but not by CD1c proteins, supporting the cellular antigen processing hypothesis. Furthermore, the synthetic analogs elicited T cell responses that were cross-reactive with nature-identical MPM, fulfilling important requirements for future clinical use.

Dutch policy stipulates that people with dementia should remain at home for as long as possible. If they need care, they must preferably appeal to family, friends and neighbours. Professional help and nursing homes are deemed last resorts. Therefore, case managers must coproduce their public services increasingly in healthcare triads with both people with dementia (PWDs) and their informal caregivers. Case managers are professionals who provide and coordinate care and support for PWDs and their informal caregivers during the entire trajectory from (suspected) diagnosis until institutionalisation. The literature on coproduction has focused on the bilateral interactions between service providers and users rather than the multilateral collaborative relationships through which many public services are currently delivered, as is the case in dementia care. Little is known about how frontline workers, case managers in this study, handle conflicts in these healthcare triads. Our study addresses this gap in the coproduction literature and explores the action strategies case managers use to handle conflicts. We interviewed 19 Dutch case managers and observed 10 of their home visits between January and May 2017. We focused on the end stage of dementia at home, just before admission to a nursing home, as we assumed that most conflicts occur in that phase. The findings reveal that the case managers use a variety of action strategies to resolve and intervene in these conflicts. Their initial strategies are in line with the ideals underlying coproduction; however, their successive strategies abandon those ideals and are more focused on production or result from their own lack of power. We also found that current reforms create new dilemmas for case managers. Future research should focus on the boundaries of coproducing public services in triadic relationships and the effects of current welfare reforms aimed at coproducing public services in healthcare triads.

The highly lipophilic outer barrier of mycobacteria, such as M. tuberculosis and M. leprae, is key to their virulence and intrinsic antibiotic resistance. Various components of this mycomembrane interact with the host immune system but many of these interactions remain ill-understood. This review covers several chemical syntheses of one of these components, mycobacterial phenolic glycolipids (PGLs), and outlines the interaction of these PGLs with the human immune system, as established using these well-defined pure compounds.

Introduction We investigated the impact of the Corona Virus Disease 2019 (COVID-19) pandemic and the resulting lockdown on reperfusion treatments and door-to-treatment times during the first surge in Dutch comprehensive stroke centers. Furthermore, we studied the association between COVID-19-status and treatment times. Methods We included all patients receiving reperfusion treatment in 17 Dutch stroke centers from May 11th, 2017, until May 11th, 2020. We collected baseline characteristics, National Institutes of Health Stroke Scale (NIHSS) at admission, onset-to-door time (ODT), door-to-needle time (DNT), door-to-groin time (DGT) and COVID-19-status at admission. Parameters during the lockdown (March 15th, 2020 until May 11th, 2020) were compared with those in the same period in 2019, and between groups stratified by COVID-19-status. We used nationwide data and extrapolated our findings to the increasing trend of EVT numbers since May 2017. Results A decline of 14% was seen in reperfusion treatments during lockdown, with a decline in both IVT and EVT delivery. DGT increased by 12 min (50 to 62 min, p-value of Conclusions During the first surge of the COVID-19 pandemic, a decline in acute reperfusion treatments and a delay in DGT was seen, which indicates a target for attention. It also appeared that COVID-19-positive or -suspected patients had more severe neurologic symptoms, whereas their EVT-workflow was not affected.

Complement deficient patients are susceptible to rare meningococcal serogroups. A 6-year-old girl presented with serogroup Z meningitis. This led to identification of a C8 deficiency. The MenB-4C vaccine induced cross-reactive antibodies to serogroup Z and increased in vitro opsonophagocytic killing and may thus protect complement deficient patients.

Wall teichoic acids (WTAs) are important components of the cell wall of the opportunistic Gram‐positive bacterium Staphylococcus aureus. WTAs are composed of repeating ribitol phosphate (RboP) residues that are decorated with d‐alanine and N‐acetyl‐d‐glucosamine (GlcNAc) modifications, in a seemingly random manner. These WTA‐modifications play an important role in shaping the interactions of WTA with the host immune system. Due to the structural heterogeneity of WTAs, it is impossible to isolate pure and well‐defined WTA molecules from bacterial sources. Therefore, here synthetic chemistry to assemble a broad library of WTA‐fragments, incorporating all possible glycosylation modifications (α‐GlcNAc at the RboP C4; β‐GlcNAc at the RboP C4; β‐GlcNAc at the RboP C3) described for S. aureus WTAs, is reported. DNA‐type chemistry, employing ribitol phosphoramidite building blocks, protected with a dimethoxy trityl group, was used to efficiently generate a library of WTA‐hexamers. Automated solid phase syntheses were used to assemble a WTA‐dodecamer and glycosylated WTA‐hexamer. The synthetic fragments have been fully characterized and diagnostic signals were identified to discriminate the different glycosylation patterns. The different glycosylated WTA‐fragments were used to probe binding of monoclonal antibodies using WTA‐functionalized magnetic beads, revealing the binding specificity of these WTA‐specific antibodies and the importance of the specific location of the GlcNAc modifications on the WTA‐chains., By using tailor‐made teichoic acid building blocks, a library of glycosylated ribitol phosphate Staphylococcus aureus wall teichoic acids (WTA) has been assembled, ranging in length, and varying in N‐acetyl glucosamine substitution pattern. The library of well‐defined WTAs has been used to develop a magnetic bead binding assay to interrogate monoclonal antibody binding, revealing the preference of the various antibodies for different glycosylation patterns.

Simultaneous triggering of Toll-like receptors (TLRs) and NOD-like receptors (NLRs) has previously been shown to synergistically activate monocytes, dendritic cells and macrophages. We applied these properties in a T-cell vaccine setting by conjugating the NOD2-ligand muramyl-dipeptide (MDP) and TLR2-ligand Pam3CSK4 to a synthetic peptide derived from a model antigen. Stimulation of human DCs with the MDP-peptide-Pam3CSK4 conjugate led to a strongly increased secretion of pro-inflammatory and Th1-type cytokines and chemokines. We further show that the conjugated ligands retain their ability to trigger their respective receptors, while even improving NOD2-triggering. Also, activation of murine DCs was enhanced by the dual triggering, ultimately leading to effective induction of vaccine-specific T cells expressing IFNγ, IL-2 and TNFα. Together, these data indicate that the dual MDP-SLP-Pam3CSK4 conjugate constitutes a chemically well-defined vaccine approach that holds promise for the use in the treatment of virus infections and cancer.

Gaucher disease is caused by inherited deficiency in glucocerebrosidase (GBA, a retaining β-glucosidase), and deficiency in GBA constitutes the largest known genetic risk factor for Parkinson's disease. In the past, animal models of Gaucher disease have been generated by treatment with the mechanism-based GBA inhibitors, conduritol B epoxide (CBE), and cyclophellitol. Both compounds, however, also target other retaining glycosidases, rendering generation and interpretation of such chemical knockout models complicated. Here we demonstrate that cyclophellitol derivatives carrying a bulky hydrophobic substituent at C8 are potent and selective GBA inhibitors and that an unambiguous Gaucher animal model can be readily generated by treatment of zebrafish with these.

Although nontypeable Haemophilus influenzae (NTHi) is a human-specific nasopharyngeal commensal bacterium, it also causes upper respiratory tract infections in children and lower respiratory tract infections in the elderly, resulting in frequent antibiotic use. The transition from symbiotic colonizing bacterium to opportunistic pathogen is not completely understood. Incorporation of sialic acids into lipooligosaccharides is thought to play an important role in bacterial virulence. It has been known for more than 25 years that sialic acids increase resistance to complement-mediated killing; however, the mechanism of action has not been elucidated thus far. Here, we provide evidence that growth of NTHi in the presence of sialic acids Neu5Ac and Neu5Gc decreases complement-mediated killing through abrogating the classical pathway of complement activation by preventing mainly IgM antibody binding to the bacterial surface. Therefore, strategies that interfere with uptake or incorporation of sialic acids into the lipooligosaccharide, such as novel antibiotics and vaccines, might be worth exploring to prevent or treat NTHi infections.

We thank The Netherlands Organization for Scientific Research (NWO-CW, ChemThem grant to J.M.F.G.A. and H.S.O.), the European Research Council (ERC-2011-AdG290836 “Chembiosphing”, to H.S.O.) and Sanofi Genzyme (research grant to J.M.F.G.A. and H.S.O. and postdoctoral contract to M.A.). We acknowledge ChemAxon for kindly providing the Instant JChem software to manage our compound library. T.M.G., V.O. and S.A.M. are supported by a Wellcome Trust Research Career Development Fellowship and Institutional Strategic Support Funding. Cyclophellitol aziridines are potent irreversible inhibitors of retaining glycosidases and versatile intermediates in the synthesis of activity‐based glycosidase probes (ABPs). Direct 3‐amino‐2‐(trifluoromethyl)quinazolin‐4(3H)‐one‐mediated aziridination of ʟ‐ido‐configured cyclohexene has enabled the synthesis of new covalent inhibitors and ABPs of α‐ʟ‐iduronidase, deficiency of which underlies the lysosomal storage disorder mucopolysaccharidosis type I (MPS I). The iduronidase ABPs react covalently and irreversibly in an activity‐based manner with human recombinant α‐ʟ‐iduronidase (rIDUA, Aldurazyme®). The structures of IDUA when complexed with the inhibitors in a non‐covalent transition state mimicking form and a covalent enzyme‐bound form provide insights into its conformational itinerary. Inhibitors 1–3 adopt a half‐chair conformation in solution (4H3 and 3H4), as predicted by DFT calculations, which is different from the conformation of the Michaelis complex observed by crystallographic studies. Consequently, 1–3 may need to overcome an energy barrier in order to switch from the 4H3 conformation to the transition state (2, 5B) binding conformation before reacting and adopting a covalent 5S1 conformation. rIDUA can be labeled with fluorescent Cy5 ABP 2 , which allows monitoring of the delivery of therapeutic recombinant enzyme to lysosomes, as is intended in enzyme replacement therapy for the treatment of MPS I patients. Publisher PDF

Currently, many policymakers try to encourage client involvement during the public service delivery process and make it a co‐production. Clients are encouraged to act as active agents and embrace an integrated approach to address their problems to empower them. However, different studies have raised questions regarding to what extent these ambitions are appropriate for clients with vulnerabilities, such as clients with multiple problems. Aiming to further explore this issue, we studied the expectations of clients with multiple problems concerning the co‐production of public services. We interviewed 46 clients with multiple problems at the start of their support trajectory. All 46 participants lived in five districts in Rotterdam, the Netherlands, and were recruited via community‐based primary care teams. Our study indicates that co‐production ambitions might not resonate with clients with multiple problems. The study shows that these clients’ expectations are driven by their feelings of being overwhelmed and stressed out by their situation, feelings of being a victim of circumstances, bad experiences with public services in the past, their evaluation of what counts as a problem and the envisioned solutions. These clients expect public service providers to take over, fix their main problem(s) and not interfere with other aspects of their lives (not an integrated approach). Although participants seek a ‘normal’ life with, e.g., a house, work, partner, children, holidays, a pet, and no stress (a white picket fence life) as ideal, they do not feel that this is attainable for them. More insight into the rationale behind these expectations could help to bridge the gap between policymakers’ ambitions and clients’ expectations.

Background New hemocytometric parameters can be used to differentiate causes of acute febrile illness (AFI). We evaluated a software algorithm–Infection Manager System (IMS)—which uses hemocytometric data generated by Sysmex hematology analyzers, for its accuracy to detect bacteremia in AFI patients with and without malaria in Burkina Faso. Secondary aims included comparing the accuracy of IMS with C-reactive protein (CRP) and procalcitonin (PCT). Methods In a prospective observational study, patients of ≥ three-month-old (range 3 months– 90 years) presenting with AFI were enrolled. IMS, blood culture and malaria diagnostics were done upon inclusion and additional diagnostics on clinical indication. CRP, PCT, viral multiplex PCR on nasopharyngeal swabs and bacterial- and malaria PCR were batch-tested retrospectively. Diagnostic classification was done retrospectively using all available data except IMS, CRP and PCT results. Findings A diagnosis was affirmed in 549/914 (60.1%) patients and included malaria (n = 191) bacteremia (n = 69), viral infections (n = 145), and malaria-bacteremia co-infections (n = 47). The overall sensitivity, specificity, and negative predictive value (NPV) of IMS for detection of bacteremia in patients of ≥ 5 years were 97.0% (95% CI: 89.8–99.6), 68.2% (95% CI: 55.6–79.1) and 95.7% (95% CI: 85.5–99.5) respectively, compared to 93.9% (95% CI: 85.2–98.3), 39.4% (95% CI: 27.6–52.2), and 86.7% (95% CI: 69.3–96.2) for CRP at ≥20mg/L. The sensitivity, specificity and NPV of PCT at 0.5 ng/ml were lower at respectively 72.7% (95% CI: 60.4–83.0), 50.0% (95% CI: 37.4–62.6) and 64.7% (95% CI: 50.1–77.6) The diagnostic accuracy of IMS was lower among malaria cases and patients, Author summary This study describes the diagnostic accuracy of the Infection Manager System (IMS), a novel diagnostic algorithm for febrile illnesses that is equipped on a routine hematology analyzer. The latest generation hematology analyzers allow better differentiation between leukocyte subsets and their phenotype. The IMS was created, using differences in immune cell subsets (their activation status for instance), to differentiate viral from bacterial etiologies of fever. Such a tool may guide clinicians in their decision the initiate or withhold antimicrobial therapy. The study was carried out among febrile patients aged 3 months and older in rural Burkina Faso, a sub-Saharan African setting where malaria is endemic. Standard microbiological techniques such as blood culture were used as a reference to assess the diagnostic accuracy of IMS. We then compared the diagnostic accuracy of the IMS with the marketed biomarkers C-reactive protein (CRP) and procalcitonin (PCT). Our study showed that the diagnostic performance of the IMS was similar to CRP and better than PCT to detect bacteremia in patients with and without malaria co-infection. Further studies are needed to see if the IMS can be safely used to guide initial antimicrobial treatment and help to reduce further spread of antimicrobial resistance.

Staphylococcus aureus is the leading cause of skin and soft tissue infections. It remains incompletely understood how skin-resident immune cells respond to invading S. aureus and contribute to an effective immune response. Langerhans cells (LCs), the only professional antigen-presenting cell type in the epidermis, sense S. aureus through their pattern-recognition receptor langerin, triggering a proinflammatory response. Langerin recognizes the β-1,4-linked N-acetylglucosamine (β1,4-GlcNAc) but not α-1,4-linked GlcNAc (α1,4-GlcNAc) modifications, which are added by dedicated glycosyltransferases TarS and TarM, respectively, on the cell wall glycopolymer wall teichoic acid (WTA). Recently, an alternative WTA glycosyltransferase, TarP, was identified, which also modifies WTA with β-GlcNAc but at the C-3 position (β1,3-GlcNAc) of the WTA ribitol phosphate (RboP) subunit. Here, we aimed to unravel the impact of β-GlcNAc linkage position for langerin binding and LC activation. Using genetically modified S. aureus strains, we observed that langerin similarly recognized bacteria that produce either TarS- or TarP-modified WTA, yet tarP-expressing S. aureus induced increased cytokine production and maturation of in vitro-generated LCs compared to tarS-expressing S. aureus. Chemically synthesized WTA molecules, representative of the different S. aureus WTA glycosylation patterns, were used to identify langerin-WTA binding requirements. We established that β-GlcNAc is sufficient to confer langerin binding, thereby presenting synthetic WTA molecules as a novel glycobiology tool for structure-binding studies and for elucidating S. aureus molecular pathogenesis. Overall, our data suggest that LCs are able to sense all β-GlcNAc-WTA producing S. aureus strains, likely performing an important role as first responders upon S. aureus skin invasion.

Amylases are key enzymes in the processing of starch in many kingdoms of life. They are important catalysts in industrial biotechnology where they are applied in, among others, food processing and the production of detergents. In man amylases are the first enzymes in the digestion of starch to glucose and arguably also the preferred target in therapeutic strategies aimed at the treatment of type 2 diabetes patients through down-tuning glucose assimilation. Efficient and sensitive assays that report selectively on retaining amylase activities irrespective of the nature and complexity of the biomaterial studied are of great value both in finding new and effective human amylase inhibitors and in the discovery of new microbial amylases with potentially advantageous features for biotechnological application. Activity-based protein profiling (ABPP) of retaining glycosidases is inherently suited for the development of such an assay format. We here report on the design and synthesis of 1,6-epi-cyclophellitol-based pseudodisaccharides equipped with a suite of reporter entities and their use in ABPP of retaining amylases from human saliva, murine tissue as well as secretomes from fungi grown on starch. The activity and efficiency of the inhibitors and probes are substantiated by extensive biochemical analysis, and the selectivity for amylases over related retaining endoglycosidases is validated by structural studies.

OBJECTIVES: Areas with declining malaria transmission in sub-Saharan Africa have recently witnessed important changes in the aetiology of childhood acute febrile illness (AFI). We describe the aetiology of AFI in a high malaria transmission area in rural Burkina Faso. METHODS: In a prospective hospital-based diagnostic study, children aged 3 months to 15 years with AFI were recruited and assessed using a systematic diagnostic protocol, including blood cultures, whole blood PCR on a selection of bacterial pathogens, malaria diagnostics and a multiplex PCR on nasopharyngeal swabs targeting 21 viral and 4 bacterial respiratory pathogens. RESULTS: A total of 589 children with AFI were enrolled from whom an infectious disease was considered in 575 cases. Acute respiratory tract infections, malaria and invasive bacterial infections (IBI) accounted for 179 (31.1%), 175 (30.4%) and 75 (13%) of AFI cases respectively; 16 (21.3%) of IBI cases also had malarial parasitaemia. A viral pathogen was demonstrated from the nasopharynx in 157 children (90.7%) with respiratory tract symptoms. Of all children with viral respiratory tract infections, 154 (92.4% received antibiotics, whereas no antibiotic was provided in 13 (17%) of IBI cases. CONCLUSIONS: Viral respiratory infections are a common cause of childhood AFI in high malaria transmission areas, next to malaria and IBI. These findings highlight the importance of interventions to improve targeted treatment with antimicrobials. Most patients with viral infections received antibiotics unnecessarily, while a considerable number with IBI did not receive antibiotics. ispartof: CLINICAL MICROBIOLOGY AND INFECTION vol:27 issue:4 pages:590-596 ispartof: location:England status: published

20 pags., 7 figs., 2 tabs., Genetic variants arising from within-patient evolution shed light on bacterial adaptation during chronic infection. Contingency loci generate high levels of genetic variation in bacterial genomes, enabling adaptation to the stringent selective pressures exerted by the host. A significant gap in our understanding of phase-variable contingency loci is the extent of their contribution to natural infections. The human-adapted pathogen nontypeable Haemophilus influenzae (NTHi) causes persistent infections, which contribute to underlying disease progression. The phase-variable high-molecular-weight (HMW) adhesins located on the NTHi surface mediate adherence to respiratory epithelial cells and, depending on the allelic variant, can also confer high epithelial invasiveness or hyperinvasion. In this study, we characterize the dynamics of HMW-mediated hyperinvasion in living cells and identify a specific HMW binding domain shared by hyperinvasive NTHi isolates of distinct pathological origins. Moreover, we observed that HMW expression decreased over time by using a longitudinal set of persistent NTHi strains collected from chronic obstructive pulmonary disease (COPD) patients, resulting from increased numbers of simple-sequence repeats (SSRs) downstream of the functional P2hmw1A promoter, which is the one primarily driving HMW expression. Notably, the increased SSR numbers at the hmw1 promoter region also control a phenotypic switch toward lower bacterial intracellular invasion and higher biofilm formation, likely conferring adaptive advantages during chronic airway infection by NTHi. Overall, we reveal novel molecular mechanisms of NTHi pathoadaptation based on within-patient lifestyle switching controlled by phase variation. IMPORTANCE Human-adapted bacterial pathogens have evolved specific mechanisms to colonize their host niche. Phase variation is a contingency strategy to allow adaptation to changing conditions, as phase-variable bacterial loci rapidly and reversibly switch their expression. Several NTHi adhesins are phase variable. These adhesins are required for colonization but also immunogenic, in such a way that bacteria with lower adhesin levels are better equipped to survive an immune response, making their contribution to natural infections unclear. We show here that the major NTHi adhesin HMW1A displays allelic variation, which can drive a phase-variable epithelial hyperinvasion phenotype. Over time, hmw1A phase variation lowers adhesin expression, which controls an NTHi lifestyle switch from high epithelial invasiveness to lower invasion and higher biofilm formation. This reversible loss of function aligns with the previously stated notion that epithelial infection is essential for NTHi infection establishment, but once established, persistence favors gene inactivation, in this case facilitating biofilm growth., This work has been funded by grants SAF2015-66520-R and RTI2018-096369-B-I00 from MINECO, 875/2019 from SEPAR, and PC150-151-152 from the Gobierno de Navarra to J.G. and RTI2018-099985-B-I00 to M.M. CIBER is an initiative from the Instituto de Salud Carlos III (ISCIII), Madrid, Spain. A.T.-A. is funded by a Ministry of Science and Innovation grant (PID2019-105216GB-I00)

Synthetic vaccines, based on antigenic peptides that comprise MHC−I and MHC‐II T‐cell epitopes expressed by tumors, show great promise for the immunotherapy of cancer. For optimal immunogenicity, the synthetic peptides (SPs) should be adjuvanted with suitable immunostimulatory additives. Previously, we have shown that improved immunogenicity in vivo is obtained with vaccine modalities in which an SP is covalently connected to an adjuvanting moiety, typically a ligand to Toll‐like receptor 2 (TLR2). SPs were covalently attached to UPam, which is a derivative of the classic TLR2 ligand Pam3CysSK4. A disadvantage of the triply palmitoylated UPam is its high lipophilicity, which precludes universal adoption of this adjuvant for covalent modification of various antigenic peptides as it renders the synthetic vaccine insoluble in several cases. Here, we report a novel conjugatable TLR2 ligand, mini‐UPam, which contains only one palmitoyl chain, rather than three, and therefore has less impact on the solubility and other physicochemical properties of a synthetic peptide. In this study, we used SPs that contain the clinically relevant neoepitopes identified in a melanoma patient who completely recovered after T‐cell therapy. Homogeneous mini‐UPam‐SP conjugates have been prepared in good yields by stepwise solid‐phase synthesis that employed a mini‐UPam building block pre‐prepared in solution and the standard set of Fmoc‐amino acids. The immunogenicity of the novel mini‐UPam‐SP conjugates was demonstrated by using the cancer patient's T‐cells., Less lipophilic ligands: A novel TLR2 ligand has been designed and synthesized to be subsequently conjugated with synthetic peptides containing clinically relevant neoepitopes. By treating HEK‐TLR2 cells, human moDCs and antigen specific human T‐cells with these new constructs, the immunogenic potential of the new TLR2 ligand and the conjugation of TLR ligands with neoepitopes was assessed.

Point‐of‐care (POC) diagnostic tests for the rapid detection of individuals infected with Mycobacterium leprae, the causative pathogen of leprosy, represent efficient tools to guide therapeutic and prophylactic treatment strategies in leprosy control programs, thus positively contributing to clinical outcome and reducing transmission of this infectious disease. Levels of antibodies directed against the M. leprae‐specific phenolic glycolipid I (PGL−I) closely correlate with an individual's bacterial load and a higher risk of developing leprosy. We describe herein the assembly of a set of PGL glycans carrying the characteristic phenol aglycon and featuring different methylation patterns. The PGL trisaccharides were applied to construct neoglycoproteins that were used to detect anti‐PGL IgM antibodies in leprosy patients. ELISAs and quantitative lateral‐flow assays based on up‐converting nanoparticles (UCP‐LFAs) showed that the generated PGL−I and PGL‐II trisaccharide neoglycoconjugates can be applied for the detection of anti M. leprae IgM antibodies in POC tests., Getting to the point of care: Trisaccharides of phenolic glycolipids (PGL) originating from M. leprae were evaluated for their diagnostic potential as neoglycoproteins in ELISA and an up‐converting particle lateral flow assay (UCP‐LFA). Two naturally occurring trisaccharides were found to be suitable for this purpose and can thus be used for future field studies in leprosy endemic areas.

Anthracycline anticancer drugs doxorubicin and aclarubicin have been used in the clinic for several decades to treat various cancers. Although closely related structures, their molecular mode of action diverges, which is reflected in their biological activity profile. For a better understanding of the structure-function relationship of these drugs, we synthesized ten doxorubicin/aclarubicin hybrids varying in three distinct features: aglycon, glycan, and amine substitution pattern. We continued to evaluate their capacity to induce DNA breaks, histone eviction, and relocated topoisomerase II alpha in living cells. Furthermore, we assessed their cytotoxicity in various human tumor cell lines. Our findings underscore that histone eviction alone, rather than DNA breaks, contributes strongly to the overall cytotoxicity of anthracyclines, and structures containing N,N-dimethylamine at the reducing sugar prove that are more cytotoxic than their nonmethylated counterparts. This structural information will support further development of novel anthracycline variants with improved anticancer activity.

Mannose‐6‐phosphate (M6P) is recognized by the mannose‐6‐phosphate receptor and plays an important role in the transport of cargo to the endosomes, making it an attractive tool to improve endosomal trafficking of vaccines. We describe herein the assembly of peptide antigen conjugates carrying clusters of mannose‐6‐C‐phosphonates (M6Po). The M6Po's are stable M6P mimics that are resistant to cleavage of the phosphate group by endogenous phosphatases. Two different strategies for the incorporation of the M6Po clusters in the conjugate have been developed: the first relies on a “post‐assembly” click approach employing an M6Po bearing an alkyne functionality; the second hinges on an M6Po C‐glycoside amino acid building block that can be used in solid‐phase peptide synthesis. The generated conjugates were further equipped with a TLR7 ligand to stimulate dendritic cell (DC) maturation. While antigen presentation is hindered by the presence of the M6Po clusters, the incorporation of the M6Po clusters leads to increased activation of DCs, thus demonstrating their potential in improving vaccine adjuvanticity by intraendosomally active TLR ligands., Enhancing the immune response: Two mannose‐6‐C‐phosphonate (M6Po) building blocks have been synthesized that can be incorporated into a conjugate by a “post‐assembly” click or an online SPPS approach to improve endosomal trafficking of vaccines. The generated M6Po conjugates, which were also equipped with a TLR7 ligand, were able to enhance the immune response by activation and maturation of dendritic cells.

INTRODUCTION: China has made considerable progress with health system reforms in recent years. Rural China, however, has lagged behind as the diversity of needs of China's 3,000 rural counties were not always well addressed by national top-down reforms. China's Rural Health Reform Project Health XI (HXI) piloted a hybrid process of top down and bottom up implementation of health system reforms which were tailored to rural county level needs and covered a population of more than 21 million. Different studies provide evidence that HXI counties have achieved substantial benefits given the relatively limited investment. The Effectiveness of HXI subsequently raises the question how the hybrid approach may have resulted in effective implementation of interventions. We answer this question to advance understanding of hybrid approaches in general and in the rural Chinese context in particular, where the bottom-up elements might match poorly with the traditional organisational culture and learning style.MATERIALS & METHODS: We conducted an in-depth qualitative analysis in three 'best practice' counties, performing document-analyses, observations, semi-structured individual and group interviews. In alignment with the research question, this study is of an explorative nature and follows a sequence of deductive and inductive steps.RESULTS: HXI struggled initially as counties had difficulties to take initiative and autonomously select and adapt their own reforms. The initial reforms required multiple improvement iterations before achieving the planned results. The effectiveness of these bottom up reform processes has been aided by tight top down supervision and extensive domestic expert involvement. County level leadership is seen as essential to align the top down and bottom up structures and processes. Where successful, HXI has changed mind-sets and counties developed generic health improvement capabilities.CONCLUSION: Tailoring innovations to fit local needs formed a severe challenge for the three 'best practice' counties studied. A 'change of mindset' to actively take initiative and assume autonomy was needed to advance. Top down supervision and extensive support of experts was required to overcome the barriers. The studied counties finally achieved sustainable improvements and developed double loop learning capabilities beyond HXI objectives. Taken together, the above findings suggest that the continuum of healthcare reform implementation approaches in which hybrid approaches reside-from bottom up to top down-has two dimensions: a content dimension and a procedural dimension. Enabled by top down procedures, counties were able to bottom up tailor the content of best practice innovations to fit local needs.

Golgi mannosidase II (GMII) catalyzes the sequential hydrolysis of two mannosyl residues from GlcNAcMan

Background Neutrophil accumulation in the skin is a hallmark of psoriasis. Novel insights on neutrophil phenotypic and functional heterogeneity raise the question to what extent these cells contribute to the sustained inflammatory skin reaction. Objective We sought to examine the phenotype and functional properties of neutrophils in blood and skin of patients with psoriasis, and the effect of TNF-α and p40(IL-12/IL-23) antibody therapy on circulating neutrophils. Methods Thirty-two patients with psoriasis were enrolled in an observational study performed in 2 university hospitals. We evaluated neutrophil phenotype and function using in vitro (co)culture stimulation assays, flow cytometry, multiplex immunohistochemistry, and multispectral imaging of patient-derived blood and skin samples. Results Cluster of differentiation (CD)10pos and CD10neg neutrophils were increased in peripheral blood of patients with psoriasis. In CD10neg neutrophils, different maturation stages were observed, including a subset resembling aged neutrophils that was 3 times more abundant than in healthy individuals. These aged neutrophils displayed suboptimal canonical neutrophil functions and induced IL-17 and IFN-γ production by T cells in vitro, mediated by neutrophil extracellular trap formation. Also, mature and aged neutrophils were present in psoriatic skin and were found in the vicinity of T cells. Upon antibody therapy, numbers of these cells in circulation decreased. Conclusions Patients with psoriasis reveal a unique neutrophil profile in circulation, and 2 distinct neutrophil subsets are present in psoriatic skin. Targeted biological treatment may aid in the containment of sustained neutrophil-mediated inflammation.

Objective To examine the association between bacterial-viral co-occurrence in the nasopharynx and the risk of community acquired pneumonia (CAP) in young children living in resource-limited settings. Methods A case-control study was conducted between January and December 2017 in Moshi, Tanzania. Children 2–59 months with CAP and healthy controls were enrolled. RSV and Influenza A/B were detected with a standardized polymerase chain reaction (PCR) method, and a simplified real-time quantitative PCR method, without sample pre-processing, was developed to detect bacterial pathogens in nasopharyngeal samples. Results A total of 109 CAP patients and 324 healthy controls were enrolled. Co-detection of H. influenzae and S. pneumoniae in nasopharyngeal swabs was linked with higher odds of CAP (aOR=3.2, 95% CI=1.1–9.5). The majority of the H. influenzae isolated in cases and controls (95.8%) were non-typeable. Of the viruses examined, respiratory syncytial virus (RSV) was most common (n = 31, 7.2%) in cases and controls. Children with RSV had 8.4 times higher odds to develop pneumonia than healthy children (aOR=8.4, 95%CI= 3.2 – 22.1). Conclusions Co-occurence of H. influenzae and S. pneumoniae in the nasopharynx was strongly associated with CAP. The high prevalence of non-typeable H. influenzae might be a sign of replacement as a consequence of Haemophilus influenzae type b vaccination.

BackgroundA high variety of team interventions aims to improve team performance outcomes. In 2008, we conducted a systematic review to provide an overview of the scientific studies focused on these interventions. However, over the past decade, the literature on team interventions has rapidly evolved. An updated overview is therefore required, and it will focus on all possible team interventions without restrictions to a type of intervention, setting, or research design.ObjectivesTo review the literature from the past decade on interventions with the goal of improving team effectiveness within healthcare organizations and identify the “evidence base” levels of the research.MethodsSeven major databases were systematically searched for relevant articles published between 2008 and July 2018. Of the original search yield of 6025 studies, 297 studies met the inclusion criteria according to three independent authors and were subsequently included for analysis. The Grading of Recommendations, Assessment, Development, and Evaluation Scale was used to assess the level of empirical evidence.ResultsThree types of interventions were distinguished: (1)Training, which is sub-divided into training that is based on predefined principles (i.e. CRM: crew resource management and TeamSTEPPS: Team Strategies and Tools to Enhance Performance and Patient Safety), on a specific method (i.e. simulation), or on general team training. (2)Toolscovers tools that structure (i.e. SBAR: Situation, Background, Assessment, and Recommendation, (de)briefing checklists, and rounds), facilitate (through communication technology), or trigger (through monitoring and feedback) teamwork. (3)Organizational (re)designis about (re)designing structures to stimulate team processes and team functioning.(4) Aprogrammeis a combination of the previous types. The majority of studies evaluated a training focused on the (acute) hospital care setting. Most of the evaluated interventions focused on improving non-technical skills and provided evidence of improvements.ConclusionOver the last decade, the number of studies on team interventions has increased exponentially. At the same time, research tends to focus on certain interventions, settings, and/or outcomes. Principle-based training (i.e. CRM and TeamSTEPPS) and simulation-based training seem to provide the greatest opportunities for reaching the improvement goals in team functioning.

Background: Most interventions to improve patient safety (Patient Safety Practices (PSPs)), are introduced without engaging front-line professionals. Administrative staff, managers and sometimes a few professionals, representing only one or two disciplines, decide what to change and how. Consequently, PSPs are not fully adapted to the professionals' needs or to the local context and as a result, adoption is low. To support adoption, two theoretical concepts, Participatory Design and Experiential Learning were combined in a new model: Adaptive Design. The aim was to explore whether Adaptive Design supports adaptation and adoption of PSPs by engaging all professionals and creating time to (re) design, reflect and learn as a team. The Time Out Procedure (TOP) and Debriefing (plus) for improving patient safety in the operating theatre (OT) was used as PSP. Methods: Qualitative exploratory multi-site study using participatory action research as a research design. The implementation process consisted of four phases: 1) start-up: providing information by presentations and team meetings, 2) pilot: testing the prototype with 100 surgical procedures, 3) small scale implementation: with one or two surgical disciplines, 4) implementation hospital-wide: including all surgical disciplines. In iterations, teams (re) designed, tested, evaluated, and if necessary adapted TOPplus. Gradually all professionals were included. Adaptations in content, process and layout of TOPplus were measured following each iteration. Adoption was monitored until final implementation in every hospital's OT. Results: 10 Dutch hospitals participated. Adaptations varied per hospital, but all hospitals adapted both procedures. Adaptations concerned the content, process and layout of TOPplus. Both procedures were adopted in all OTs, but user participation and time to include all users varied between hospitals. Ultimately all users were actively involved and TOPplus was implemented in all OTs. Conclusions: Engaging all professionals in a structured bottom-up implementation approach with a focus on learning, improves adaptation and adoption of a PSP. As a result, all 10 participating hospitals implemented TOPplus with all surgical disciplines in all OTs. Adaptive Design gives professionals the opportunity to adapt the PSP to their own needs and their specific local context. All hospitals adapted TOPplus, but without compromising the essential features for its effectiveness.

Haemophilus influenzae is a Gram-negative bacterium that can be classified into typeable (types a through f) and nontypeable (NTHi) groups. This opportunistic pathogen asymptomatically colonizes the mucosal epithelium of the upper respiratory tract, from where it spreads to other neighboring regions, potentially leading to disease. Infection with NTHi can cause otitis media, sinusitis, conjunctivitis, exacerbations of chronic obstructive pulmonary disease, and pneumonia, but it is increasingly causing invasive disease, including bacteremia and meningitis. Invasive NTHi strains are more resistant to complement-mediated killing. However, the mechanisms of complement resistance have never been studied in large numbers of invasive NTHi strains. In this study, we determined the relationship between binding of IgG or IgM and the bacterial survival in normal human serum for 267 invasive H. influenzae strains from Spain, Portugal, and the Netherlands, of which the majority (200 [75%]) were NTHi. NTHi bacteria opsonized with high levels of IgM had the lowest survival in human serum. IgM binding to the bacterial surface, but not IgG binding, was shown to be associated with complement-mediated killing of NTHi strains. We conclude that evasion of IgM binding by NTHi strains increases survival in blood, thereby potentially contributing to their ability to cause severe invasive diseases. info:eu-repo/semantics/publishedVersion

Contains fulltext : 218852.pdf (Publisher’s version ) (Open Access) Streptococcus pneumoniae and Neisseria meningitidis are pathogens that frequently colonize the nasopharynx in an asymptomatic manner but are also a cause of invasive bacterial infections mainly in young children. The complement system plays a crucial role in humoral immunity, complementing the ability of antibodies to clear microbes, thereby protecting the host against bacterial infections, including S. pneumoniae and N. meningitidis. While it is widely accepted that complement deficiencies due to rare genetic variants increase the risk for invasive bacterial infection, not much is known about the common genetic variants in the complement system in relation to disease susceptibility. In this review, we provide an overview of the effects of common genetic variants on complement activation and on complement-mediated inflammation.

The aim of this study was to determine whether the extent of peritoneal metastases (PMs) on preoperative diffusion-weighted magnetic resonance imaging (DW-MRI) can be used as a biomarker of disease-free and overall survival in patients with colorectal cancer who are considered for cytoreductive surgery and hyperthermic intraperitoneal chemotherapy (CRS/HIPEC).For this retrospective cohort study, patients with PMs considered for CRS/HIPEC who underwent DW-MRI for preoperative staging in 2016-2017 were included. The DW-MRI protocol consisted of diffusion-weighted, T2-weighted, and pre- and post-gadolinium T1-weighted imaging of the chest, abdomen, and pelvis. DW-MRI images were evaluated by two independent readers to determine the extent of PMs represented by the Peritoneal Cancer Index (MRI-PCI), as well as extraperitoneal metastases. Cox regression and Kaplan-Meier analysis was performed to determine the prognostic value of DW-MRI for overall and disease-free survival.Seventy-eight patients were included. CRS/HIPEC was planned for 53 patients and completed in 50 patients (60.5%). Median follow-up after DW-MRI was 23 months (interquartile range 13-24). The MRI-PCI of both readers showed prognostic value for overall survival, independently of whether R1 resection was achieved (hazard ratio [HR] 1.06-1.08; p 0.05). For the patients who received successful CRS/HIPEC, the MRI-PCI also showed independent prognostic value for disease-free survival for both readers (HR 1.09-1.10; p 0.05).The extent of PMs on preoperative DW-MRI is an independent predictor of overall and disease-free survival and should therefore be considered as a non-invasive prognostic biomarker.

Hypoxia and inflammation are closely intertwined phenomena. Critically ill patients often suffer from systemic inflammatory conditions and concurrently experience short-lived hypoxia. We evaluated the effects of short-term hypoxia on systemic inflammation, and show that it potently attenuates pro-inflammatory cytokine responses during murine endotoxemia. These effects are independent of hypoxia-inducible factors (HIFs), but involve augmented adenosine levels, in turn resulting in an adenosine 2B receptor-mediated post-transcriptional increase of interleukin (IL)-10 production. We translated our findings to humans using the experimental endotoxemia model, where short-term hypoxia resulted in enhanced plasma concentrations of adenosine, augmentation of endotoxin-induced circulating IL-10 levels, and concurrent attenuation of the pro-inflammatory cytokine response. Again, HIFs were shown not to be involved. Taken together, we demonstrate that short-term hypoxia dampens the systemic pro-inflammatory cytokine response through enhanced purinergic signaling in mice and men. These effects may contribute to outcome and provide leads for immunomodulatory treatment strategies for critically ill patients., Highlights • Short-term hypoxia attenuates the systemic pro-inflammatory cytokine response in vivo in both mice and men • The underlying mechanism involves adenosine 2B receptor-mediated enhanced production of anti-inflammatory interleukin-10 • These effects may contribute to outcome and provide leads for novel treatment strategies for critically ill patients Inflammation and short bouts of low oxygen levels are frequently encountered phenomena in severely ill patients admitted to the Intensive Care unit. This study investigated the effect of short-term low oxygen levels on the inflammatory response in both mice and humans. The results show that a short period of low levels of oxygen potently dampens inflammation, and identify the underlying mechanisms. These effects of low oxygen levels may contribute to the outcome of severely ill patients. Furthermore, the increased understanding of the underlying mechanisms provided by this study can be used to develop therapies to dampen inflammation in patients.

Antibody replacement therapy for patients with antibody deficiencies contains only IgG. As a result, concurrent IgM and IgA deficiency present in a large proportion of antibody deficient patients persists. Especially patients with IgM deficiency remain at risk for recurrent infections of the gastrointestinal and respiratory tract. The lack of IgM in the current IgG replacement therapy is likely to contribute to the persistence of these mucosal infections because this antibody class is especially important for complement activation on the mucosal surface. We evaluated whether supplementation with IgM increased serum bactericidal capacity in vitro. Serum was collected from a patient with agammaglobulinemia and supplemented with purified serum IgM to normal levels. Antibody and complement deposition on the bacterial surface was determined by multi-color flow cytometry. Bacterial survival in serum was determined by colony-forming unit counts. We present a patient previously diagnosed with agammaglobulinemia due to CD79A (Igα) deficiency revealing a novel pathogenic insertion variant in the CD79a gene (NM_001783.3:c.353_354insT). Despite IgG replacement therapy and antibiotic prophylaxis, this patient developed a Campylobacter jejuni spondylodiscitis of lumbar vertebrae L4–L5. We found that serum IgM significantly contributes to complement activation on the bacterial surface of C. jejuni. Furthermore, supplementation of serum IgM augmented serum bactericidal activity significantly. In conclusion, supplementation of intravenous IgG replacement therapy with IgM may potentially offer greater protection against bacterial infections, also in the context of increasing antibiotic resistance.

Contains fulltext : 177354.pdf (Publisher’s version ) (Open Access) Vaccination has been one of the most successful strategies to reduce morbidity and mortality caused by respiratory infections. Recent evidence suggests that differences in the host genetic background and environmental factors may contribute to heterogeneity in the immune response to vaccination. During pre-clinical testing, vaccines are often evaluated in a single mouse inbred strain, which may not translate well to the heterogeneous human population. Here, we examined the influence of host genetic background on vaccine-induced protection against pneumococcal colonization in two commonly used inbred mouse strains, i.e. C57BL/6 and BALB/cas well as the F1 cross of these two strains. Groups of mice were vaccinated intranasally with a vaccine formulation containing a model pneumococcal antigen, i.e. pneumococcal surface protein A (PspA), adjuvanted with cholera toxin subunit B (CTB). Even in the absence of vaccination, differences in colonization density were observed between mouse strains. Although vaccination significantly reduced pneumococcal density in all mouse strains, differences were observed in the magnitude of protection. We therefore examined immunological parameters known to be involved in vaccine-induced mucosal clearance of S. pneumoniae. We found that PspA-specific IgG levels in nasal tissue differed between mouse strains, but in all cases it correlated significantly with a reduction in colonization. Furthermore, increased mucosal IL17A, but not IFNgamma, IL10, or IL4, was found to be mouse strain specific. This suggests that the reduction of bacterial load may be accompanied by a Th17 response in all genetic backgrounds, although the cytokine dynamics may differ. Increased insight into the different immune mechanisms that affect pneumococcal carriage will contribute to development of future vaccines against S. pneumoniae.

Contains fulltext : 177218.pdf (Publisher’s version ) (Open Access) Metabolic incorporation of azide- or alkyne-modified sialic acids into the cellular glycosylation pathway enables the study of sialoglycan expression, localization, and trafficking via bioorthogonal chemistry. Herein, we report that such modifications of the sialic acid sugar can have a profound influence on their hydrolysis by neuraminidases (sialidase). Azidoacetyl (Az)-modified sialic acids were prone to neuraminidase cleavage, whereas propargyloxycarbonyl (Poc)-modified sialic acids were largely resistant to cleavage. Because the influenza virus infection cycle depends on the hydrolysis of host-cell-surface sialic acids, influenza cell-to-cell transmission was strongly reduced in Poc sialic acid glycoengineered host cells. The use of Poc sialic acids may disturb biological processes involving neuraminidase cleavage but also provides perspective for use in applications in which sialic acid hydrolysis is not desired, such as antibody modification, viral infection, etc. 5 p.

Contains fulltext : 171105.pdf (Publisher’s version ) (Open Access) Bacterial pathogens not only stimulate innate immune receptors, but also activate the complement system. Crosstalk between complement C5a receptor (C5aR) and other innate immune receptors is known to enhance the proinflammatory cytokine response. An important determinant of the magnitude of complement activation is the activity of the alternative pathway, which serves as an amplification mechanism for complement activation. Both alternative pathway activity as well as plasma levels of factor H, a key inhibitor of the alternative pathway, show large variation within the human population. Here, we studied the effect of factor H-mediated regulation of the alternative pathway on bacterial-induced proinflammatory cytokine responses. We used the human pathogen Streptococcus pneumoniae as a model stimulus to induce proinflammatory cytokine responses in human peripheral blood mononuclear cells. Serum containing active complement enhanced pneumococcal induced proinflammatory cytokine production through C5a release and C5aR crosstalk. We found that inhibition of the alternative pathway by factor H, with a concentration equivalent to a high physiological level, strongly reduced C5a levels and decreased proinflammatory cytokine production in human peripheral blood mononuclear cells. This suggests that variation in alternative pathway activity due to variation in factor H plasma levels affects individual cytokine responses during infection.

Nontypeable Haemophilus influenzae (NTHi) bacteria express various molecules that contribute to their virulence. The presence of phosphocholine (PCho) on NTHi lipooligosaccharide increases adhesion to epithelial cells and is an advantage for the bacterium, enabling nasopharyngeal colonization, as measured in humans and animal models. However, when PCho is expressed on the lipooligosaccharide, it is also recognized by the acute-phase protein C-reactive protein (CRP) and PCho-specific antibodies, both of which are potent initiators of the classical pathway of complement activation. In this study, we show that blood isolates, which are exposed to CRP and PCho-specific antibodies in the bloodstream, have a higher survival in serum than oropharyngeal isolates, which was associated with a decreased presence of PCho. PCho(low) strains showed decreased IgM, CRP, and complement C3 deposition, which was associated with increased survival in human serum. Consistent with the case for the PCho(low) strains, removal of PCho expression by licA gene deletion decreased IgM, CRP, and complement C3 deposition, which increased survival in human serum. Complement-mediated killing of PCho(high) strains was mainly dependent on binding of IgM to the bacterial surface. These data support the hypothesis that a PCho(low) phenotype was selected in blood during invasive disease, which increased resistance to serum killing, mainly due to lowered IgM and CRP binding to the bacterial surface.

Summary Vaccination against meningococcal serogroup B is recommended for patients with a complement deficiency; however, although immunogenicity in this patient group has been shown, efficacy has not yet been established. In this study, we collected serum from children with a complement deficiency in the alternative pathway or in late terminal pathway before and after vaccination with multi‐component meningococcal serogroup B (MenB)‐4C. MenB‐4C is a multi‐component, protein‐based vaccine against MenB consisting of factor H‐binding protein, Neisserial heparin‐binding protein, Neisserial adhesion A and outer membrane vesicles containing Porin A. We assessed the vaccine immunogenicity and vaccine‐mediated protection by a whole cell enzyme‐linked immunosorbent assay with Neisseria meningitidis serogroup B strains H44/76, 5/99 and NZ98/254, which shows that vaccination induced antibody titers against meningococcus. We show that the classical serum bactericidal activity assay with exogenous serum indicates the presence of vaccine‐induced antibodies and capacity to activate complement‐mediated pathogen lysis. However, in children with a late terminal pathway deficiency, no complement‐mediated pathogen lysis was observed when autologous serum was applied in the serum bactericidal activity assay, demonstrating a lack of serum bactericidal activity in children with complement deficiencies. However, MenB‐4C vaccination still induced effective complement‐dependent opsonophagocytic killing against N. meningitidis serogroup B in reconstituted whole blood with autologous serum from children with an alternative pathway or late terminal pathway deficiency. These findings support the recommendation to vaccinate all complement‐deficient children against MenB., Patients with a complement deficiency are extensively vaccinated against Meningococcal infection, including with the 4C‐MenB vaccine against N. meningitidis serogroup B. We show that vaccination induces functional antibodies, but does not result in bacterial killing via the complement system in patients with a late terminal pathway deficiency. Despite the complement deficiency, killing can occur via opsonophagocytosis.

Nontypeable Haemophilus influenzae (NTHi) colonizes the human upper respiratory tract without causing disease symptoms, but it is also a major cause of upper and lower respiratory tract infections in children and elderly, respectively. NTHi synthesizes various molecules to decorate its lipooligosaccharide (LOS), which modulates the level of virulence. The presence of phosphorylcholine (PCho) on NTHi LOS increases adhesion to epithelial cells, which is an advantage for the bacterium enabling nasopharyngeal colonization. However, when PCho is incorporated on the LOS of NTHi, it is recognized by the acute-phase C-reactive protein (CRP) and PCho-specific antibodies, both potent initiators of the classical pathway of complement activation. We determined the presence of PCho and binding of IgG and IgM to the bacterial surface for 319 NTHi strains collected from the nasopharynx/oropharynx, middle ear, and lower respiratory tract. PCho detection was higher for NTHi strains collected from the nasopharynx/oropharynx, which was associated with increased binding of IgM and IgG to the bacterial surface. Binding of CRP and IgM to the bacterial surface of PCho(high) NTHi strains increased complement-mediated killing, which was largely dependent on PCho-specific IgM. The levels of PCho-specific IgM varied in sera from 12 healthy individuals, and higher PCho-specific IgM levels were associated with increased complement-mediated killing of a PCho(high) NTHi strain. In conclusion, incorporation of PCho on the LOS of NTHi marks the bacterium for binding of CRP and IgM, resulting in complement-mediated killing. Therefore, having a lower PCho might be beneficial in situations where sufficient PCho-specific antibodies and complement are present.

Covalent linking of immunogenic oligopeptides with synthetic Toll-like receptor ligands is a useful approach to develop self-adjuvanting vaccines. In particular, small-molecule based agonists of Toll-like receptor 7 (TLR7) that are derived from 8-oxo-adenine core are potentially promising because these chemically robust TLR7 ligands can be connected to peptide T-cell epitopes via straightforward solid-phase peptide synthesis. In this contribution we present the synthesis of a Boc-protected 9-benzyl-2-alkoxy-8-oxo-adenine building block and its application in the online solid phase synthesis of three peptide conjugates that differ in the position of the TLR7 ligand within the peptide. The conjugates are able to induce dendritic cell maturation and T cell proliferation while the position of the ligand impacts T cell proliferation potency.