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Contains fulltext : 293706.pdf (Publisher’s version ) (Open Access)

Background The rs72824905 single-nucleotide polymorphism in the PLCG2 gene, encoding the p.P522R residue change in Phospholipase C gamma 2 (PLCγ2), associates with protection against several dementia subtypes and with increased likelihood of longevity. Cell lines and animal models indicated that p.P522R is a functional hypermorph. We aimed to confirm this in human circulating peripheral immune cells. Methods We compared effects of p.P522R on immune system function between carriers and non-carriers (aged 59-103y), using in-depth immunophenotyping, functional B-cell and myeloid cell assays, and in vivo SARS-CoV-2 vaccination. Results In line with expectations, p.P522R impacts immune cell function only slightly, but it does so across a wide array of immune cell types. Upon B-cell stimulation, we observed increased PLCγ2 phosphorylation and calcium release, suggesting increased B-cell sensitivity upon antigen recognition. Further, p.P522R-carriers had higher numbers of CD20++CD21-CD24+ naive B cells and IgG1+ memory B cells. In myeloid cells, normalized ROS production was higher upon PLCγ2-dependent stimulation. On classical monocytes, CD33 levels were elevated. Furthermore, carriers expressed lower levels of allergy-related FcεRI on several immune cell subsets. Nevertheless, carriers and non-carriers had similar serological responses to SARS-CoV-2 vaccination. Conclusion The immune system from p.P522R-carriers is slightly more responsive to stimulation than in non-carriers.

Background Older age is associated with increased severity and death from respiratory infections, including coronavirus disease 2019 (COVID-19). The tuberculosis BCG vaccine may provide heterologous protection against nontuberculous infections and has been proposed as a potential preventive strategy against COVID-19. Methods In this multicenter, placebo-controlled trial, we randomly assigned older adults (aged ≥60 years; n = 2014) to intracutaneous vaccination with BCG vaccine (n = 1008) or placebo (n = 1006). The primary end point was the cumulative incidence of respiratory tract infections (RTIs) that required medical intervention, during 12 months of follow-up. Secondary end points included the incidence of COVID-19, and the effect of BCG vaccination on the cellular and humoral immune responses. Results The cumulative incidence of RTIs requiring medical intervention was 0.029 in the BCG-vaccinated group and 0.024 in the control group (subdistribution hazard ratio, 1.26 [98.2% confidence interval, .65–2.44]). In the BCG vaccine and placebo groups, 51 and 48 individuals, respectively tested positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with polymerase chain reaction (subdistribution hazard ratio, 1.053 [95% confidence interval, .71–1.56]). No difference was observed in the frequency of adverse events. BCG vaccination was associated with enhanced cytokine responses after influenza, and also partially associated after SARS-CoV-2 stimulation. In patients diagnosed with COVID-19, antibody responses after infection were significantly stronger if the volunteers had previously received BCG vaccine. Conclusions BCG vaccination had no effect on the incidence of RTIs, including SARS-CoV-2 infection, in older adult volunteers. However, it improved cytokine responses stimulated by influenza and SARS-CoV-2 and induced stronger antibody titers after COVID-19 infection. Clinical trials registration EU Clinical Trials Register 2020-001591-15 ClinicalTrials.gov NCT04417335.

Contains fulltext : 291405.pdf (Publisher’s version ) (Open Access) INTRODUCTION: Several studies have shown that intradermal vaccination leads to improved immune responses. In addition, lowering vaccine doses will reduce costs and therefore potentially increase coverage. To determine whether intradermal delivery enhances the antibody responses against the 13-valent pneumococcal conjugate vaccine (PCV13), we compared intradermally and intramuscularly vaccinated mice. METHODS: Mice were immunized with PCV13, either intradermally or intramuscularly and CFU-counts in the nasal tissue were determined three or seven days after intranasal colonization with a serotype 4 clinical strain. Antibody concentrations against all thirteen polysaccharides were measured in blood and mucosal samples using a fluorescent-bead-based multiplex immunoassay. RESULTS: Antibody levels in both serum and mucosal samples were higher in the intramuscularly vaccinated group as compared to the intradermally vaccinated group. No protection against S. pneumoniae intranasal colonization was observed for either vaccination route. CONCLUSIONS: Intradermal vaccination was inferior to intramuscular immunization in inducing serotype-specific antibodies.

Background Nasopharyngeal swabs are taken to determine the causative agent of community acquired pneumonia (CAP), while the reliability of upper respiratory tract sampling as a proxy for lower respiratory tract infections is still unclear. Methods Nasopharyngeal (NP) swabs, bronchoalveolar lavage (BAL) fluid samples and clinical data were collected from 153 hospitalized children between 3 months and 14 years of age with severe CAP, enrolled from March to June 2019. Written informed consent for the storage and use of the samples for further studies was obtained from the parents or caregivers. Putative pathogens were detected using a sensitive, high-throughput GeXP-based multiplex PCR and qPCR. Results The same bacterial species in paired samples were found in 29 (23.4%) and the same viral species in 52 (27.5%) of the patients. moderate concordance was found for Mycoplasma pneumoniae (ĸ=0.64), followed by Haemophilus influenzae (ĸ=0.42). The strongest discordance was observed for human adenovirus and also for Pseudomonas aeruginosa, the latter was exclusively detected in BAL samples. In the adenovirus cases strong concordance was associated with high viral loads in the NP swabs. Conclusion The variation in concordance in pathogen detection in the upper and lower respiratory tract of children with severe pneumonia is generally high but varies depending on the species. Novel and impactful insights are the concordance between NP and BAL detection for M. pneumoniae and H. influenzae and the strong correlation between high adenoviral loads in NP swabs and detection in BAL fluid.

Background:Phospholipase C gamma 2 (PLCγ2) is encoded by thePLCG2gene. A single-nucleotide polymorphism (p.P522R) associates with protection against several dementia subtypes and with increased likelihood of longevity. Cell lines and animal models indicated that p.P522R is a functional hypermorph. We aimed to confirm this in human peripheral immune cells.Methods:We compared effects of p.P522R on immune system function between carriers and non-carriers (aged 59-103y), using in-depth immunophenotyping, functional B-cell and myeloid-cell assays, andin vivoSARS-CoV-2 vaccination.Results:As expected, effects of p.P522R on immune cell function were small. Immune cell numbers in p.P522R carriers better resembled a younger reference cohort than those of non-carriers. Moreover, carriers expressed lower levels of FcεRI on several immune cell subsets and elevated CD33 levels on classical monocytes. Upon B-cell stimulation, PLCγ2 phosphorylation and calcium release were increased in carriers compared to non-carriers. Normalized ROS production in myeloid cells was higher upon PLCγ2-dependent stimulation, but lower upon PLCγ2-independent stimulation. Carriers and non-carriers had similar serological responses to SARS-CoV-2 vaccination.Conclusion:Compared to non-carriers, immune profiles from carriers more closely resembled those from younger individuals, suggesting that p.P522R associates with resilience against immunological aging.

A range of public health measures have been implemented to suppress local transmission of coronavirus disease 2019 (COVID-19) in Shenzhen. We examined the effect of these measures on the prevalence of respiratory pathogens in children. Clinical and respiratory pathogen data were collected for routine care from hospitalized children with acute respiratory infections in Shenzhen Children’s Hospital from July 2018 to January 2022. Nasopharyngeal swabs were collected and respiratory pathogens were detected using standardized clinical diagnostics as part of routine care. Data were analyzed to describe the effects of COVID-19 prevention procedures on other common pathogens. A total of 56,325 children under 14 years of age were hospitalized with an acute respiratory infection during the study period, 33,909 were tested from July 2018 to January 2020 (pre-lockdown), 1168 from February 2020 to May 2020 (lockdown) and 21,248 from July 2020 to January 2022 (post-lockdown). We observed a 37.3% decline of routine care in respiratory infection associated hospital admission in the 19 months’ post-lockdown vs. the 19 months’ pre-lockdown. There were 99.4%, 16.0% and 1.26% reductions measured for Mycoplasma pneumoniae, influenza virus A and adenovirus, respectively. However, a 118.7% and 75.8% rise was found for respiratory syncytial virus (RSV) and human para-influenza virus (HPIV) during the 19 months’ post-lockdown in comparison to the pre-pandemic period. The detection of RSV especially increased in toddlers after the lockdown. Lockdown measures during the COVID-19 pandemic led to a significant reduction of Mycoplasma pneumoniae, influenza virus A and adenovirus infection. In contrast, RSV and HPIV infection increased.

Purpose of review The human upper respiratory tract is colonized with a variety of bacterial microorganisms including Haemophilus influenzae. The species H. influenzae consists of typeable and nontypeable H. influenzae (NTHi) variants. Typeable H. influenzae are subdivided into types a through f, based on the polysaccharide capsule, whereas the NTHi strains do not express a polysaccharide capsule. In this review, we highlight the current advances in the field of H. influenzae, with the focus on bacterial virulence mechanisms that facilitate bacterial colonization and disease, particularly for NTHi. Recent findings In the past decade, it has become apparent that NTHi has the ability to cause invasive infections. Recently, a number of adhesins have been shown to be crucial for bacterial colonization and invasion and these proteins were investigated as vaccine antigens. Although NTHi lacks a polysaccharide capsule, it expresses lipooligosaccharide that contribute to adhesion and evasion of complement-mediated killing, both contributing to bacterial virulence, which could potentially be targeted by novel antimicrobial drugs or vaccines. Summary The unraveling of H. influenzae virulence mechanisms resulted in the identification of promising targets for novel antimicrobials and vaccine antigens aiming to prevent or treat both typeable and nontypeable H. influenzae infections.

Objective: To describe the treatment of community-acquired pneumonia (CAP) in children under five years in Tanzania. Methods: Between January and December 2017, children aged 2–59 months with chest radiography-confirmed CAP were enrolled. The parents were interviewed to collect information on the patients and home-based medication. Clinical information was derived from the patient files. Nasopharyngeal swab and blood samples were collected for isolation of the causative pathogens. Swab samples were analysed by quantitative PCR whereas blood samples were tested using BacT/Alert 3D. Results: Overall, 109 children with CAP were included in this analysis. Provision of care to most children was delayed (median = 4.6 days). A quarter (26.6%) were given unprescribed/leftover antibiotics at home. Only one child had positive bacterial culture. Referrals were associated with nasopharyngeal carriage of Streptococcus pneumoniae (p = 0.003) and Haemophilus influenzae (p = 0.004). Of all admitted children, more than a quarter (n = 29) did not need to be hospitalised and inappropriately received injectable instead of oral antibiotics. Conclusion: We found high rates of home treatment, particularly with antibiotics. Appropriate health care was delayed for most children because of home treatment. Efforts are needed at the community level to improve awareness of antimicrobial resistance. Keywords: Home treatment, Community-acquired pneumonia, Children under five years, Delays, Unnecessary hospitalisation

Cystic fibrosis (CF) is a common autosomal recessive disease causing thick, viscous secretions leading to pulmonary infections with pathogenic bacteria. As part of routine patient care, colonization and infection with these bacteria is monitored with cough swab or sputum cultures and sometimes bronchoalveolar lavage. In this cross-sectional proof-of-concept study in a cohort of CF patients we collected swabs or sputa and exhaled breath samples with the modular breath sampler (MBS), a newly developed two-way non-rebreathing sampling device. Pathogen specific polymerase chain reactions (PCRs) were performed on the MBS samples and compared with the results obtained with conventional diagnostics (i.e. culturing of swabs and sputa). A control group of stable asthma patients was used as negative control for the MBS measurements. The pathogens detected using MBS and conventional culturing differed: S. aureus was found more often in swab or sputum samples whereas Pseudomonas aeruginosa and S. pneumoniae were found more often in MBS samples. We hypothesize that this is due to sampling of different compartments, MBS samples are derived from the lower respiratory tract while cultures from cough swabs and sputa are dominated by pathogens residing in the upper respiratory tract. Another important difference is the readout, i.e. culture versus PCR. The majority of CF patients in whom P. aeruginosa was found did not have recent positive cultures suggesting higher sensitivity of MBS-based than conventional diagnostics. The majority of parents/patients found the MBS easy to use and less of a burden than respiratory sampling.

CCR5 is the main HIV co-receptor. We aimed to (1) compare CCR5 expression on immune cells between people living with HIV (PLHIV) using combination antiretroviral therapy (cART) and HIV-uninfected controls, (2) relate CCR5 expression to viral reservoir size and (3) assess determinants of CCR5 expression. This cross-sectional study included 209 PLHIV and 323 controls. Percentages of CCR5+ cells (%) and CCR5 mean fluorescence intensity assessed by flow cytometry in monocytes and lymphocyte subsets were correlated to host factors, HIV-1 cell-associated (CA)-RNA and CA-DNA, plasma inflammation markers and metabolites. Metabolic pathways were identified. PLHIV displayed higher percentages of CCR5+ monocytes and several CD8+ T cell subsets, but lower percentages of CCR5+ naive CD4+ T cells and regulatory T cells (Tregs). HIV-1 CA-DNA and CA-RNA correlated positively with percentages of CCR5+ lymphocytes. Metabolome analysis revealed three pathways involved in energy metabolism associated with percentage of CCR5+ CD8+ T cells in PLHIV. Our results indicate that CCR5 is differently expressed on various circulating immune cells in PLHIV. Hence, cell-trafficking of CD8+ T cells and Tregs may be altered in PLHIV. Associations between energy pathways and percentage of CCR5+ CD8+ T cells in PLHIV suggest higher energy demand of these cells in PLHIV.

Bordetella pertussis (Bp), the causative agent of pertussis, continues to circulate despite widespread vaccination programs. An important question is whether and how (sub)clinical infections shape immune memory to Bp, particularly in populations primed with acellular pertussis vaccines (aP). Here, we examine the prevalence of mucosal antibodies against non-vaccine antigens in aP-primed children and adolescents of the BERT study (NCT03697798), using antibody binding to a Bp mutant strain lacking aP antigens (Bp_mut). Our study identifies increased levels of mucosal IgG and IgA binding to Bp_mut in older aP-primed individuals, suggesting different Bp exposure between aP-primed birth cohorts, in line with pertussis disease incidence data. To examine whether Bp exposure influences vaccination responses, we measured mucosal antibody responses to aP booster vaccination as a secondary study outcome. Although booster vaccination induces significant increases in mucosal antibodies to Bp in both cohorts, the older age group that had higher baseline antibodies to Bp_ mut shows increased persistence of antibodies after vaccination.

Contains fulltext : 248218.pdf (Publisher’s version ) (Open Access) In situ labelling and encapsulation of biological entities, such as of single viruses, may provide a versatile approach to modulate their functionality and facilitate their detection at single particle level. Here, we introduce a novel virus metallization approach based on in situ coating of viruses in solution with silver nanoparticles (AgNP) in a two-step synthetic process, i.e. surface activation with a tannic acid - Sn(II) coordination complex, which subsequently induces silver ion (I) reduction. The metalic coating on the virus surface opens the opportunity for electrochemical quantification of the AgNP-tagged viruses by nano-impact electrochemistry on a microelectrode with single particle sensitivity, i.e. enable the detection of particles oherwise undetectable. We show that the silver coating of the virus particles impacting the electrode can be oxidized to produce distinct current peaks the frequency of which show a linear correlation with the virus count. The proof of the concept was done with inactivated Influenza A (H3N2) viruses resulting in their quantitation down to the femtomolar concentrations (ca. 5 × 10(7) particles per mL) using 50 s counting sequences.

Synthetic sugar analogs are widely applied in metabolic oligosaccharide engineering (MOE) and as novel drugs to interfere with glycoconjugate biosynthesis. However, mechanistic insights on their exact cellular metabolism over time are mostly lacking. We combined ion-pair UHPLC-QqQ mass spectrometry using tributyl- and triethylamine buffers for sensitive analysis of sugar metabolites in cells and organisms and identified low abundant nucleotide sugars, such as UDP-arabinose in human cell lines and CMP-sialic acid (CMP-NeuNAc) in Drosophila. Furthermore, MOE revealed that propargyloxycarbonyl (Poc) labeled ManNPoc was metabolized to both CMP-NeuNPoc and UDP-GlcNPoc. Finally, time-course analysis of the effect of antitumor compound 3Fax-NeuNAc by incubation of B16-F10 melanoma cells with N-acetyl-D-[UL-13C6]glucosamine revealed full depletion of endogenous ManNAc 6-phosphate and CMP-NeuNAc within 24 hour. Thus, dynamic tracing of sugar metabolic pathways provides a general approach to reveal time-dependent insights into the metabolism of synthetic sugars, which is important for the rational design of analogs with optimized effects., Glycobiology, 32 (3), ISSN:0959-6658

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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

Congenital disorders of glycosylation (CDG) are inherited metabolic diseases characterized by mutations in enzymes involved in different steps of protein glycosylation, leading to aberrant synthesis, attachment or processing of glycans. Recently, immunological dysfunctions in several CDG types have been increasingly documented. Despite these observations, detailed studies on immune cell dysfunction in PMM2-CDG and other CDG types are still scarce. Studying PMM2-CDG patient immune cells is challenging due to limited availability of patient material, which is a result of the low incidence of the disease and the often young age of the subjects. Dedicated immune cell models, mimicking PMM2-CDG, could circumvent many of these problems and facilitate research into the mechanisms of immune dysfunction. Here we provide initial observations about the immunophenotype and the phagocytic function of primary PMM2-CDG monocytes. Furthermore, we assessed the suitability of two different glycosylation-impaired human monocyte models: tunicamycin-treated THP-1 monocytes and PMM2 knockdown THP-1 monocytes induced by shRNAs. We found no significant differences in primary monocyte subpopulations of PMM2-CDG patients as compared to healthy individuals but we did observe anomalous surface glycosylation patterns in PMM2-CDG patient monocytes as determined using fluorescent lectin binding. We also looked at the capacity of monocytes to bind and internalize fungal particles and found a slightly increased uptake of C. albicans by PMM2-CDG monocytes as compared to healthy monocytes. Tunicamycin-treated THP-1 monocytes showed a highly decreased uptake of fungal particles, accompanied by a strong decrease in glycosylation levels and a high induction of ER stress. In contrast and despite a drastic reduction of the PMM2 enzyme activity, PMM2 knockdown THP-1 monocytes showed no changes in global surface glycosylation levels, levels of fungal particle uptake similar to control monocytes, and no ER stress induction. Collectively, these initial observations suggest that the absence of ER stress in PMM2 knockdown THP-1 cells make this model superior over tunicamycin-treated THP-1 cells and more comparable to primary PMM2-CDG monocytes. Further development and exploitation of CDG monocyte models will be essential for future in-depth studies to ultimately unravel the mechanisms of immune dysfunction in CDG.

Knowledge about contagiousness is key to accurate management of hospitalized COVID-19 patients. Epidemiological studies suggest that in addition to transmission through droplets, aerogenic SARS-CoV-2 transmission contributes to the spread of infection. However, the presence of virus in exhaled air has not yet been sufficiently demonstrated. In pandemic situations low tech disposable and user-friendly bedside devices are required, while commercially available samplers are unsuitable for application in patients with respiratory distress. We included 49 hospitalized COVID-19 patients and used a disposable modular breath sampler to measure SARS-CoV-2 RNA load in exhaled air samples and compared these to SARS-CoV-2 RNA load of combined nasopharyngeal throat swabs and saliva. Exhaled air sampling using the modular breath sampler has proven feasible in a clinical COVID-19 setting and demonstrated viral detection in 25% of the patients.

Contains fulltext : 248219.pdf (Publisher’s version ) (Open Access) Trained immunity is a de facto memory of innate immune cells, resulting in a long-term increase in innate host defense mechanisms after infection. The long-term heterologous protection conferred by trained immunity is mediated through epigenetic and functional reprogramming of hematopoietic stem and progenitor cells. Because the spleen is a reservoir of undifferentiated monocytes and is considered the prime organ for extramedullary hematopoiesis, we investigated the role of the spleen in the establishment of trained immunity. A β-glucan-induced trained immunity mouse model was performed in previously sham-operated or splenectomized animals. Removal of the spleen did not modulate the proinflammatory cytokine production of in vivo trained peritoneal cells, nor did it ablate the increased percentage of proinflammatory circulatory monocytes and natural killer cells seen in trained animals. However, spleen removal prevented neutrophilia, an important characteristic of trained immunity. These data point to a limited role of the spleen in trained immunity. The pathophysiologic relevance of the spleen in the induction of neutrophilia during trained immunity remains to be fully explored.

The majority of COVID-19 patients experience mild to moderate disease course and recover within a few weeks. An increasing number of studies characterized the long-term changes in the specific anti-SARS-CoV-2 immune responses, but how COVID-19 shapes the innate and heterologous adaptive immune system after recovery is less well known. To comprehensively investigate the post-SARS-CoV-2 infection sequelae on the immune system, we performed a multi-omics study by integrating single-cell RNA-sequencing, single-cell ATAC-sequencing, genome-wide DNA methylation profiling, and functional validation experiments in 14 convalescent COVID-19 and 15 healthy individuals. We showed that immune responses generally recover without major sequelae after COVID-19. However, subtle differences persist at the transcriptomic level in monocytes, with downregulation of the interferon pathway, while DNA methylation also displays minor changes in convalescent COVID-19 individuals. However, these differences did not affect the cytokine production capacity of PBMCs upon different bacterial, viral, and fungal stimuli, although baseline release of IL-1Ra and IFN-γ was higher in convalescent individuals. In conclusion, we propose that despite minor differences in epigenetic and transcriptional programs, the immune system of convalescent COVID-19 patients largely recovers to the homeostatic level of healthy individuals.

Background Recent research suggests that the microbiota affects susceptibility to both respiratory tract infections (RTIs) and gastrointestinal infections (GIIs). In order to optimize global treatment options, it is important to characterize microbiota profiles across different niches and geographic/socioeconomic areas where RTI and GII prevalences are high. Methods We performed 16S sequencing of nasopharyngeal swabs from 209 Venezuelan Amerindian children aged 6 weeks–59 months who were participating in a 13-valent pneumococcal conjugate vaccine (PCV13) study. Using random forest models, differential abundance testing, and regression analysis, we determined whether specific bacteria were associated with RTIs or GIIs and variation in PCV13 response. Results Microbiota compositions differed between children with or without RTIs (P = .018) or GIIs (P = .001). Several species were associated with the absence of infections. Some of these health-associated bacteria are also observed in developed regions, such as Corynebacterium (log2(fold change [FC]) = 3.30 for RTIs and log2(FC) = 1.71 for GIIs), while others are not commonly observed in developed regions, such as Acinetobacter (log2(FC) = 2.82 and log2(FC) = 5.06, respectively). Klebsiella spp. presence was associated with both RTIs (log2(FC) = 5.48) and GIIs (log2(FC) = 7.20). Conclusions The nasopharyngeal microbiota of rural Venezuelan children included several bacteria that thrive in tropical humid climates. Interestingly, nasopharyngeal microbiota composition not only differed in children with an RTI but also in those with a GII, which suggests a reciprocal interplay between the 2 environments. Knowledge of region-specific microbiota patterns enables tailoring of preventive and therapeutic approaches., The nasopharyngeal microbiota of rural Venezuelan children included several bacteria that thrive in tropical humid climates. Nasopharyngeal microbiota composition not only differed in children with a respiratory infection but also in those with a gastrointestinal infection.

The last decade has witnessed a renewed interest in space exploration. Public and private institutions are investing considerable effort toward the direct exploration of the Moon and Mars, as well as more distant bodies in the solar system. Both automated and human-crewed spacecraft are being considered in these efforts. As inevitable fellow travelers on the bodies of astronauts, spaceships, or equipment, terrestrial microorganisms will undoubtedly come into contact with extraterrestrial environments, despite stringent decontamination. These microorganisms could eventually adapt and grow in their new habitats, where they might potentially recolonize and lead to the infection of the human space travelers. In this article, we demonstrate that clinically relevant bacterial species found in the environment are able to grow in minimal media with sugar compounds identified in extraterrestrial carbon sources. As a surrogate model, we used carbohydrates previously isolated from carbonaceous meteorites. The bacteria underwent an adaptation process that caused structural modifications in the cell envelope that sparked changes in pathogenic potential, both in vitro and in vivo. Understanding the adaptation of microorganisms exposed to extraterrestrial environments, with subsequent changes in their immunogenicity and virulence, requires a comprehensive analysis of such scenarios to ensure the safety of major space expeditions in the decades to come.

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.

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. Keywords: Targeted mass spectrometry, Multiple reaction monitoring (MRM), Complement system, Infectious disease, C-reactive protein (CRP), Clusterin

Despite the availability of vaccines and antibiotics, pneumonia remains the leading cause of mortality among children under 5 years of age. The objective of this study was to identify modifiable risk factors for community-acquired pneumonia (CAP) in children under 5 years of age in a vaccinated population.A case-control study was conducted between January and December 2017. The cases included children aged 2-59 months with X-ray-confirmed pneumonia, whereas the controls were children from the community with no history of respiratory infection. A multivariable logistic regression model was used to determine the modifiable risk factors for CAP.A total of 113 children with X-ray-confirmed pneumonia and 350 healthy children were enrolled in this study. The median ages for the cases and controls were 13.7 (IQR = 7.2-25.3) and 13.4 (IQR = 6.0-24.8) months respectively. One (0.9%) child died after the enrolment. The independent predictors of CAP included a lack of exclusive breastfeeding for 6 months (aOR = 1.7, 95% CI = 1.0-2.9), underweight (aOR = 2.1, 95% CI = 1.0-4.5), unclean cooking fuel (aOR = 1.8, 95% CI = 1.0-3.3) and low income (aOR = 2.9, 95% CI = 1.6-5.4). No association was found between vaccination status and CAP.In addition to a lack of exclusive breastfeeding, children from families of low-economic status were at risk of contracting CAP. Since the risk factors are complex, the study results call for more concerted efforts by and collaboration among the health, agriculture and development sectors to address mortality caused by CAP.Malgré la disponibilité des vaccins et des antibiotiques, la pneumonie reste la principale cause de mortalité chez les enfants de moins de cinq ans. L'objectif de cette étude était d'identifier les facteurs de risque modifiables pour la pneumonie acquise en communauté (PAC) chez les enfants de moins de cinq ans dans une population vaccinée. MÉTHODES: Une étude cas-témoins a été menée entre janvier et décembre 2017. Les cas concernaient des enfants âgés de 2 à 59 mois atteints de pneumonie confirmée par la radiographie, alors que les témoins étaient des enfants de la communauté sans antécédents d'infection respiratoire. Un modèle de régression logistique multivariée a été utilisé pour déterminer les facteurs de risque modifiables pour la PAC. RÉSULTATS: Au total, 113 enfants atteints de pneumonie confirmée par la radiographie et 350 enfants en bonne santé ont été inclus dans cette étude. Les âges médians pour les cas et les témoins étaient respectivement de 13,7 (IQR = 7,2 - 25,3) et de 13,4 (IQR = 6,0 - 24,8) mois. Un enfant (0,9%) est décédé après l'inscription. Les prédicteurs indépendants de la PAC comprenaient une absence d'allaitement exclusif pendant six mois (aOR = 1,7; IC95%: 1,0 - 2,9), un poids insuffisant (aOR = 2,1; IC95%: 1,0 - 4,5), un combustible de cuisson non propre (aOR = 1,8; IC95%: 1,0 - 3,3) et un faible revenu (aOR = 2,9; IC95%: 1,6 - 5,4). Aucune association n'a été trouvée entre le statut de vaccination et la PAC.Outre l'absence d'allaitement maternel exclusif, les enfants issus de familles à faible statut économique étaient à risque de contracter la PAC. Les facteurs de risque étant complexes, les résultats de l’étude appellent à des efforts plus concertés et une collaboration accrue entre les secteurs de la santé, de l'agriculture et du développement afin de lutter contre la mortalité causée par la PAC.

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.

Acellular pertussis (aP) booster vaccines are central to pertussis immunization programs, although their effectiveness varies. The Bacille Calmette-Guérin (BCG) vaccine is a prototype inducer of trained immunity, which enhances immune responses to subsequent infections or vaccinations. While previous clinical studies have demonstrated that trained immunity can protect against heterologous infections, its effect on aP vaccines in humans is unknown. We conducted a clinical study in order to determine the immunological effects of trained immunity on pertussis vaccination. Healthy female volunteers were randomly assigned to either receive BCG followed by a booster dose of tetanus-diphteria-pertussis inactivated polio vaccine (Tdap-IPV) 3 months later (BCG-trained), BCG + Tdap-IPV concurrently, or Tdap-IPV followed by BCG 3 months later. Primary outcomes were pertussis-specific humoral, T- and B-cell responses and were quantified at baseline of Tdap-IPV vaccination and 2 weeks thereafter. As a secondary outcome in the BCG-trained cohort, ex vivo leukocyte responses were measured in response to unrelated stimuli before and after BCG vaccination. BCG vaccination 3 months prior to, but not concurrent with, Tdap-IPV improves pertussis-specific Th1-cell and humoral responses, and also increases total memory B cell responses. These responses were correlated with enhanced IL-6 and IL-1β production at the baseline of Tdap-IPV vaccination in the BCG-trained cohort. Our study demonstrates that prior BCG vaccination potentiates immune responses to pertussis vaccines and that biomarkers of trained immunity are the most reliable correlates of those responses.

The mRNA-based BNT162b2 protects against severe disease and mortality caused by SARS-CoV-2 through induction of specific antibody and T-cell responses. Much less is known about its broad effects on immune responses against other pathogens. In the present study, we investigated the specific adaptive immune responses induced by BNT162b2 vaccination against various SARS-CoV-2 variants, as well as its effects on the responsiveness of human immune cells upon stimulation with heterologous viral, bacterial, and fungal pathogens. BNT162b2 vaccination induced effective humoral and cellular immunity against SARS-CoV-2 that started to wane after six months. We also observed long-term transcriptional changes in immune cells after vaccination, as assessed by RNA sequencing. Additionally, vaccination with BNT162b2 modulated innate immune responses as measured by the production of inflammatory cytokines when stimulated with various microbial stimuli other than SARS-CoV-2, including higher IL-1/IL-6 release and decreased production of IFN-α. Altogether, these data expand our knowledge regarding the overall immunological effects of this new class of vaccines and underline the need of additional studies to elucidate their effects on both innate and adaptive immune responses.

Objectives Complete deficiency of alternative pathway (AP) complement factors, explained by homozygous mutations, is a well‐known risk factor for invasive bacterial infections; however, this is less obvious for heterozygous mutations. We describe two siblings with a heterozygous NM_001928.3(CFD):c.125C>A p.(Ser42*) mutation in the complement factor D (fD) gene having a history of recurrent bacterial infections. We determined the effect of heterozygous fD deficiency on AP complement activity. Methods We determined the effect of fD levels on complement activation as measured by AP activity, complement C3 binding to the bacterial surface of Neisseria meningitidis (Nm), Streptococcus pneumoniae (Sp) and non‐typeable Haemophilus influenzae (NTHi), and complement‐mediated killing of Nm and NTHi. In addition, we measured the effect of vaccination of complement C3 binding to the bacterial surface and killing of Nm. Results Reconstitution of fD‐deficient serum with fD increased AP activity in a dose‐ and time‐dependent way. Reconstitution of patient serum with fD to normal levels increased complement C3 binding to Sp, Nm and NTHi, as well as complement‐mediated killing of Nm and NTHi. Vaccination increased complement C3 binding and resulted in complete killing of Nm without fD reconstitution. Conclusion We conclude that low fD serum levels (, In this study, we show that low complement factor D levels (

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.