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Competing Interests: The authors declare that there are no competing interests.

Coronavirus Disease-19 (COVID-19) symptoms range from mild to severe illness; the cause for this differential response to infection remains unknown. Unravelling the immune mechanisms acting at different levels of the colonization process might be key to understand these differences. We carried out a multi-tissue (nasal, buccal and blood; n = 156) gene expression analysis of immune-related genes from patients affected by different COVID-19 severities, and healthy controls through the nCounter technology. Mild and asymptomatic cases showed a powerful innate antiviral response in nasal epithelium, characterized by activation of interferon (IFN) pathway and downstream cascades, successfully controlling the infection at local level. In contrast, weak macrophage/monocyte driven innate antiviral response and lack of IFN signalling activity were present in severe cases. Consequently, oral mucosa from severe patients showed signals of viral activity, cell arresting and viral dissemination to the lower respiratory tract, which ultimately could explain the exacerbated innate immune response and impaired adaptative immune responses observed at systemic level. Results from saliva transcriptome suggest that the buccal cavity might play a key role in SARS-CoV-2 infection and dissemination in patients with worse prognosis. Co-expression network analysis adds further support to these findings, by detecting modules specifically correlated with severity involved in the abovementioned biological routes; this analysis also provides new candidate genes that might be tested as biomarkers in future studies. We also found tissue specific severity-related signatures mainly represented by genes involved in the innate immune system and cytokine/chemokine signalling. Local immune response could be key to determine the course of the systemic response and thus COVID-19 severity. Our findings provide a framework to investigate severity host gene biomarkers and pathways that might be relevant to diagnosis, prognosis, and therapy., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Respiratory syncytial virus (RSV) infection has been associated with the subsequent development of recurrent wheezing and asthma, although the mechanisms involved are still unknown. We investigate the role of epigenetics in the respiratory morbidity after infection by comparing methylation patterns from children who develop recurrent wheezing (RW-RSV), subsequent asthma (AS-RVS), and those experiencing complete recovery (CR-RSV)., Methods: Prospective, observational study of infants aged < 2 years with RSV respiratory infection admitted to hospital and followed-up after discharge for at least three years. According to their clinical course, patients were categorized into subgroups: RW-RSV ( n = 36), AS-RSV ( n = 9), and CR-RSV ( n = 32). The DNA genome-wide methylation pattern was analyzed in whole blood samples, collected during the acute phase of the infection, using the Illumina Infinium Methylation EPIC BeadChip (850K CpG sites). Differences in methylation were determined through a linear regression model adjusted for age, gender and cell composition., Results: Patients who developed respiratory sequelae showed a statistically significant higher proportion of NK and CD8T cells (inferred through a deconvolution approach) than those with complete recovery. We identified 5,097 significant differentially methylated positions (DMPs) when comparing RW-RSV and AS-RVS together against CR-RSV. Methylation profiles affect several genes involved in airway inflammation processes. The most significant DMPs were found to be hypomethylated in cases and therefore generally leading to overexpression of affected genes. The lead CpG position (cg24509398) falls at the gene body of EYA3 ( P -value = 2.77×10 -10 ), a tyrosine phosphatase connected with pulmonary vascular remodeling, a key process in the asthma pathology. Logistic regression analysis resulted in a diagnostic epigenetic signature of 3-DMPs (involving genes ZNF2698 , LOC102723354 and RPL15 / NKIRAS1 ) that allows to efficiently differentiate sequelae cases from CR-RSV patients (AUC = 1.00). Enrichment pathway analysis reveals the role of the cell cycle checkpoint (FDR P -value = 4.71×10 -2 ), DNA damage (FD P -value = 2.53×10 -2 ), and DNA integrity checkpoint (FDR P -value = 2.56×10 -2 ) in differentiating sequelae from CR-RSV patients., Conclusions: Epigenetic mechanisms might play a fundamental role in the long-term sequelae after RSV infection, contributing to explain the different phenotypes observed., Competing Interests: IR-C has received honoraria from GSK, Pfizer, Sanofi Pasteur and MSD for taking part in advisory boards and expert meetings and for acting as a speaker in congresses outside the scope of the submitted work. IR-C has also acted as subinvestigator in randomized controlled trials of Ablynx, Abbot, Seqirus, Sanofi Pasteur MSD, Sanofi Pasteur, Cubist, Wyeth, Merck, Pfizer, Roche, Regeneron, Jansen, Medimmune, Novavax, Novartis and GSK. FM-T has received honoraria from GSK group of companies, Pfizer Inc, Sanofi Pasteur, MSD, Seqirus, Biofabri and Janssen for taking part in advisory boards and expert meetings and for acting as a speaker in congresses outside the scope of the submitted work. FM-T has also acted as principal investigator in randomized controlled trials of the above-mentioned companies as well as Ablynx, Gilead, Regeneron, Roche, Abbott, Novavax, and MedImmune, with honoraria paid to his institution. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Pischedda, Rivero-Calle, Gómez-Carballa, Cebey-López, Barral-Arca, Gómez-Rial, Pardo-Seco, Curras-Tuala, Viz-Lasheras, Bello, Crujeiras, Diaz-Lagares, González-López, Martinón-Torres, Salas and GENDRES consortium.)

Background: Pediatric osteoarticular infections (POAIs) are serious diseases requiring early diagnosis and treatment., Methods: In this prospective multicenter cohort study, children with POAIs were selected from the European Union Childhood Life-threatening Infectious Diseases Study (EUCLIDS) database to analyze their demographic, clinical, and microbiological data., Results: A cohort of 380 patients with POAIs, 203 with osteomyelitis (OM), 158 with septic arthritis (SA), and 19 with both OM and SA, was analyzed. Thirty-five patients were admitted to the Pediatric Intensive Care Unit; out of these, six suffered from shock, one needed an amputation of the right foot and of four left toes, and two had skin transplantation. According to the Pediatric Overall Performance Score, 36 (10.5%) showed a mild overall disability, 3 (0.8%) a moderate, and 1 (0.2%) a severe overall disability at discharge. A causative organism was detected in 65% (247/380) of patients. Staphylococcus aureus ( S. aureus ) was identified in 57.1% (141/247) of microbiological confirmed cases, including 1 (0.7%) methicillin-resistant S. aureus (MRSA) and 6 (4.2%) Panton-Valentine leukocidin (PVL)-producing S. aureus , followed by Group A Streptococcus (18.2%) and Kingella kingae (8.9%). K. kingae and PVL production in S. aureus were less frequently reported than expected from the literature., Conclusion: POAIs are associated with a substantial morbidity in European children, with S. aureus being the major detected pathogen. In one-third of patients, no causative organism is identified. Our observations show an urgent need for the development of a vaccine against S. aureus and for the development of new microbiologic diagnostic guidelines for POAIs in European pediatric hospitals., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Trobisch, Schweintzger, Kohlfürst, Sagmeister, Sperl, Grisold, Feierl, Herberg, Carrol, Paulus, Emonts, van der Flier, de Groot, Cebey-López, Rivero-Calle, Boeddha, Agapow, Secka, Anderson, Behrends, Wintergerst, Reiter, Martinon-Torres, Levin, Zenz and the EUCLIDS consortium.)

Progressive osseous heteroplasia (POH; OMIM 166350) is a rare autosomal-dominant genetic disorder in which extra-skeletal bone forms within skin and muscle tissue. POH is one of the clinical manifestations of an inactivating mutation in the GNAS gene. GNAS gene alterations are difficult matter to address, as GNAS alleles show genetic imprinting and produce several transcript products, and the same mutation may lead to strikingly different phenotypes. Also, most of the publications concerning POH patients are either clinical depictions of a case (or a case series), descriptions of their genetic background, or a tentative correlation of both clinical and molecular findings. Treatment for POH is rarely addressed, and POH still lacks therapeutic options. We describe a unique case of POH in two monochorionic twins, who presented an almost asymptomatic vs. the severe clinical course, despite sharing the same mutation and genetic background. We also report the results of the therapeutic interventions currently available for heterotopic ossification in the patient with the severe course. This article not only critically supports the assumption that the POH course is strongly influenced by factors beyond genetic background but also remarks the lack of options for patients suffering an orphan disease, even after testing drugs with promising in vitro results., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Justicia-Grande, Gómez-Ríal, Rivero-Calle, Pischedda, Curras-Tuala, Gómez-Carballa, Cebey-López, Pardo-Seco, Méndez-Gallart, Fernández-Seara, Salas and Martinón-Torres.)

The fight against the spread of antibiotic resistance is one of the most important challenges facing health systems worldwide. Given the limitations of current diagnostic methods, the development of fast and accurate tests for the diagnosis of viral and bacterial infections would improve patient management and treatment, as well as contribute to reducing antibiotic misuse in clinical settings. In this scenario, analysis of host transcriptomics constitutes a promising target to develop new diagnostic tests based on the host-specific response to infections. We carried out a multi-cohort meta-analysis of blood transcriptomic data available in public databases, including 11 different studies and 1209 samples from virus- ( n = 695) and bacteria- ( n = 514) infected patients. We applied a Parallel Regularized Regression Model Search (PReMS) on a set of previously reported genes that distinguished viral from bacterial infection to find a minimum gene expression bio-signature. This strategy allowed us to detect three genes, namely BAFT , ISG15 and DNMT1 , that clearly differentiate groups of infection with high accuracy (training set: area under the curve (AUC) 0.86 (sensitivity: 0.81; specificity: 0.87); testing set: AUC 0.87 (sensitivity: 0.82; specificity: 0.86)). BAFT and ISG15 are involved in processes related to immune response, while DNMT1 is related to the preservation of methylation patterns, and its expression is modulated by pathogen infections. We successfully tested this three-transcript signature in the 11 independent studies, demonstrating its high performance under different scenarios. The main advantage of this three-gene signature is the low number of genes needed to differentiate both groups of patient categories.

Background: Rotavirus (RV) is an enteric pathogen that has devastating impact on childhood morbidity and mortality worldwide. The immunologic mechanism underlying the protection achieved after RV vaccination is not yet fully understood., Methods: We compared the transcriptome of children affected by community-acquired RV infection and children immunized with a live attenuated RV vaccine (RotaTeq ® )., Results: RV vaccination mimics the wild type infection causing similar changes in children's transcriptome, including transcripts associated with cell cycle, diarrhea, nausea, vomiting, intussusception, and abnormal morphology of midgut. A machine learning approach allowed to detect a combination of nine-transcripts that differentiates vaccinated from convalescent-naturally infected children (AUC: 90%; 95%CI: 70-100) and distinguishes between acute-infected and healthy control children (in both cases, AUC: 100%; 95%CI: 100-100). We identified a miRNA hsa-mir-149 that seems to play a role in the host defense against viral pathogens and may have an antiviral role., Discussion: Our findings might shed further light in the understanding of RV infection, its functional link to intussusception causes, as well as guide development of antiviral treatments and safer and more effective vaccines. The nine-transcript signature may constitute a marker of vaccine protection and helps to differentiate vaccinated from naturally infected or susceptible children., Competing Interests: FM-T has received honoraria from GSK, Pfizer, Sanofi Pasteur, MSD, Seqirus, and Janssen for taking part in advisory boards and expert meetings, and for acting as speaker in congresses outside the scope of the submitted work. JG-R has received honoraria from GSK, Pfizer, and MSD for taking part in advisory boards and expert meetings, and for acting as speaker in congresses outside the scope of the submitted work. FM-T has also acted as principal investigator in RCTs of the above-mentioned companies as well as Ablynx, Regeneron, Roche, Abbott, Novavax, and MedImmune, with honoraria paid to his institution. The remaining authors declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Gómez-Carballa, Barral-Arca, Cebey-López, Currás-Tuala, Pischedda, Gómez-Rial, Habgood-Coote, Herberg, Kaforou, Martinón-Torres and Salas.)

Background: Emerging evidence indicates a potential role for monocytes in COVID-19 immunopathology. We investigated two soluble markers of monocyte activation, sCD14 and sCD163, in COVID-19 patients, with the aim of characterizing their potential role in monocyte-macrophage disease immunopathology. To the best of our knowledge, this is the first study of its kind., Methods: Fifty-nine SARS-Cov-2 positive hospitalized patients, classified according to ICU or non-ICU admission requirement, were prospectively recruited and analyzed by ELISA for levels of sCD14 and sCD163, along with other laboratory parameters, and compared to a healthy control group., Results: sCD14 and sCD163 levels were significantly higher among COVID-19 patients, independently of ICU admission requirement, compared to the control group. We found a significant correlation between sCD14 levels and other inflammatory markers, particularly Interleukin-6, in the non-ICU patients group. sCD163 showed a moderate positive correlation with the time lapsed from admission to sampling, independently of severity group. Treatment with corticoids showed an interference with sCD14 levels, whereas hydroxychloroquine and tocilizumab did not., Conclusions: Monocyte-macrophage activation markers are increased and correlate with other inflammatory markers in SARS-Cov-2 infection, in association to hospital admission. These data suggest a preponderant role for monocyte-macrophage activation in the development of immunopathology of COVID-19 patients., (Copyright © 2020 Gómez-Rial, Currás-Tuala, Rivero-Calle, Gómez-Carballa, Cebey-López, Rodríguez-Tenreiro, Dacosta-Urbieta, Rivero-Velasco, Rodríguez-Núñez, Trastoy-Pena, Rodríguez-García, Salas and Martinón-Torres.)

There is a growing interest in unraveling gene expression mechanisms leading to viral host invasion and infection progression. Current findings reveal that long non-coding RNAs (lncRNAs) are implicated in the regulation of the immune system by influencing gene expression through a wide range of mechanisms. By mining whole-transcriptome shotgun sequencing (RNA-seq) data using machine learning approaches, we detected two lncRNAs (ENSG00000254680 and ENSG00000273149) that are downregulated in a wide range of viral infections and different cell types, including blood monocluclear cells, umbilical vein endothelial cells, and dermal fibroblasts. The efficiency of these two lncRNAs was positively validated in different viral phenotypic scenarios. These two lncRNAs showed a strong downregulation in virus-infected patients when compared to healthy control transcriptomes, indicating that these biomarkers are promising targets for infection diagnosis. To the best of our knowledge, this is the very first study using host lncRNAs biomarkers for the diagnosis of human viral infections.

Respiratory syncytial virus (RSV) is one of the major causes of acute lower respiratory tract infection worldwide. The absence of a commercial vaccine and the limited success of current therapeutic strategies against RSV make further research necessary. We used a multi-cohort analysis approach to investigate host transcriptomic biomarkers and shed further light on the molecular mechanism underlying RSV-host interactions. We meta-analyzed seven transcriptome microarray studies from the public Gene Expression Omnibus (GEO) repository containing a total of 922 samples, including RSV, healthy controls, coronaviruses, enteroviruses, influenzas, rhinoviruses, and coinfections, from both adult and pediatric patients. We identified > 1500 genes differentially expressed when comparing the transcriptomes of RSV-infected patients against healthy controls. Functional enrichment analysis showed several pathways significantly altered, including immunologic response mediated by RSV infection, pattern recognition receptors, cell cycle, and olfactory signaling. In addition, we identified a minimal 17-transcript host signature specific for RSV infection by comparing transcriptomic profiles against other respiratory viruses. These multi-genic signatures might help to investigate future drug targets against RSV infection.

Background: Diabetes is one of the underlying risk factors for developing community-acquired pneumonia (CAP). The high prevalence of diabetes among population and the rising incidence of this illness, converts it as an important disease to better control and manage, to prevent its secondary consequences as CAP. The objective of this research is to describe the characteristics of the patients with diabetes and the differences with the no diabetes who have had an episode of CAP in the context of the primary care field., Methods: A retrospective, observational study in adult patients (> 18 years-old) who suffer from CAP and attended at primary care in Spain between 2009 and 2013 was developed using the Computerized Database for Pharmacoepidemiological Studies in Primary Care (BIFAP). We carried out a descriptive analysis of the first episodes of CAP, in patients with or without diabetes as comorbidity. Other morbidity (CVA, Anaemia, Arthritis, Asthma, Heart disease, Dementia, Depression, Dysphagia, Multiple sclerosis, Epilepsy, COPD, Liver disease, Arthrosis, Parkinson's disease, Kidney disease, HIV) and life-style factors were also included in the study., Results: A total of 51,185 patients were included in the study as they suffer from the first episode of CAP. Of these, 8012 had diabetes as comorbidity. There were differences between sex and age in patients with diabetes. Patients without diabetes were younger, and had less comorbidities including those related to lifestyles such as smoking, alcoholism, social and dental problems than patients with diabetes., Conclusions: Patients who developed an episode of CAP with diabetes have more risk factors which could be reduced with an appropriate intervention, including vaccination to prevent successive CAP episodes and hospitalization. The burden of associated factors in these patients can produce an accumulation of risk. Health care professional should know this for treating and control these patients in order to avoid complications. Diabetes and those other risk factors associated could be reduced with an appropriate intervention, including vaccination to prevent the first and successive CAP episodes and the subsequent hospitalization in severe cases.

The diagnosis of bacterial infections in hospital settings is currently performed using bacterial culture from sterile site, but they are lengthy and limited. Transcriptomic biomarkers are becoming promising tools for diagnosis with potential applicability in clinical settings. We evaluated a RT-qPCR assay for a 2-transcript host expression signature (FAM89A and IFI44L genes) inferred from microarray data that allow to differentiate between viral and bacterial infection in febrile children. This assay was able to discriminate viral from bacterial infections (P-value = 1.04 × 10 -4 ; AUC = 92.2%; sensitivity = 90.9%; specificity = 85.7%) and showed very high reproducibility regardless of the reference gene(s) used to normalize the data. Unexpectedly, the monogenic IFI44L expression signature yielded better results than those obtained from the 2-transcript test (P-value = 3.59 × 10 -5 ; AUC = 94.1%; sensitivity = 90.9%; specificity = 92.8%). We validated this IFI44L signature in previously published microarray and whole-transcriptome data from patients affected by different types of viral and bacterial infections, confirming that this gene alone differentiates between both groups, thus saving time, effort, and costs. Herein, we demonstrate that host expression microarray data can be successfully translated into a fast, highly accurate and relatively inexpensive in vitro assay that could be implemented in the clinical routine.

Background: Sepsis and severe focal infections represent a substantial disease burden in children admitted to hospital. We aimed to understand the burden of disease and outcomes in children with life-threatening bacterial infections in Europe., Methods: The European Union Childhood Life-threatening Infectious Disease Study (EUCLIDS) was a prospective, multicentre, cohort study done in six countries in Europe. Patients aged 1 month to 18 years with sepsis (or suspected sepsis) or severe focal infections, admitted to 98 participating hospitals in the UK, Austria, Germany, Lithuania, Spain, and the Netherlands were prospectively recruited between July 1, 2012, and Dec 31, 2015. To assess disease burden and outcomes, we collected demographic and clinical data using a secured web-based platform and obtained microbiological data using locally available clinical diagnostic procedures., Findings: 2844 patients were recruited and included in the analysis. 1512 (53·2%) of 2841 patients were male and median age was 39·1 months (IQR 12·4-93·9). 1229 (43·2%) patients had sepsis and 1615 (56·8%) had severe focal infections. Patients diagnosed with sepsis had a median age of 27·6 months (IQR 9·0-80·2), whereas those diagnosed with severe focal infections had a median age of 46·5 months (15·8-100·4; p<0·0001). Of 2844 patients in the entire cohort, the main clinical syndromes were pneumonia (511 [18·0%] patients), CNS infection (469 [16·5%]), and skin and soft tissue infection (247 [8·7%]). The causal microorganism was identified in 1359 (47·8%) children, with the most prevalent ones being Neisseria meningitidis (in 259 [9·1%] patients), followed by Staphylococcus aureus (in 222 [7·8%]), Streptococcus pneumoniae (in 219 [7·7%]), and group A streptococcus (in 162 [5·7%]). 1070 (37·6%) patients required admission to a paediatric intensive care unit. Of 2469 patients with outcome data, 57 (2·2%) deaths occurred: seven were in patients with severe focal infections and 50 in those with sepsis., Interpretation: Mortality in children admitted to hospital for sepsis or severe focal infections is low in Europe. The disease burden is mainly in children younger than 5 years and is largely due to vaccine-preventable meningococcal and pneumococcal infections. Despite the availability and application of clinical procedures for microbiological diagnosis, the causative organism remained unidentified in approximately 50% of patients., Funding: European Union's Seventh Framework programme., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

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.

Pneumonia is the leading cause of death amongst infectious diseases. Streptococcus pneumoniae is responsible for about 25% of pneumonia cases worldwide, and it is a major cause of childhood mortality. We carried out a whole exome sequencing (WES) study in eight patients with complicated cases of pneumococcal pneumonia (empyema). An initial assessment of statistical association of WES variation with pneumonia was carried out using data from the 1000 Genomes Project (1000G) for the Iberian Peninsula (IBS) as reference controls. Pseudo-replication statistical analyses were carried out using different European control groups. Association tests pointed to single nucleotide polymorphism (SNP) rs201967957 (gene MEIS1 ; chromosome 2; p -value IBS = 3.71 × 10 -13 ) and rs576099063 (gene TSPAN15 ; chromosome 10; p -value IBS = 2.36 × 10 -8 ) as the best candidate variants associated to pneumococcal pneumonia. A burden gene test of pathogenicity signaled four genes, namely, OR9G9 , MUC6 , MUC3A and APOB , which carry significantly increased pathogenic variation when compared to controls. By analyzing various transcriptomic data repositories, we found strong supportive evidence for the role of MEIS1, TSPAN15 and APOBR (encoding the receptor of the APOB protein) in pneumonia in mouse and human models. Furthermore, the association of the olfactory receptor gene OR9G9 has recently been related to some viral infectious diseases, while the role of mucin genes ( MUC6 and MUC3A ), encoding mucin glycoproteins, are well-known factors related to chronic obstructive airway disease. WES emerges as a promising technique to disentangle the genetic basis of host genome susceptibility to infectious respiratory diseases.

Respiratory syncytial virus (RSV) is an important cause of serious lower respiratory tract disease in infants. Several studies have shown evidence pointing to the genome of the host as an important factor determining susceptibility to respiratory disease caused by RSV. We sequenced the complete exomes of 54 patients infected by RSV that needed hospitalization due to development of severe bronchiolitis. The Iberian sample (IBS) from The 1000 Genomes Project (1000G) was used as control group; all the association results were pseudo-replicated using other 1000G-European controls and Spanish controls. The study points to SNP rs199665292 in the olfactory receptor (OR) gene OR13C5 as the best candidate variant (P-value = 1.16 × 10 -12 ; OR = 5.56). Genetic variants at HLA genes (HLA-DQA1, HLA-DPB1), and in the mucin 4 gene (MUC4) also emerge as susceptibility candidates. By collapsing rare variants in genes and weighing by pathogenicity, we obtained confirmatory signals of association in the OR gene OR8U1/OR8U8, the taste receptor TAS2R19, and another mucin gene (MUC6). Overall, we identified new predisposition variants and genes related to RSV infection. Of special interest is the association of RSV to olfactory and taste receptors; this finding is in line with recent evidence pointing to their role in viral infectious diseases.

The mechanisms of rotavirus (RV) infection have been analyzed from different angles but the way in which RV modifies the transcriptome of the host is still unknown. Whole transcriptome shotgun sequencing of peripheral blood samples was used to reveal patterns of expression from the genome of RV-infected patients. RV provokes global changes in the transcriptome of infected cells, involving an over-expression of genes involved in cell cycle and chromatin condensation. While interferon IFI27 was hyper-activated, interferon type II was not suggesting that RV has developed mechanisms to evade the innate response by host cells after virus infection. Most interesting was the inhibition of genes of the glycophorins A and B (GYPA/B) family, which are the major sialoglycoproteins of the human erythrocyte membrane and receptor of several viruses for host invasion. RV infection induces a complex and global response in the host. The strong inhibition of glycophorins suggests a novel defense mechanism of the host to prevent viral infection, inhibiting the expression of receptors used by the virus for infection. The present results add further support to the systemic nature of RV infection., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Importance: Because clinical features do not reliably distinguish bacterial from viral infection, many children worldwide receive unnecessary antibiotic treatment, while bacterial infection is missed in others., Objective: To identify a blood RNA expression signature that distinguishes bacterial from viral infection in febrile children., Design, Setting, and Participants: Febrile children presenting to participating hospitals in the United Kingdom, Spain, the Netherlands, and the United States between 2009-2013 were prospectively recruited, comprising a discovery group and validation group. Each group was classified after microbiological investigation as having definite bacterial infection, definite viral infection, or indeterminate infection. RNA expression signatures distinguishing definite bacterial from viral infection were identified in the discovery group and diagnostic performance assessed in the validation group. Additional validation was undertaken in separate studies of children with meningococcal disease (n = 24) and inflammatory diseases (n = 48) and on published gene expression datasets., Exposures: A 2-transcript RNA expression signature distinguishing bacterial infection from viral infection was evaluated against clinical and microbiological diagnosis., Main Outcomes and Measures: Definite bacterial and viral infection was confirmed by culture or molecular detection of the pathogens. Performance of the RNA signature was evaluated in the definite bacterial and viral group and in the indeterminate infection group., Results: The discovery group of 240 children (median age, 19 months; 62% male) included 52 with definite bacterial infection, of whom 36 (69%) required intensive care, and 92 with definite viral infection, of whom 32 (35%) required intensive care. Ninety-six children had indeterminate infection. Analysis of RNA expression data identified a 38-transcript signature distinguishing bacterial from viral infection. A smaller (2-transcript) signature (FAM89A and IFI44L) was identified by removing highly correlated transcripts. When this 2-transcript signature was implemented as a disease risk score in the validation group (130 children, with 23 definite bacterial, 28 definite viral, and 79 indeterminate infections; median age, 17 months; 57% male), all 23 patients with microbiologically confirmed definite bacterial infection were classified as bacterial (sensitivity, 100% [95% CI, 100%-100%]) and 27 of 28 patients with definite viral infection were classified as viral (specificity, 96.4% [95% CI, 89.3%-100%]). When applied to additional validation datasets from patients with meningococcal and inflammatory diseases, bacterial infection was identified with a sensitivity of 91.7% (95% CI, 79.2%-100%) and 90.0% (95% CI, 70.0%-100%), respectively, and with specificity of 96.0% (95% CI, 88.0%-100%) and 95.8% (95% CI, 89.6%-100%). Of the children in the indeterminate groups, 46.3% (63/136) were classified as having bacterial infection, although 94.9% (129/136) received antibiotic treatment., Conclusions and Relevance: This study provides preliminary data regarding test accuracy of a 2-transcript host RNA signature discriminating bacterial from viral infection in febrile children. Further studies are needed in diverse groups of patients to assess accuracy and clinical utility of this test in different clinical settings.

Background and Aims: A properly validated scoring system allowing objective categorization of infants with acute respiratory infections (ARIs), avoiding the need for in-person assessment and that could also be used by non-health professionals is currently not available. We aimed to develop a new clinical assessment scale meeting these specifications., Methods: We designed a clinical scale (ReSVinet scale) based on seven parameters (feeding intolerance, medical intervention, respiratory difficulty, respiratory frequency, apnoea, general condition, fever) that were assigned different values (from 0 to 3) for a total of 20 points.170 children under two years of age with ARI were assessed independently by three pediatricians using this scale. Parents also evaluated their offspring with an adapted version of the scale in a subset of 61 cases. The scale was tested for internal consistency (Cronbach's alpha), Pearson correlation coefficient for the items in the scale, inter-observer reliability (kappa index) and floor-ceiling effect., Results: Internal consistency was good for all the observers, with the lowest Cronbach's alpha being 0.72. There was a strong correlation between the investigators (r-value ranged 0.76-0.83) and also between the results obtained by the parents and the investigators(r = 0.73). Light's kappa for the observations of the three investigators was 0.74. Weighted kappa in the group evaluated by the parents was 0.73. The final score was correlated with length of hospital stay, PICU admission and Wood-Downes Score., Conclusions: The ReSVinet scale may be useful and reliable in the evaluation of infants with ARI, particularly acute bronchiolitis, even with data obtained from medical records and when employed by parents. Although further studies are necessary, ReSVinet scale already complies with more score validation criteria than the vast majority of the alternatives currently available and used in the clinical practice.

Background: Multiple viruses are often detected in children with respiratory infection but the significance of co-infection in pathogenesis, severity and outcome is unclear., Objectives: To correlate the presence of viral co-infection with clinical phenotype in children admitted with acute respiratory infections (ARI)., Methods: We collected detailed clinical information on severity for children admitted with ARI as part of a Spanish prospective multicenter study (GENDRES network) between 2011-2013. A nested polymerase chain reaction (PCR) approach was used to detect respiratory viruses in respiratory secretions. Findings were compared to an independent cohort collected in the UK., Results: 204 children were recruited in the main cohort and 97 in the replication cohort. The number of detected viruses did not correlate with any markers of severity. However, bacterial superinfection was associated with increased severity (OR: 4.356; P-value = 0.005), PICU admission (OR: 3.342; P-value = 0.006), higher clinical score (1.988; P-value = 0.002) respiratory support requirement (OR: 7.484; P-value < 0.001) and longer hospital length of stay (OR: 1.468; P-value < 0.001). In addition, pneumococcal vaccination was found to be a protective factor in terms of degree of respiratory distress (OR: 2.917; P-value = 0.035), PICU admission (OR: 0.301; P-value = 0.011), lower clinical score (-1.499; P-value = 0.021) respiratory support requirement (OR: 0.324; P-value = 0.016) and oxygen necessity (OR: 0.328; P-value = 0.001). All these findings were replicated in the UK cohort., Conclusion: The presence of more than one virus in hospitalized children with ARI is very frequent but it does not seem to have a major clinical impact in terms of severity. However bacterial superinfection increases the severity of the disease course. On the contrary, pneumococcal vaccination plays a protective role.

Background: Vitamin D is known to have modulatory actions in the immune system. Its influence on the severity of lower tract acute respiratory infections (LT-ARIs) is unclear., Objectives: The aim of the present study was to evaluate the role of vitamin D on LT-ARI in paediatric patients., Methods: Children admitted to hospital with LT-ARI were prospectively recruited through the GENDRES network (March 2009-May 2013). The 25-hydroxyvitamin D (25-OHD) levels were measured by immunoassay. The severity of the illness was evaluated according to clinical scales, length of hospital stay, ventilatory requirements, and pediatric intensive care unit admission., Results: A total of 347 patients with a median (interquartile range) age of 8.4 (2.6-21.1) months were included. The mean (SD) 25-OHD levels in our series were 27.1 (11.3) ng/mL. In this study, a cutoff value of ≥30 ng/mL was considered optimal vitamin status. Patients with 25-OHD levels <20 ng/mL were at a higher risk of showing severe signs of respiratory difficulties (OR 5.065, 95% confidence interval 1.998-12.842; P = 0.001) than patients with normal values, and had a 117% higher risk of oxygen necessity and 217% higher risk of ventilatory requirement than those patients with normal values. An inverse correlation was found between 25-OHD levels and the severity in the evaluated scales. 25-OHD levels did not influence PICU admission rate or length of hospital stay., Conclusions: 25-OHD levels of children admitted because of a LT-ARI are <30 ng/mL. Lower levels of 25-OHD were found to be correlated with severity of the disease. The possible role of abnormal 25-OHD levels as a facilitator or consequence of the infection needs further evaluation.

Background: The risk of bacteremia is considered low in children with acute bronchiolitis. However the rate of occult bacteremia in infants with RSV infection is not well established. The aim was to determine the actual rate and predictive factors of bacteremia in children admitted to hospital due to confirmed RSV acute respiratory illness (ARI), using both conventional culture and molecular techniques., Methods: A prospective multicenter study (GENDRES-network) was conducted between 2011-2013 in children under the age of two admitted to hospital because of an ARI. Among those RSV-positive, bacterial presence in blood was assessed using PCR for Meningococcus, Streptococcus pneumoniae, Haemophilus influenzae, Streptococcus pyogenes, Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus, in addition to conventional cultures., Results: 66 children with positive RSV respiratory illness were included. In 10.6% patients, bacterial presence was detected: H. influenzae (n = 4) and S. pneumoniae (n = 2). In those patients with bacteremia, there was a previous suspicion of bacterial superinfection and had received empirical antibiotic treatment 6 out of 7 (85.7%) patients. There were significant differences in terms of severity between children with positive bacterial PCR and those with negative results: PICU admission (100% vs. 50%, P-value = 0.015); respiratory support necessity (100% vs. 18.6%, P-value < 0.001); Wood-Downes score (mean = 8.7 vs. 4.8 points, P-value < 0.001); GENVIP scale (mean = 17 vs. 10.1, P-value < 0.001); and length of hospitalization (mean = 12.1 vs. 7.5 days, P-value = 0.007)., Conclusion: Bacteremia is not frequent in infants hospitalized with RSV respiratory infection, however, it should be considered in the most severe cases.

Background: Molecular techniques can often reveal a broader range of pathogens in respiratory infections. We aim to investigate the prevalence and age pattern of viral co-infection in children hospitalized with lower tract acute respiratory infection (LT-ARI), using molecular techniques., Methods: A nested polymerase chain reaction approach was used to detect Influenza (A, B), metapneumovirus, respiratory syncytial virus (RSV), parainfluenza (1-4), rhinovirus, adenovirus (A-F), bocavirus and coronaviruses (NL63, 229E, OC43) in respiratory samples of children with acute respiratory infection prospectively admitted to any of the GENDRES network hospitals between 2011-2013. The results were corroborated in an independent cohort collected in the UK., Results: A total of 204 and 97 nasopharyngeal samples were collected in the GENDRES and UK cohorts, respectively. In both cohorts, RSV was the most frequent pathogen (52.9% and 36.1% of the cohorts, respectively). Co-infection with multiple viruses was found in 92 samples (45.1%) and 29 samples (29.9%), respectively; this was most frequent in the 12-24 months age group. The most frequently observed co-infection patterns were RSV-Rhinovirus (23 patients, 11.3%, GENDRES cohort) and RSV-bocavirus / bocavirus-influenza (5 patients, 5.2%, UK cohort)., Conclusion: The presence of more than one virus in pediatric patients admitted to hospital with LT-ARI is very frequent and seems to peak at 12-24 months of age. The clinical significance of these findings is unclear but should warrant further analysis.

Background: Rotavirus vaccine (RV) might reduce the risk of hospitalization due to childhood seizures (CS). We aimed to identify and assess variations in the incidence of hospitalizations for CS among children <5 years of age before and after RV introduction., Methods: Annual hospitalization rates for any kind of CS, before and after RV introduction in 2007, were calculated using the official surveillance system for hospitalization data., Results: Our study cohort totaled 6149 children <5 years of age admitted to the hospital between 2003 and 2013 with any kind of CS (780.3* + 779.0* + 333.2* + 345* ICD-9-CM code). The annual hospitalization rates for any kind of CS in children <5 years of age were correlated with RV coverage (r = -0.673; P = 0.033) and rotavirus acute gastroenteritis admission rates (ρ = 0.506; P = 0.001), with decrease rates ranging from 16.2% (95% confidence interval: 8.3-23.5%) in 2007 to 34.0% (27.3-40.1%) in 2010, as compared with the median rate of the pre-vaccination period (2003 to 2006). Similarly, for convulsions (780.3*ICD-9-CM code), the decrease seen in children <5 years of age was significantly correlated with the increase in RV coverage (r = -0.747; P = 0.013) and rotavirus acute gastroenteritis admission rates (ρ = 0.543; P < 0.001), with decrease rates ranging from 18.7% (9.6-26.8%) in 2007 to 42.5% (35.3-48.9%) in 2012. Significant results were also obtained for infants <12 months and infants 1-2 years of age. In the remaining age groups or diagnostic categories analyzed, changes were either not significant or not related to vaccination changes or rotavirus acute gastroenteritis admission rates., Conclusions: Our results show that rotavirus vaccination may have a significant impact in the decrease in seizure-related hospitalizations in childhood. This additional benefit of rotavirus vaccination seems more marked in the youngest infants.