Adoption of C-reactive protein rapid tests for the management of acute childhood infections in hospitals in the Netherlands and England: a comparative health systems analysis.

BACKGROUND: The adoption of C-reactive protein point-of-care tests (CRP POCTs) in hospitals varies across Europe. We aimed to understand the factors that contribute to different levels of adoption of CRP POCTs for the management of acute childhood infections in two countries. METHODS: Comparative qualitative analysis of the implementation of CRP POCTs in the Netherlands and England. The study was informed by the non-adoption, abandonment, spread, scale-up, and sustainability (NASSS) framework. Data were collected through document analysis and qualitative interviews with stakeholders. Documents were identified by a scoping literature review, search of websites, and through the stakeholders. Stakeholders were sampled purposively initially, and then by snowballing. Data were analysed thematically. RESULTS: Forty-one documents resulted from the search and 46 interviews were conducted. Most hospital healthcare workers in the Netherlands were familiar with CRP POCTs as the tests were widely used and trusted in primary care. Moreover, although diagnostics were funded through similar Diagnosis Related Group reimbursement mechanisms in both countries, the actual funding for each hospital was more constrained in England. Compared to primary care, laboratory-based CRP tests were usually available in hospitals and their use was encouraged in both countries because they were cheaper. However, CRP POCTs were perceived as useful in some hospitals of the two countries in which the laboratory could not provide CRP measures 24/7 or within a short timeframe, and/or in emergency departments where expediting patient care was important. CONCLUSIONS: CRP POCTs are more available in hospitals in the Netherlands because of the greater familiarity of Dutch healthcare workers with the tests which are widely used in primary care in their country and because there are more funding constraints in England. However, most hospitals in the Netherlands and England have not adopted CRP POCTs because the alternative CRP measurements from the hospital laboratory are available in a few hours and at a lower cost.

Primary Exposure to SARS-CoV-2 via Infection or Vaccination Determines Mucosal Antibody-Dependent ACE2 Binding Inhibition.

BACKGROUND: Mucosal antibodies play a critical role in preventing SARS-CoV-2 infections or reinfections by blocking the interaction of the receptor-binding domain (RBD) with the angiotensin-converting enzyme 2 (ACE2) receptor on the cell surface. In this study, we investigated the difference between the mucosal antibody response after primary infection and vaccination. METHODS: We assessed longitudinal changes in the quantity and capacity of nasal antibodies to neutralize the interaction of RBD with the ACE2 receptor using the spike protein and RBD from ancestral SARS-CoV-2 (Wuhan-Hu-1), as well as the RBD from the Delta and Omicron variants. RESULTS: Significantly higher mucosal IgA concentrations were detected postinfection vs postvaccination, while vaccination induced higher IgG concentrations. However, ACE2-inhibiting activity did not differ between the cohorts. Regarding whether IgA or IgG drove ACE2 inhibition, infection-induced binding inhibition was driven by both isotypes, while postvaccination binding inhibition was mainly driven by IgG. CONCLUSIONS: Our study provides new insights into the relationship between antibody isotypes and neutralization by using a sensitive and high-throughput ACE2 binding inhibition assay. Key differences are highlighted between vaccination and infection at the mucosal level, showing that despite differences in the response quantity, postinfection and postvaccination ACE2 binding inhibition capacity did not differ.

Diagnosis of childhood febrile illness using a multi-class blood RNA molecular signature.

BACKGROUND: Appropriate treatment and management of children presenting with fever depend on accurate and timely diagnosis, but current diagnostic tests lack sensitivity and specificity and are frequently too slow to inform initial treatment. As an alternative to pathogen detection, host gene expression signatures in blood have shown promise in discriminating several infectious and inflammatory diseases in a dichotomous manner. However, differential diagnosis requires simultaneous consideration of multiple diseases. Here, we show that diverse infectious and inflammatory diseases can be discriminated by the expression levels of a single panel of genes in blood. METHODS: A multi-class supervised machine-learning approach, incorporating clinical consequence of misdiagnosis as a "cost" weighting, was applied to a whole-blood transcriptomic microarray dataset, incorporating 12 publicly available datasets, including 1,212 children with 18 infectious or inflammatory diseases. The transcriptional panel identified was further validated in a new RNA sequencing dataset comprising 411 febrile children. FINDINGS: We identified 161 transcripts that classified patients into 18 disease categories, reflecting individual causative pathogen and specific disease, as well as reliable prediction of broad classes comprising bacterial infection, viral infection, malaria, tuberculosis, or inflammatory disease. The transcriptional panel was validated in an independent cohort and benchmarked against existing dichotomous RNA signatures. CONCLUSIONS: Our data suggest that classification of febrile illness can be achieved with a single blood sample and opens the way for a new approach for clinical diagnosis. FUNDING: European Union's Seventh Framework no. 279185; Horizon2020 no. 668303 PERFORM; Wellcome Trust (206508/Z/17/Z); Medical Research Foundation (MRF-160-0008-ELP-KAFO-C0801); NIHR Imperial BRC.

Impact of infection on proteome-wide glycosylation revealed by distinct signatures for bacterial and viral pathogens.

Mechanisms of infection and pathogenesis have predominantly been studied based on differential gene or protein expression. Less is known about posttranslational modifications, which are essential for protein functional diversity. We applied an innovative glycoproteomics method to study the systemic proteome-wide glycosylation in response to infection. The protein site-specific glycosylation was characterized in plasma derived from well-defined controls and patients. We found 3862 unique features, of which we identified 463 distinct intact glycopeptides, that could be mapped to more than 30 different proteins. Statistical analyses were used to derive a glycopeptide signature that enabled significant differentiation between patients with a bacterial or viral infection. Furthermore, supported by a machine learning algorithm, we demonstrated the ability to identify the causative pathogens based on the distinctive host blood plasma glycopeptide signatures. These results illustrate that glycoproteomics holds enormous potential as an innovative approach to improve the interpretation of relevant biological changes in response to infection.

Adoption of C-reactive protein point-of-care tests for the management of acute childhood infections in primary care in the Netherlands and England: a comparative health systems analysis.

BACKGROUND: The use of point of care (POC) tests varies across Europe, but research into what drives this variability is lacking. Focusing on CRP POC tests, we aimed to understand what factors contribute to high versus low adoption of the tests, and also to explore whether they are used in children. METHODS: We used a comparative qualitative case study approach to explore the implementation of CRP POC tests in the Netherlands and England. These countries were selected because although they have similar primary healthcare systems, the availability of CRP POC tests in General Practices is very different, being very high in the former and rare in the latter. The study design and analysis were informed by the non-adoption, abandonment, spread, scale-up and sustainability (NASSS) framework. Data were collected through a review of documents and interviews with stakeholders. Documents were identified through a scoping literature review, search of websites, and stakeholder recommendation. Stakeholders were selected purposively initially, and then by snowballing. Data were analysed thematically. RESULTS: Sixty-five documents were reviewed and 21 interviews were conducted. The difference in the availability of CRP POC tests is mainly because of differences at the wider national context level. In the two countries, early adopters of the tests advocated for their implementation through the generation of robust evidence and by engaging with all relevant stakeholders. This led to the inclusion of CRP POC tests in clinical guidelines in both countries. In the Netherlands, this mandated their reimbursement in accordance with Dutch regulations. Moreover, the prevailing better integration of health services enabled operational support from laboratories to GP practices. In England, the funding constraints of the National Health Service and the prioritization of alternative and less expensive antimicrobial stewardship interventions prevented the development of a reimbursement scheme. In addition, the lack of integration between health services limits the operational support to GP practices. In both countries, the availability of CRP POC tests for the management of children is a by-product of the test being available for adults. The tests are less used in children mainly because of concerns regarding their accuracy in this age-group. CONCLUSIONS: The engagement of early adopters combined with a more favourable and receptive macro level environment, including the role of clinical guidelines and their developers in determining which interventions are reimbursed and the operational support from laboratories to GP practices, led to the greater adoption of the tests in the Netherlands. In both countries, CRP POC tests, when available, are less used less in children. Organisations considering introducing POC tests into primary care settings need to consider how their implementation fits into the wider health system context to ensure achievable plans.

SARS-CoV-2 mucosal antibody development and persistence and their relation to viral load and COVID-19 symptoms.

Although serological studies have shown that antibodies against SARS-CoV-2 play an important role in protection against (re)infection, the dynamics of mucosal antibodies during primary infection and their potential impact on viral load and the resolution of disease symptoms remain unclear. During the first pandemic wave, we assessed the longitudinal nasal antibody response in index cases with mild COVID-19 and their household contacts. Nasal and serum antibody responses were analysed for up to nine months. Higher nasal receptor binding domain and spike protein-specific antibody levels at study inclusion were associated with lower viral load. Older age was correlated with more frequent COVID-19 related symptoms. Receptor binding domain and spike protein-specific mucosal antibodies were associated with the resolution of systemic, but not respiratory symptoms. Finally, receptor binding domain and spike protein-specific mucosal antibodies remained elevated up to nine months after symptom onset.

Biosynthetic homeostasis and resilience of the complement system in health and infectious disease.

BACKGROUND: The complement system is a central component of the innate immune system. Constitutive biosynthesis of complement proteins is essential for homeostasis. Dysregulation as a consequence of genetic or environmental cues can lead to inflammatory syndromes or increased susceptibility to infection. However, very little is known about steady state levels in children or its kinetics during infection. METHODS: With a newly developed multiplex mass spectrometry-based method we analyzed the levels of 32 complement proteins in healthy individuals and in a group of pediatric patients infected with bacterial or viral pathogens. FINDINGS: In plasma from young infants we found reduced levels of C4BP, ficolin-3, factor B, classical pathway components C1QA, C1QB, C1QC, C1R, and terminal pathway components C5, C8, C9, as compared to healthy adults; whereas the majority of complement regulating (inhibitory) proteins reach adult levels at very young age. Both viral and bacterial infections in children generally lead to a slight overall increase in complement levels, with some exceptions. The kinetics of complement levels during invasive bacterial infections only showed minor changes, except for a significant increase and decrease of CRP and clusterin, respectively. INTERPRETATION: The combination of lower levels of activating and higher levels of regulating complement proteins, would potentially raise the threshold of activation, which might lead to suppressed complement activation in the first phase of life. There is hardly any measurable complement consumption during bacterial or viral infection. Altogether, expression of the complement proteins appears surprisingly stable, which suggests that the system is continuously replenished. FUND: European Union's Horizon 2020, project PERFORM, grant agreement No. 668303.

Nationwide Study on the Course of Influenza A (H1N1) Infections in Hospitalized Children in the Netherlands During the Pandemic 2009-2010.

BACKGROUND: The influenza H1N1 pandemic of 2009-2010, provided a unique opportunity to assess the course of disease, as well as the analysis of risk factors for severe disease in hospitalized children (< 18 years). METHODS: Retrospective national chart study on hospitalized children with H1N1 infection during the 2009-2010 pH1N1 outbreak. RESULTS: Nine hundred forty patients (56% boys), median age 3.0 years, were enrolled; the majority were previously healthy. Treatment consisted of supplemental oxygen (24%), mechanical ventilation (5%) and antiviral therapy (63%). Fifteen patients died (1.6%), 5 of whom were previously healthy. Multivariable analyses confirmed pre-existent heart and lung disease as risk factors for intensive care unit admission. Risk factors for mortality included children with a neurologic or oncologic disease and psychomotor retardation. CONCLUSIONS: This nationwide overview of hospitalized children confirms known risk groups for severe influenza infections. However, most of the acute and severe presentations of influenza occurred in previously healthy children.

Mortality and morbidity in community-acquired sepsis in European pediatric intensive care units: a prospective cohort study from the European Childhood Life-threatening Infectious Disease Study (EUCLIDS).

BACKGROUND: Sepsis is one of the main reasons for non-elective admission to pediatric intensive care units (PICUs), but little is known about determinants influencing outcome. We characterized children admitted with community-acquired sepsis to European PICUs and studied risk factors for mortality and disability. METHODS: Data were collected within the collaborative Seventh Framework Programme (FP7)-funded EUCLIDS study, which is a prospective multicenter cohort study aiming to evaluate genetic determinants of susceptibility and/or severity in sepsis. This report includes 795 children admitted with community-acquired sepsis to 52 PICUs from seven European countries between July 2012 and January 2016. The primary outcome measure was in-hospital death. Secondary outcome measures were PICU-free days censured at day 28, hospital length of stay, and disability. Independent predictors were identified by multivariate regression analysis. RESULTS: Patients most commonly presented clinically with sepsis without a source (n = 278, 35%), meningitis/encephalitis (n = 182, 23%), or pneumonia (n = 149, 19%). Of 428 (54%) patients with confirmed bacterial infection, Neisseria meningitidis (n = 131, 31%) and Streptococcus pneumoniae (n = 78, 18%) were the main pathogens. Mortality was 6% (51/795), increasing to 10% in the presence of septic shock (45/466). Of the survivors, 31% were discharged with disability, including 24% of previously healthy children who survived with disability. Mortality and disability were independently associated with S. pneumoniae infections (mortality OR 4.1, 95% CI 1.1-16.0, P = 0.04; disability OR 5.4, 95% CI 1.8-15.8, P < 0.01) and illness severity as measured by Pediatric Index of Mortality (PIM2) score (mortality OR 2.8, 95% CI 1.3-6.1, P < 0.01; disability OR 3.4, 95% CI 1.8-6.4, P < 0.001). CONCLUSIONS: Despite widespread immunization campaigns, invasive bacterial disease remains responsible for substantial morbidity and mortality in critically ill children in high-income countries. Almost one third of sepsis survivors admitted to the PICU were discharged with some disability. More research is required to delineate the long-term outcome of pediatric sepsis and to identify interventional targets. Our findings emphasize the importance of improved early sepsis-recognition programs to address the high burden of disease.

Mice deficient for soluble adenylyl cyclase are infertile because of a severe sperm-motility defect.

To acquire the ability to fertilize, spermatozoa undergo complex, but at present poorly understood, activation processes. The intracellular rise of cAMP produced by the bicarbonate-dependent soluble adenylyl cyclase (sAC) has been suggested to play a central role in initiating the cascade of the events that culminates in spermatozoa maturation. Here, we show that targeted disruption of the sAC gene does not affect spermatogenesis but dramatically impairs sperm motility, leading to male sterility. sAC mutant spermatozoa are characterized by a total loss of forward motility and are unable to fertilize oocytes in vitro. Interestingly, motility in sAC mutant spermatozoa can be restored on cAMP loading, indicating that the motility defect observed is not caused by a structural defect. We, therefore, conclude that sAC plays an essential and nonredundant role in the activation of the signaling cascade controlling motility and, therefore, in fertility. The crucial role of sAC in fertility and the absence of any other obvious pathological abnormalities in sAC-deficient mice may provide a rationale for developing inhibitors that can be applied as a human male contraceptive.