Immunogenicity, reactogenicity, and IgE-mediated immune responses of a mixed whole-cell and acellular pertussis vaccine schedule in Australian infants: A randomised, double-blind, noninferiority trial.

BACKGROUND: In many countries, infant vaccination with acellular pertussis (aP) vaccines has replaced use of more reactogenic whole-cell pertussis (wP) vaccines. Based on immunological and epidemiological evidence, we hypothesised that substituting the first aP dose in the routine vaccination schedule with wP vaccine might protect against IgE-mediated food allergy. We aimed to compare reactogenicity, immunogenicity, and IgE-mediated responses of a mixed wP/aP primary schedule versus the standard aP-only schedule. METHODS AND FINDINGS: OPTIMUM is a Bayesian, 2-stage, double-blind, randomised trial. In stage one, infants were assigned (1:1) to either a first dose of a pentavalent wP combination vaccine (DTwP-Hib-HepB, Pentabio PT Bio Farma, Indonesia) or a hexavalent aP vaccine (DTaP-Hib-HepB-IPV, Infanrix hexa, GlaxoSmithKline, Australia) at approximately 6 weeks old. Subsequently, all infants received the hexavalent aP vaccine at 4 and 6 months old as well as an aP vaccine at 18 months old (DTaP-IPV, Infanrix-IPV, GlaxoSmithKline, Australia). Stage two is ongoing and follows the above randomisation strategy and vaccination schedule. Ahead of ascertainment of the primary clinical outcome of allergist-confirmed IgE-mediated food allergy by 12 months old, here we present the results of secondary immunogenicity, reactogenicity, tetanus toxoid IgE-mediated immune responses, and parental acceptability endpoints. Serum IgG responses to diphtheria, tetanus, and pertussis antigens were measured using a multiplex fluorescent bead-based immunoassay; total and specific IgE were measured in plasma by means of the ImmunoCAP assay (Thermo Fisher Scientific). The immunogenicity of the mixed schedule was defined as being noninferior to that of the aP-only schedule using a noninferiority margin of 2/3 on the ratio of the geometric mean concentrations (GMR) of pertussis toxin (PT)-IgG 1 month after the 6-month aP. Solicited adverse reactions were summarised by study arm and included all children who received the first dose of either wP or aP. Parental acceptance was assessed using a 5-point Likert scale. The primary analyses were based on intention-to-treat (ITT); secondary per-protocol (PP) analyses were also performed. The trial is registered with ANZCTR (ACTRN12617000065392p). Between March 7, 2018 and January 13, 2020, 150 infants were randomised (75 per arm). PT-IgG responses of the mixed schedule were noninferior to the aP-only schedule at approximately 1 month after the 6-month aP dose [GMR = 0·98, 95% credible interval (0·77 to 1·26); probability (GMR > 2/3) > 0·99; ITT analysis]. At 7 months old, the posterior median probability of quantitation for tetanus toxoid IgE was 0·22 (95% credible interval 0·12 to 0·34) in both the mixed schedule group and in the aP-only group. Despite exclusions, the results were consistent in the PP analysis. At 6 weeks old, irritability was the most common systemic solicited reaction reported in wP (65 [88%] of 74) versus aP (59 [82%] of 72) vaccinees. At the same age, severe systemic reactions were reported among 14 (19%) of 74 infants after wP and 8 (11%) of 72 infants after aP. There were 7 SAEs among 5 participants within the first 6 months of follow-up; on blinded assessment, none were deemed to be related to the study vaccines. Parental acceptance of mixed and aP-only schedules was high (71 [97%] of 73 versus 69 [96%] of 72 would agree to have the same schedule again). CONCLUSIONS: Compared to the aP-only schedule, the mixed schedule evoked noninferior PT-IgG responses, was associated with more severe reactions, but was well accepted by parents. Tetanus toxoid IgE responses did not differ across the study groups. TRIAL REGISTRATION: Trial registered at the Australian and New Zealand Clinical 207 Trial Registry (ACTRN12617000065392p).

Mapping Lung Hematopoietic Progenitors: Developmental Kinetics and Response to Influenza A Infection.

The bone marrow is a specialized niche responsible for the maintenance of hematopoietic stem and progenitor cells during homeostasis and inflammation. Recent studies, however, have extended this essential role to the extramedullary and extravascular lung microenvironment. Here, we provide further evidence for a reservoir of hematopoietic stem and progenitor cells within the lung from Embryonic Day 18.5 until adulthood. These lung progenitors display distinct microenvironment-specific developmental kinetics compared with their bone marrow counterparts, exemplified by a rapid shift from a common myeloid to a megakaryocyte-erythrocyte progenitor-dominated niche with increasing age. In adult mice, influenza A viral infection results in a transient reduction in multipotent progenitors within the lungs, with a parallel increase in downstream granulocyte-monocyte progenitors and dendritic cell populations associated with acute viral infections. Our findings suggest that lung hematopoietic progenitors play a role in reestablishing immunological homeostasis in the respiratory mucosa, which may have significant clinical implications for maintaining pulmonary health after inflammatory perturbation.

Impaired interferon response in plasmacytoid dendritic cells from children with persistent wheeze.

BACKGROUND: Impaired interferon response and allergic sensitization may contribute to virus-induced wheeze and asthma development in young children. Plasmacytoid dendritic cells (pDCs) play a key role in antiviral immunity as critical producers of type I interferons. pDCs also express the high-affinity IgE receptor through which type I interferon production may be negatively regulated. Whether antiviral function of pDCs is associated with recurrent episodes of wheeze in young children is not well understood. OBJECTIVE: We sought to evaluate the phenotype and function of circulating pDCs in children with a longitudinally defined wheezing phenotype. METHODS: We performed multiparameter flow cytometry on PBMCs from 38 children presenting to the emergency department with an acute episode of respiratory wheeze and 19 controls. RNA sequencing on isolated pDCs from the same individuals was also performed. For each subject, their longitudinal exacerbation phenotype was determined using the Western Australia public hospital database. RESULTS: We observed a significant depletion of circulating pDCs in young children with a persistent phenotype of wheeze. The same individuals also displayed upregulation of the FcεRI on their pDCs. Based on transcriptomic analysis, pDCs from these individuals did not mount a robust systemic antiviral response as observed in children who displayed a nonrecurrent wheezing phenotype. CONCLUSIONS: Our data suggest that circulating pDC phenotype and function are altered in young children with a persistent longitudinal exacerbation phenotype. Expression of high-affinity IgE receptor is increased and their function as major interferon producers is impaired during acute exacerbations of wheeze.

Single cell transcriptomics reveals cell type specific features of developmentally regulated responses to lipopolysaccharide between birth and 5 years.

Background: Human perinatal life is characterized by a period of extraordinary change during which newborns encounter abundant environmental stimuli and exposure to potential pathogens. To meet such challenges, the neonatal immune system is equipped with unique functional characteristics that adapt to changing conditions as development progresses across the early years of life, but the molecular characteristics of such adaptations remain poorly understood. The application of single cell genomics to birth cohorts provides an opportunity to investigate changes in gene expression programs elicited downstream of innate immune activation across early life at unprecedented resolution. Methods: In this study, we performed single cell RNA-sequencing of mononuclear cells collected from matched birth cord blood and 5-year peripheral blood samples following stimulation (18hrs) with two well-characterized innate stimuli; lipopolysaccharide (LPS) and Polyinosinic:polycytidylic acid (Poly(I:C)). Results: We found that the transcriptional response to LPS was constrained at birth and predominantly partitioned into classical proinflammatory gene upregulation primarily by monocytes and Interferon (IFN)-signaling gene upregulation by lymphocytes. Moreover, these responses featured substantial cell-to-cell communication which appeared markedly strengthened between birth and 5 years. In contrast, stimulation with Poly(I:C) induced a robust IFN-signalling response across all cell types identified at birth and 5 years. Analysis of gene regulatory networks revealed IRF1 and STAT1 were key drivers of the LPS-induced IFN-signaling response in lymphocytes with a potential developmental role for IRF7 regulation. Conclusion: Additionally, we observed distinct activation trajectory endpoints for monocytes derived from LPS-treated cord and 5-year blood, which was not apparent among Poly(I:C)-induced monocytes. Taken together, our findings provide new insight into the gene regulatory landscape of immune cell function between birth and 5 years and point to regulatory mechanisms relevant to future investigation of infection susceptibility in early life.

LPS binding protein and activation signatures are upregulated during asthma exacerbations in children.

Asthma exacerbations in children are associated with respiratory viral infection and atopy, resulting in systemic immune activation and infiltration of immune cells into the airways. The gene networks driving the immune activation and subsequent migration of immune cells into the airways remains incompletely understood. Cellular and molecular profiling of PBMC was employed on paired samples obtained from atopic asthmatic children (n = 19) during acute virus-associated exacerbations and later during convalescence. Systems level analyses were employed to identify coexpression networks and infer the drivers of these networks, and validation was subsequently obtained via independent samples from asthmatic children. During exacerbations, PBMC exhibited significant changes in immune cell abundance and upregulation of complex interlinked networks of coexpressed genes. These were associated with priming of innate immunity, inflammatory and remodelling functions. We identified activation signatures downstream of bacterial LPS, glucocorticoids and TGFB1. We also confirmed that LPS binding protein was upregulated at the protein-level in plasma. Multiple gene networks known to be involved positively or negatively in asthma pathogenesis, are upregulated in circulating PBMC during acute exacerbations, supporting the hypothesis that systemic pre-programming of potentially pathogenic as well as protective functions of circulating immune cells preceeds migration into the airways. Enhanced sensitivity to LPS is likely to modulate the severity of acute asthma exacerbations through exposure to environmental LPS.

Maternal diet modulates the infant microbiome and intestinal Flt3L necessary for dendritic cell development and immunity to respiratory infection.

Poor maternal diet during pregnancy is a risk factor for severe lower respiratory infections (sLRIs) in the offspring, but the underlying mechanisms remain elusive. Here, we demonstrate that in mice a maternal low-fiber diet (LFD) led to enhanced LRI severity in infants because of delayed plasmacytoid dendritic cell (pDC) recruitment and perturbation of regulatory T cell expansion in the lungs. LFD altered the composition of the maternal milk microbiome and assembling infant gut microbiome. These microbial changes reduced the secretion of the DC growth factor Flt3L by neonatal intestinal epithelial cells and impaired downstream pDC hematopoiesis. Therapy with a propionate-producing bacteria isolated from the milk of high-fiber diet-fed mothers, or supplementation with propionate, conferred protection against sLRI by restoring gut Flt3L expression and pDC hematopoiesis. Our findings identify a microbiome-dependent Flt3L axis in the gut that promotes pDC hematopoiesis in early life and confers disease resistance against sLRIs.

Asthma and allergies in a cohort of adolescents conceived with ART.

RESEARCH QUESTION: Are asthma and allergies more common in adolescents conceived with assisted reproductive technologies (ART) compared with adolescents conceived without? DESIGN: The Growing Up Healthy Study (GUHS) is a prospective cohort study including ART-conceived offspring born between 1991 and 2001 in Perth, Australia. Their long-term health outcomes, including asthma and allergy parameters, were compared with those of their counterparts conceived without ART from the Raine Study Generation 2 (Gen2), born in 1989-1991. At age 14, 152 GUHS and 1845 Gen2 participants completed the following assessments: the International Studies of Asthma and Allergies in Childhood (ISAAC) questionnaire, spirometry, methacholine challenge testing and skin prick testing (SPT). RESULTS: No differences were detected in the prevalence of current asthma (7.7% versus 10.8%, adjusted odds ratio [aOR] 0.82 (95% CI 0.44-1.52), P = 0.530). Spirometry-measured lung volumes were larger in the ART adolescents. Bronchial hyperresponsiveness was less prevalent in the ART cohort (8.8 versus 18.6%, P = 0.006). Current allergic rhinoconjunctivitis (ARC) rates were significantly higher in the ART cohort (32.4% versus 25.2%, aOR 1.52 [95% CI 1.03-2.26], P = 0.036), with no cohort differences in atopic dermatitis. Food allergies were more prevalent in the ART cohort (20.7 versus 10.9%, aOR 1.89 [95% CI 1.17-3.06], P = 0.010) with more adolescents having a positive SPT (68.0% versus 45.4%, aOR 3.03 [95% 1.99-4.63], P < 0.001). CONCLUSIONS: This study reports no differences in asthma prevalence, slightly altered lung function, an increase in ARC, food allergies and positive SPT in the ART-conceived adolescents. These findings are important to families and healthcare providers and may open up possibilities for targeted screening and treatment. Further studies are required to confirm these findings.

Lipopolysaccharide-induced interferon response networks at birth are predictive of severe viral lower respiratory infections in the first year of life.

Appropriate innate immune function is essential to limit pathogenesis and severity of severe lower respiratory infections (sLRI) during infancy, a leading cause of hospitalization and risk factor for subsequent asthma in this age group. Employing a systems biology approach to analysis of multi-omic profiles generated from a high-risk cohort (n=50), we found that the intensity of activation of an LPS-induced interferon gene network at birth was predictive of sLRI risk in infancy (AUC=0.724). Connectivity patterns within this network were stronger among susceptible individuals, and a systems biology approach identified IRF1 as a putative master regulator of this response. These findings were specific to the LPS-induced interferon response and were not observed following activation of viral nucleic acid sensing pathways. Comparison of responses at birth versus age 5 demonstrated that LPS-induced interferon responses but not responses triggered by viral nucleic acid sensing pathways may be subject to strong developmental regulation. These data suggest that the risk of sLRI in early life is in part already determined at birth, and additionally that the developmental status of LPS-induced interferon responses may be a key determinant of susceptibility. Our findings provide a rationale for the identification of at-risk infants for early intervention aimed at sLRI prevention and identifies targets which may be relevant for drug development.

Remission of peanut allergy is associated with rewiring of allergen-driven T helper 2-related gene networks.

BACKGROUND: The immunological changes underpinning acquisition of remission (also called sustained unresponsiveness) following food immunotherapy remain poorly defined. Limited access to effective therapies and biosamples from treatment responders has prevented progress. Probiotic peanut oral immunotherapy is highly effective at inducing remission, providing an opportunity to investigate immune changes. METHODS: Using a systems biology approach, we examined gene co-expression network patterns in peanut-specific CD4+ T cell responses before and after probiotic and peanut oral immunotherapy in subjects enrolled in the PPOIT-001 randomized trial: Responders who attained remission (n = 16), placebo-treated who remained allergic (n = 16). RESULTS: Acquisition of remission was associated with rewiring of gene network patterns, which was characterized by integration of T helper 2 and interferon signalling modules, markedly reduced T helper 2 gene connectivity, and shutdown in co-expression activity between T helper 2 effectors and cell cycle regulators. CONCLUSION: The immunological changes underlying remission following peanut oral immunotherapy are mediated by reprogramming of T helper 2-associated gene networks in the CD4+ T cell compartment. Findings provide insight into immune mechanisms driving the acquisition of remission following oral immunotherapy, paving the way for the development of improved approaches to induce remission/sustained unresponsiveness in patients with food allergy.

Protection against severe infant lower respiratory tract infections by immune training: Mechanistic studies.

BACKGROUND: Results from recent clinical studies suggest potential efficacy of immune training (IT)-based approaches for protection against severe lower respiratory tract infections in infants, but underlying mechanisms are unclear. OBJECTIVE: We used systems-level analyses to elucidate IT mechanisms in infants in a clinical trial setting. METHODS: Pre- and posttreatment peripheral blood mononuclear cells from a placebo-controlled trial in which winter treatment with the IT agent OM85 reduced infant respiratory infection frequency and/or duration were stimulated for 24 hours with the virus/bacteria mimics polyinosinic:polycytidylic acid/lipopolysaccharide. Transcriptomic profiling via RNA sequencing, pathway and upstream regulator analyses, and systems-level gene coexpression network analyses were used sequentially to elucidate and compare responses in treatment and placebo groups. RESULTS: In contrast to subtle changes in antivirus-associated polyinosinic:polycytidylic acid response profiles, the bacterial lipopolysaccharide-triggered gene coexpression network responses exhibited OM85 treatment-associated upregulation of IFN signaling. This was accompanied by network rewiring resulting in increased coordination of TLR4 expression with IFN pathway-associated genes (especially master regulator IRF7); segregation of TNF and IFN-γ (which potentially synergize to exaggerate inflammatory sequelae) into separate expression modules; and reduced size/complexity of the main proinflammatory network module (containing, eg, IL-1,IL-6, and CCL3). Finally, we observed a reduced capacity for lipopolysaccharide-induced inflammatory cytokine (eg, IL-6 and TNF) production in the OM85 group. CONCLUSION: These changes are consistent with treatment-induced enhancement of bacterial pathogen detection/clearance capabilities concomitant with enhanced capacity to regulate ensuing inflammatory response intensity and duration. We posit that IT agents exemplified by OM85 potentially protect against severe lower respiratory tract infections in infants principally by effects on innate immune responses targeting the bacterial components of the mixed respiratory viral/bacterial infections that are characteristic of this age group.

Rare variant analysis in eczema identifies exonic variants in DUSP1, NOTCH4 and SLC9A4.

Previous genome-wide association studies revealed multiple common variants involved in eczema but the role of rare variants remains to be elucidated. Here, we investigate the role of rare variants in eczema susceptibility. We meta-analyze 21 study populations including 20,016 eczema cases and 380,433 controls. Rare variants are imputed with high accuracy using large population-based reference panels. We identify rare exonic variants in DUSP1, NOTCH4, and SLC9A4 to be associated with eczema. In DUSP1 and NOTCH4 missense variants are predicted to impact conserved functional domains. In addition, five novel common variants at SATB1-AS1/KCNH8, TRIB1/LINC00861, ZBTB1, TBX21/OSBPL7, and CSF2RB are discovered. While genes prioritized based on rare variants are significantly up-regulated in the skin, common variants point to immune cell function. Over 20% of the single nucleotide variant-based heritability is attributable to rare and low-frequency variants. The identified rare/low-frequency variants located in functional protein domains point to promising targets for novel therapeutic approaches to eczema.

IRF7-Associated Immunophenotypes Have Dichotomous Responses to Virus/Allergen Coexposure and OM-85-Induced Reprogramming.

High risk for virus-induced asthma exacerbations in children is associated with an IRF7lo immunophenotype, but the underlying mechanisms are unclear. Here, we applied a Systems Biology approach to an animal model comprising rat strains manifesting high (BN) versus low susceptibility (PVG) to experimental asthma, induced by virus/allergen coexposure, to elucidate the mechanism(s)-of-action of the high-risk asthma immunophenotype. We also investigated potential risk mitigation via pretreatment with the immune training agent OM-85. Virus/allergen coexposure in low-risk PVG rats resulted in rapid and transient airways inflammation alongside IRF7 gene network formation. In contrast, responses in high-risk BN rats were characterized by severe airways eosinophilia and exaggerated proinflammatory responses that failed to resolve, and complete absence of IRF7 gene networks. OM-85 had more profound effects in high-risk BN rats, inducing immune-related gene expression changes in lung at baseline and reducing exaggerated airway inflammatory responses to virus/allergen coexposure. In low-risk PVG rats, OM-85 boosted IRF7 gene networks in the lung but did not alter baseline gene expression or cellular influx. Distinct IRF7-associated asthma risk immunophenotypes have dichotomous responses to virus/allergen coexposure and respond differentially to OM-85 pretreatment. Extrapolating to humans, our findings suggest that the beneficial effects OM-85 pretreatment may preferentially target those in high-risk subgroups.

Protection against neonatal respiratory viral infection via maternal treatment during pregnancy with the benign immune training agent OM-85.

OBJECTIVES: Incomplete maturation of immune regulatory functions at birth is antecedent to the heightened risk for severe respiratory infections during infancy. Our forerunner animal model studies demonstrated that maternal treatment with the microbial-derived immune training agent OM-85 during pregnancy promotes accelerated postnatal maturation of mechanisms that regulate inflammatory processes in the offspring airways. Here, we aimed to provide proof of concept for a novel solution to reduce the burden and potential long-term sequelae of severe early-life respiratory viral infection through maternal oral treatment during pregnancy with OM-85, already in widespread human clinical use. METHODS: In this study, we performed flow cytometry and targeted gene expression (RT-qPCR) analysis on lungs from neonatal offspring whose mothers received oral OM-85 treatment during pregnancy. We next determined whether neonatal offspring from OM-85 treated mothers demonstrate enhanced protection against lethal lower respiratory infection with mouse-adapted rhinovirus (vMC0), and associated lung immune changes. RESULTS: Offspring from mothers treated with OM-85 during pregnancy display accelerated postnatal seeding of lung myeloid populations demonstrating upregulation of function-associated markers. Offspring from OM-85 mothers additionally exhibit enhanced expression of TLR4/7 and the IL-1ß/NLRP3 inflammasome complex within the lung. These treatment effects were associated with enhanced capacity to clear an otherwise lethal respiratory viral infection during the neonatal period, with concomitant regulation of viral-induced IFN response intensity. CONCLUSION: These results demonstrate that maternal OM-85 treatment protects offspring against lethal neonatal respiratory viral infection by accelerating development of innate immune mechanisms crucial for maintenance of local immune homeostasis in the face of pathogen challenge.

The intersect of genetics, environment, and microbiota in asthma-perspectives and challenges.

In asthma, a significant portion of the interaction between genetics and environment occurs through microbiota. The proposed mechanisms behind this interaction are complex and at times contradictory. This review covers recent developments in our understanding of this interaction: the "microbial hypothesis" and the "farm effect"; the role of endotoxin and genetic variation in pattern recognition systems; the interaction with allergen exposure; the additional involvement of host gut and airway microbiota; the role of viral respiratory infections in interaction with the 17q21 and CDHR3 genetic loci; and the importance of in utero and early-life timing of exposures. We propose a unified framework for understanding how all these phenomena interact to drive asthma pathogenesis. Finally, we point out some future challenges for continued research in this field, in particular the need for multiomic integration, as well as the potential utility of asthma endotyping.

Developmental patterns in the nasopharyngeal microbiome during infancy are associated with asthma risk.

BACKGROUND: Studies indicate that the nasal microbiome may correlate strongly with the presence or future risk of childhood asthma. OBJECTIVES: In this study, we tested whether developmental trajectories of the nasopharyngeal microbiome in early life and the composition of the microbiome during illnesses were related to risk of childhood asthma. METHODS: Children participating in the Childhood Origins of Asthma study (N = 285) provided nasopharyngeal mucus samples in the first 2 years of life, during routine healthy study visits (at 2, 4, 6, 9, 12, 18, and 24 months of age), and during episodes of respiratory illnesses, all of which were analyzed for respiratory viruses and bacteria. We identified developmental trajectories of early-life microbiome composition, as well as predominant bacteria during respiratory illnesses, and we correlated these with presence of asthma at 6, 8, 11, 13, and 18 years of age. RESULTS: Of the 4 microbiome trajectories identified, a Staphylococcus-dominant microbiome in the first 6 months of life was associated with increased risk of recurrent wheezing by age 3 years and asthma that persisted throughout childhood. In addition, this trajectory was associated with the early onset of allergic sensitization. During wheezing illnesses, detection of rhinoviruses and predominance of Moraxella were associated with asthma that persisted throughout later childhood. CONCLUSION: In infancy, the developmental composition of the microbiome during healthy periods and the predominant microbes during acute wheezing illnesses are both associated with the subsequent risk of developing persistent childhood asthma.

OPTIMUM study protocol: an adaptive randomised controlled trial of a mixed whole-cell/acellular pertussis vaccine schedule.

INTRODUCTION: Combination vaccines containing whole-cell pertussis antigens were phased out from the Australian national immunisation programme between 1997 and 1999 and replaced by the less reactogenic acellular pertussis (aP) antigens. In a large case-control study of Australian children born during the transition period, those with allergist diagnosed IgE-mediated food allergy were less likely to have received whole-cell vaccine in early infancy than matched population controls (OR: 0.77 (95% CI, 0.62 to 0.95)). We hypothesise that a single dose of whole-cell vaccine in early infancy is protective against IgE-mediated food allergy. METHODS AND ANALYSIS: This adaptive double-blind randomised controlled trial is investigating whether a mixed whole-cell/aP vaccine schedule prevents allergic disease in the first year of life. The primary outcome is IgE-mediated food allergy by 12 months of age. Secondary outcomes include new onset of atopic dermatitis by 6 or 12 months of age; sensitisation to at least one allergen by 12 months of age; seroconversion in anti-pertussis toxin IgG titres after vaccination with aP booster at 18 months of age; and solicited systemic and local adverse events following immunisation with pertussis-containing vaccines. Analyses will be performed using a Bayesian group sequential design. ETHICS AND DISSEMINATION: This study has been approved by the Child and Adolescent Health Service Human Research Ethics Committee, Perth, Western Australia (RGS 00019). The investigators will ensure that this trial is conducted in accordance with the principles of the Declaration of Helsinki and with the International Conference on Harmonisation Guidelines for Good Clinical Practice. Individual consent will be requested. Parents will be reimbursed reasonable travel and parking costs to attend the study visits. The dissemination of these research findings will follow the National Health and Medical Research Council of Australia Open Access Policy. TRIAL REGISTRATION NUMBER: ACTRN12617000065392p.

Assessing the strength of evidence for a causal effect of respiratory syncytial virus lower respiratory tract infections on subsequent wheezing illness: a systematic review and meta-analysis.

BACKGROUND: Although a positive association has been established, it is unclear whether lower respiratory tract infections (LRTIs) with respiratory syncytial virus (RSV) cause chronic wheezing illnesses. If RSV-LRTI were causal, we would expect RSV-LRTI prevention to reduce the incidence of chronic wheezing illnesses in addition to reducing acute disease. We aimed to evaluate the strength of evidence for a causal effect of RSV-LRTI on subsequent chronic wheezing illness to inform public health expectations for RSV vaccines. METHODS: We did a systematic review and meta-analysis of observational studies evaluating the association between RSV-LRTI and subsequent wheezing illness (exposure studies) and studies evaluating the association between RSV immunoprophylaxis and subsequent wheezing illness (immunoprophylaxis studies). Exposure studies were included if the exposure group members had an LRTI with laboratory-confirmed RSV and if the exposure ascertainment period began before 2 years of age and ended before 5 years of age. We required a wash-out period of more than 30 days between the index RSV-LRTI and the outcome measurement to allow for resolution of the acute illness. Comparisons between RSV-LRTI and non-RSV-LRTI were not included. Immunoprophylaxis studies were included if they measured the association with subsequent wheezing illness relative to a control group, either in a randomised controlled trial (RCT) or an observational design. For the immunoprophylaxis drugs in question, we required evidence of efficacy in targeting RSV-LRTI from at least one RCT to ensure biological plausibility. All variations of wheezing illness were combined into a single outcome that refers broadly to asthma or any other respiratory illness with wheezing symptoms. Ovid MEDLINE and Embase databases were searched from inception up to Aug 28, 2018. We evaluated whether data from exposure studies could provide evidence against the most viable non-causal theory that RSV-LRTI is a marker of respiratory illness susceptibility rather than a causal factor. Additionally, we tested whether RSV immunoprophylaxis reduces the odds of subsequent wheezing illnesses. We used a random-effects modelling framework and, to accommodate studies providing multiple correlated estimates, robust variance estimation meta-regressions. Meta-regression coefficients (b) quantify differences between exposure and comparator groups on the loge odds ratio (loge OR) scale. FINDINGS: From 14 235 records we identified 57 eligible articles that described 42 studies and provided 153 effect estimates. 35 studies estimated the direct effect of RSV-LRTI on wheezing illnesses (exposure studies) and eight evaluated the effect of RSV immunoprophylaxis (immunoprophylaxis studies). Exposure studies that adjusted for genetic influences yielded a smaller mean adjusted OR estimate (aOR+ 2·45, 95% CI 1·23-4·88) compared with those that did not (4·17, 2·36-7·37), a significant difference (b 0·53, 95% CI 0·04-1·02). Infants who were not protected with RSV immunoprophylaxis tended to have higher odds of subsequent wheezing illness, as we would expect if RSV-LRTI were causal, but the effect was not significant (OR+ 1·21, 95% CI 0·73-1·99). There was generally a high threat of confounding bias in the observational studies. Additionally, in both the observational studies and immunoprophylaxis RCTs, there was high risk of bias due to missing outcome data. INTERPRETATION: Our findings, limited to exposure and immunoprophylaxis studies, do not support basing policy decisions on an assumption that prevention of RSV-LRTI will reduce recurrent chronic wheezing illnesses. FUNDING: Bill & Melinda Gates Foundation.

Neonatal genetics of gene expression reveal potential origins of autoimmune and allergic disease risk.

Chronic immune-mediated diseases of adulthood often originate in early childhood. To investigate genetic associations between neonatal immunity and disease, we map expression quantitative trait loci (eQTLs) in resting myeloid cells and CD4+ T cells from cord blood samples, as well as in response to lipopolysaccharide (LPS) or phytohemagglutinin (PHA) stimulation, respectively. Cis-eQTLs are largely specific to cell type or stimulation, and 31% and 52% of genes with cis-eQTLs have response eQTLs (reQTLs) in myeloid cells and T cells, respectively. We identified cis regulatory factors acting as mediators of trans effects. There is extensive colocalisation between condition-specific neonatal cis-eQTLs and variants associated with immune-mediated diseases, in particular CTSH had widespread colocalisation across diseases. Mendelian randomisation shows causal neonatal gene expression effects on disease risk for BTN3A2, HLA-C and others. Our study elucidates the genetics of gene expression in neonatal immune cells, and aetiological origins of autoimmune and allergic diseases.

Rewiring of gene networks underlying mite allergen-induced CD4 + Th-cell responses during immunotherapy.

BACKGROUND: Multiple regulatory mechanisms have been identified employing conventional hypothesis-driven approaches as contributing to allergen-specific immunotherapy outcomes, but understanding of how these integrate to maintain immunological homeostasis is incomplete. OBJECTIVE: To explore the potential for unbiased systems-level gene co-expression network analysis to advance understanding of immunotherapy mechanisms. METHODS: We profiled genome-wide allergen-induced Th-cell responses prospectively during 24 months subcutaneous immunotherapy (SCIT) in 25 rhinitis, documenting changes in immunoinflammatory pathways and associated co-expression networks and their relationships to symptom scores out to 36 months. RESULTS: Prior to immunotherapy, mite-induced Th-cell response networks involved multiple discrete co-expression modules including those related to Th2-, type1 IFN-, inflammation- and FOXP3/IL2-associated signalling. A signature comprising 109 genes correlated with symptom scores, and these mapped to cytokine signalling/T-cell activation-associated pathways, with upstream drivers including hallmark Th1/Th2- and inflammation-associated genes. Reanalysis after 3.5 months SCIT updosing detected minimal changes to pathway/upstream regulator profiles despite 32.5% symptom reduction; however, network analysis revealed underlying merging of FOXP3/IL2-with inflammation-and Th2-associated modules. By 12 months SCIT, symptoms had reduced by 41% without further significant changes to pathway/upstream regulator or network profiles. Continuing SCIT to 24 months stabilized symptoms at 47% of baseline, accompanied by upregulation of the type1 IFN-associated network module and its merging into the Th2/FOXP3/IL2/inflammation module. CONCLUSIONS: Subcutaneous immunotherapy stimulates progressive integration of mite-induced Th cell-associated Th2-, FOXP3/IL2-, inflammation- and finally type1 IFN-signalling subnetworks, forming a single highly integrated co-expression network module, maximizing potential for stable homeostatic control of allergen-induced Th2 responses via cross-regulation. Th2-antagonistic type1 IFN signalling may play a key role in stabilizing clinical effects of SCIT.