Genomic attributes of airway commensal bacteria and mucosa.

Microbial communities at the airway mucosal barrier are conserved and highly ordered, in likelihood reflecting co-evolution with human host factors. Freed of selection to digest nutrients, the airway microbiome underpins cognate management of mucosal immunity and pathogen resistance. We show here the initial results of systematic culture and whole-genome sequencing of the thoracic airway bacteria, identifying 52 novel species amongst 126 organisms that constitute 75% of commensals typically present in heathy individuals. Clinically relevant genes encode antimicrobial synthesis, adhesion and biofilm formation, immune modulation, iron utilisation, nitrous oxide (NO) metabolism and sphingolipid signalling. Using whole-genome content we identify dysbiotic features that may influence asthma and chronic obstructive pulmonary disease. We match isolate gene content to transcripts and metabolites expressed late in airway epithelial differentiation, identifying pathways to sustain host interactions with microbiota. Our results provide a systematic basis for decrypting interactions between commensals, pathogens, and mucosa in lung diseases of global significance.

IL-1α is required for T cell-driven weight loss after respiratory viral infection.

Respiratory viral infections remain a major cause of hospitalization and death worldwide. Patients with respiratory infections often lose weight. While acute weight loss is speculated to be a tolerance mechanism to limit pathogen growth, severe weight loss following infection can cause quality of life deterioration. Despite the clinical relevance of respiratory infection-induced weight loss, its mechanism is not yet completely understood. We utilized a model of CD 8+ T cell-driven weight loss during respiratory syncytial virus (RSV) infection to dissect the immune regulation of post-infection weight loss. Supporting previous data, bulk RNA sequencing indicated significant enrichment of the interleukin (IL)-1 signaling pathway after RSV infection. Despite increased viral load, infection-associated weight loss was significantly reduced after IL-1α (but not IL-1ß) blockade. IL-1α depletion resulted in a reversal of the gut microbiota changes observed following RSV infection. Direct nasal instillation of IL-1α also caused weight loss. Of note, we detected IL-1α in the brain after either infection or nasal delivery. This was associated with changes in genes controlling appetite after RSV infection and corresponding changes in signaling molecules such as leptin and growth/differentiation factor 15. Together, these findings indicate a lung-brain-gut signaling axis for IL-1α in regulating weight loss after RSV infection.

Crosstalk with lung fibroblasts shapes the growth and therapeutic response of mesothelioma cells.

Mesothelioma is an aggressive cancer of the mesothelial layer associated with an extensive fibrotic response. The latter is in large part mediated by cancer-associated fibroblasts which mediate tumour progression and poor prognosis. However, understanding of the crosstalk between cancer cells and fibroblasts in this disease is mostly lacking. Here, using co-cultures of patient-derived mesothelioma cell lines and lung fibroblasts, we demonstrate that fibroblast activation is a self-propagated process producing a fibrotic extracellular matrix (ECM) and triggering drug resistance in mesothelioma cells. Following characterisation of mesothelioma cells/fibroblasts signalling crosstalk, we identify several FDA-approved targeted therapies as far more potent than standard-of-care Cisplatin/Pemetrexed in ECM-embedded co-culture spheroid models. In particular, the SRC family kinase inhibitor, Saracatinib, extends overall survival well beyond standard-of-care in a mesothelioma genetically-engineered mouse model. In short, we lay the foundation for the rational design of novel therapeutic strategies targeting mesothelioma/fibroblast communication for the treatment of mesothelioma patients.

Delayed acquisition of airway commensals in antibiotic naïve children and its relationship with wheezing in rural Ecuador.

Introduction: The hygiene hypothesis identified a relationship between living in rural areas and acquiring protective environmental factors against the development of asthma and atopy. In our previous study, we found a correlation between particular bacterial species and early-onset wheezing in infants from the rural tropics of Ecuador who were corticosteroid-naïve and had limited antibiotic exposure. We now describe a longitudinal study of infants conducted to determine the age-related changes of the microbiome and its relationship with wheezing. Methods: We performed an amplicon sequencing of the 16S rRNA bacterial gene from the oropharyngeal samples obtained from 110 infants who had a history of recurrent episodic wheezing sampled at different ages (7, 12, and 24 months) and compared it to the sequencing of the oropharyngeal samples from 150 healthy infants sampled at the same time points. Bioinformatic analyses were conducted using QIIME and R. Results: As expected, the microbiota diversity consistently increased as the infants grew older. Considering age-based microbiota changes, we found that infants with wheeze had significantly lower species richness than the healthy infants at 7 months, but not at 12 or 24 months. Most of the core and accessory organisms increased in abundance and prevalence with age, except for a few which decreased. At 7 months of age, infants with wheeze had notably higher levels of a single Streptococcus operational taxonomic unit and core microbiota member than controls. Conclusions: In a cohort with limited antibiotic and corticosteroid use, a progressively more complex and diverse respiratory microbial community develops with age. The respiratory microbiota in early life is altered in infants with wheeze, but this does not hold true in older infants.

Molecular assessment of mycobacterial burden in the treatment of nontuberculous mycobacterial disease.

Introduction: Nontuberculous pulmonary disease causes significant morbidity and mortality. Efforts to tackle infections are hampered by the lack of reliable biomarkers for diagnosis, assessment and prognostication. The aim of this study was to develop molecular assays capable of identifying and quantifying multiple nontuberculous mycobacterial (NTM) species and to examine their utility in following individual patients' clinical courses. Methods: DNA was extracted from 410 sputum samples obtained longitudinally from a cohort of 38 patients who were commencing treatment for either Mycobacterium abscessus or Mycobacterium avium complex or who were patients with bronchiectasis who had never had positive cultures for mycobacteria. NTM quantification was performed with quantitative PCR assays developed in-house. Results: The molecular assays had high in vitro sensitivity and specificity for the detection and accurate quantification of NTM species. The assays successfully identified NTM DNA from human sputum samples (in vivo sensitivity: 0.86-0.87%; specificity: 0.62-0.95%; area under the curve: 0.74-0.92). A notable association between NTM copy number and treatment (Friedman ANOVA (df)=22.8 (3), p≤0.01 for M. abscessus treatment group) was also demonstrated. Conclusion: The quantitative PCR assays developed in this study provide affordable, real-time and rapid measurement of NTM burden, with significant implications for prompt management decisions.

An invisible threat? Aspergillus positive cultures and co-infecting bacteria in airway samples.

BACKGROUND: Aspergillus fumigatus (Af) infection is associated with poor lung health in chronic suppurative lung diseases but often goes undetected. We hypothesised that inhibition of Af growth by Pseudomonas aeruginosa (Pa) increases the frequency of false-negative Af culture in co-infected people. Using a substantial group of cystic fibrosis (CF) airway samples, we assessed the relationship between Af and bacterial pathogens, additionally comparing fungal culture with next-generation sequencing. METHODS: Frequency of co-culture was assessed for 44,554 sputum/BAL cultures, from 1,367 CF patients between the years 2010-2020. In a subgroup, Internal Transcribed Spacer-2 (ITS2) fungal sequencing was used to determine sequencing-positive, culture-negative (S+/C-) rates. RESULTS: Pa+ samples were nearly 40% less likely (P<0.0001) than Pa- samples to culture Af, an effect that was also seen with some other Gram-negative isolates. This impact varied with Pa density and appeared to be moderated by Staphylococcus aureus co-infection. Sequencing identified Af-S+/C- for 40.1% of tested sputa. Samples with Pa had higher rates of Af-S+/C- (49.3%) than those without (35.7%; RR 1.38 [1.02-1.93], P<0.05). Af-S+/C- rate was not changed by other common bacterial infections. Pa did not affect the S+/C- rates of Candida, Exophiala or Scedosporium. CONCLUSIONS: Pa/ Af co-positive cultures are less common than expected in CF. Our findings suggest an Af-positive culture is less likely in the presence of Pa. Interpretation of negative cultures should be cautious, particularly in Pa-positive samples, and a companion molecular diagnostic could be useful. Further work investigating mechanisms, alternative detection techniques and other chronic suppurative lung diseases is needed.

M ycobacterium avium complex genomics and transmission in a London hospital.

BACKGROUND: Non-tuberculous mycobacteria (NTM) are environmental microorganisms and opportunistic pathogens in individuals with pre-existing lung conditions such as cystic fibrosis (CF) and non-CF bronchiectasis. While recent studies of Mycobacterium abscessus have identified transmission within single CF centres as well as nationally and globally, transmission of other NTM species is less well studied. METHODS: To investigate the potential for transmission of the Mycobacterium avium complex (MAC) we sequenced 996 isolates from 354 CF and non-CF patients at the Royal Brompton Hospital (London, UK; collected 2013-2016) and analysed them in a global context. Epidemiological links were identified from patient records. Previously published genomes were used to characterise global population structures. RESULTS: We identified putative transmission clusters in three MAC species, although few epidemiological links could be identified. For M. avium, lineages were largely limited to single countries, while for Mycobacterium chimaera, global transmission clusters previously associated with heater-cooler units (HCUs) were found. However, the immediate ancestor of the lineage causing the major HCU-associated outbreak was a lineage already circulating in patients. CONCLUSIONS: CF and non-CF patients shared transmission chains, although the lack of epidemiological links suggested that most transmission is indirect and may involve environmental intermediates or asymptomatic carriage in the wider population.

Evidence of immunometabolic dysregulation and airway dysbiosis in athletes susceptible to respiratory illness.

BACKGROUND: Respiratory tract infection (RTI) is a leading cause of training and in-competition time-loss in athlete health. The immune factors associated with RTI susceptibility remain unclear. In this study, we prospectively characterise host immune factors in elite athletes exhibiting RTI susceptibility. METHODS: Peripheral blood lymphocyte flow cytometry phenotyping and 16S rRNA microbial sequencing of oropharyngeal swabs was performed in a prospective elite athlete cohort study (n = 121). Mass cytometry, peripheral blood mononuclear cell (PBMC) stimulation and plasma metabolic profiling was performed in age-matched highly-susceptible (HS) athletes (≥4RTI in last 18 months) (n = 22) compared to non-susceptible (NS) (≤1RTI in last 18 months) (n = 23) athletes. Findings were compared to non-athletic healthy controls (HC) (n = 19). FINDINGS: Athletes (n = 121) had a reduced peripheral blood memory T regulatory cell compartment compared to HC (p = 0.02 (95%CI:0.1,1.0)) and reduced upper airway bacterial biomass compared to HC (p = 0.032, effect size r = 0.19). HS athletes (n = 22) had lower circulating memory T regulatory cells compared to NS (n = 23) athletes (p = 0.005 (95%CI:-1.5,-0.15)) and HC (p = 0.002 (95%CI:-1.9,-0.3) with PBMC microbial stimulation assays revealing a T-helper 2 skewed immune response compared to HC. Plasma metabolomic profiling showed differences in sphingolipid pathway metabolites (a class of lipids important in infection and inflammation regulation) in HS compared to NS athletes and HC, with sphingomyelin predictive of RTI infection susceptibility (p = 0.005). INTERPRETATION: Athletes susceptible to RTI have reduced circulating memory T regulatory cells, metabolic dysregulation of the sphingolipid pathway and evidence of upper airway bacterial dysbiosis. FUNDING: This study was funded by the English Institute of Sport (UK).

Airway mucins promote immunopathology in virus-exacerbated chronic obstructive pulmonary disease.

The respiratory tract surface is protected from inhaled pathogens by a secreted layer of mucus rich in mucin glycoproteins. Abnormal mucus accumulation is a cardinal feature of chronic respiratory diseases, but the relationship between mucus and pathogens during exacerbations is poorly understood. We identified elevations in airway mucin 5AC (MUC5AC) and MUC5B concentrations during spontaneous and experimentally induced chronic obstructive pulmonary disease (COPD) exacerbations. MUC5AC was more sensitive to changes in expression during exacerbation and was therefore more predictably associated with viral load, inflammation, symptom severity, decrements in lung function, and secondary bacterial infections. MUC5AC was functionally related to inflammation, as Muc5ac-deficient (Muc5ac-/-) mice had attenuated RV-induced (RV-induced) airway inflammation, and exogenous MUC5AC glycoprotein administration augmented inflammatory responses and increased the release of extracellular adenosine triphosphate (ATP) in mice and human airway epithelial cell cultures. Hydrolysis of ATP suppressed MUC5AC augmentation of RV-induced inflammation in mice. Therapeutic suppression of mucin production using an EGFR antagonist ameliorated immunopathology in a mouse COPD exacerbation model. The coordinated virus induction of MUC5AC and MUC5B expression suggests that non-Th2 mechanisms trigger mucin hypersecretion during exacerbations. Our data identified a proinflammatory role for MUC5AC during viral infection and suggest that MUC5AC inhibition may ameliorate COPD exacerbations.

Sputum Proteomics in Nontuberculous Mycobacterial Lung Disease.

BACKGROUND: Nontuberculous mycobacterial (NTM) infections are difficult to diagnose and treat. Biomarkers to identify patients with active infection or at risk of disease progression would have clinical utility. Sputum is the most frequently used matrix for the diagnosis of NTM lung disease. RESEARCH QUESTION: Can sputum proteomics be used to identify NTM-associated inflammatory profiles in sputum? STUDY DESIGN AND METHODS: Patients with NTM lung disease and a matched cohort of patients with COPD, bronchiectasis (BE), and cystic fibrosis (CF) without NTM lung disease were enrolled from two hospitals in the United Kingdom. Liquid chromatography-tandem mass spectrometry was used to identify proteomic biomarkers associated with underlying diagnosis (COPD, BE, and CF), the presence of NTM lung disease defined according to American Thoracic Society/Infectious Diseases Society of America criteria, and severity of NTM. A subset of patients receiving guideline-concordant NTM treatment were studied to identify protein changes associated with treatment response. RESULTS: This study analyzed 95 sputum samples from 55 subjects (BE, n = 21; COPD, n = 19; CF, n = 15). Underlying disease and infection with Pseudomonas aeruginosa were the strongest drivers of sputum protein profiles. Comparing protein abundance in COPD, BE, and CF found that 12 proteins were upregulated in CF compared with COPD, including MPO, AZU1, CTSG, CAT, and RNASE3, with 21 proteins downregulated, including SCGB1A1, IGFBP2, SFTPB, GC, and CFD. Across CF, BE, and COPD, NTM infection (n = 15) was not associated with statistically significant differences in sputum protein profiles compared with those without NTM. Two proteins associated with iron chelation were significantly downregulated in severe NTM disease. NTM treatment was associated with heterogeneous changes in the sputum protein profile. Patients with NTM and a decrease in immune response proteins had a subjective symptomatic improvement. INTERPRETATION: Sputum proteomics identified candidate biomarkers of NTM severity and treatment response. However, underlying lung disease and typical bacterial pathogens such as P aeruginosa are also key determinants of the sputum proteomic profile.

Integrated genomics point to immune vulnerabilities in pleural mesothelioma.

Pleural mesothelioma is an aggressive malignancy with limited effective therapies. In order to identify therapeutic targets, we integrated SNP genotyping, sequencing and transcriptomics from tumours and low-passage patient-derived cells. Previously unrecognised deletions of SUFU locus (10q24.32), observed in 21% of 118 tumours, resulted in disordered expression of transcripts from Hedgehog pathways and the T-cell synapse including VISTA. Co-deletion of Interferon Type I genes and CDKN2A was present in half of tumours and was a predictor of poor survival. We also found previously unrecognised deletions in RB1 in 26% of cases and show sub-micromolar responses to downstream PLK1, CHEK1 and Aurora Kinase inhibitors in primary mesothelioma cells. Defects in Hippo pathways that included RASSF7 amplification and NF2 or LATS1/2 mutations were present in 50% of tumours and were accompanied by micromolar responses to the YAP1 inhibitor Verteporfin. Our results suggest new therapeutic avenues in mesothelioma and indicate targets and biomarkers for immunotherapy.

Airway microbial communities, smoking and asthma in a general population sample.

BACKGROUND: Normal airway microbial communities play a central role in respiratory health but are poorly characterized. Cigarette smoking is the dominant global environmental influence on lung function, and asthma has become the most prevalent chronic respiratory disease worldwide. Both conditions have major microbial components that are incompletely defined. METHODS: We investigated airway bacterial communities in a general population sample of 529 Australian adults. Posterior oropharyngeal swabs were analyzed by sequencing of the 16S rRNA gene. The microbiota were characterized according to their prevalence, abundance and network memberships. FINDINGS: The microbiota were similar across the general population, and were strongly organized into co-abundance networks. Smoking was associated with diversity loss, negative effects on abundant taxa, profound alterations to network structure and expansion of Streptococcus spp. By contrast, the asthmatic microbiota were selectively affected by an increase in Neisseria spp. and by reduced numbers of low abundance but prevalent organisms. INTERPRETATION: Our study shows that the healthy airway microbiota in this population were contained within a highly structured ecosystem, suggesting balanced relationships between the microbiome and human host factors. The marked abnormalities in smokers may contribute to chronic obstructive pulmonary disease (COPD) and lung cancer. The narrow spectrum of abnormalities in asthmatics encourages investigation of damaging and protective effects of specific bacteria. FUNDING: The study was funded by the Asmarley Trust and a Wellcome Joint Senior Investigator Award to WOCC and MFM (WT096964MA and WT097117MA). The Busselton Healthy Ageing Study is supported by the Government of Western Australia (Office of Science, Department of Health) the City of Busselton, and private donations.

Y disruption, autosomal hypomethylation and poor male lung cancer survival.

Lung cancer is the most frequent cause of cancer death worldwide. It affects more men than women, and men generally have worse survival outcomes. We compared gene co-expression networks in affected and unaffected lung tissue from 126 consecutive patients with Stage IA-IV lung cancer undergoing surgery with curative intent. We observed marked degradation of a sex-associated transcription network in tumour tissue. This disturbance, detected in 27.7% of male tumours in the discovery dataset and 27.3% of male tumours in a further 123-sample replication dataset, was coincident with partial losses of the Y chromosome and extensive autosomal DNA hypomethylation. Central to this network was the epigenetic modifier and regulator of sexually dimorphic gene expression, KDM5D. After accounting for prognostic and epidemiological covariates including stage and histology, male patients with tumour KDM5D deficiency showed a significantly increased risk of death (Hazard Ratio [HR] 3.80, 95% CI 1.40-10.3, P = 0.009). KDM5D deficiency was confirmed as a negative prognostic indicator in a further 1100 male lung tumours (HR 1.67, 95% CI 1.4-2.0, P = 1.2 × 10-10). Our findings identify tumour deficiency of KDM5D as a prognostic marker and credible mechanism underlying sex disparity in lung cancer.

ORMDL3 regulates poly I:C induced inflammatory responses in airway epithelial cells.

BACKGROUND: Oroscomucoid 3 (ORMDL3) has been linked to susceptibility of childhood asthma and respiratory viral infection. Polyinosinic-polycytidylic acid (poly I:C) is a synthetic analog of viral double-stranded RNA, a toll-like receptor 3 (TLR3) ligand and mimic of viral infection. METHODS: To investigate the functional role of ORMDL3 in the poly I:C-induced inflammatory response in airway epithelial cells, ORMDL3 knockdown and over-expression models were established in human A549 epithelial cells and primary normal human bronchial epithelial (NHBE) cells. The cells were stimulated with poly I:C or the Th17 cytokine IL-17A. IL-6 and IL-8 levels in supernatants,  mRNA levels of genes in the TLR3 pathway and inflammatory response from cell pellets were measured. ORMDL3 knockdown models in A549 and BEAS-2B epithelial cells were then infected with live human rhinovirus (HRV16) followed by IL-6 and IL-8 measurement. RESULTS: ORMDL3 knockdown and over-expression had little influence on the transcript levels of TLR3 in airway epithelial cells. Time course studies showed that ORMDL3-deficient A549 and NHBE cells had an attenuated IL-6 and IL-8 response to poly I:C stimulation. A549 and NHBE cells over-expressing ORMDL3 released relatively more IL-6 and IL-8 following poly I:C stimulation. IL-17A exhibited a similar inflammatory response in ORMDL3 knockdown and over-expressing cells, but co-stimulation of poly I:C and IL-17A did not significantly enhance the IL-6 and IL-8 response. Transcript abundance of IFNB following poly I:C stimulation was not significantly altered by ORMDL3 knockdown or over-expression. Dampening of the IL-6 response by ORMDL3 knockdown was confirmed in HRV16 infected BEAS-2B and A549 cells. CONCLUSIONS: ORMDL3 regulates the viral inflammatory response in airway epithelial cells via mechanisms independent of the TLR3 pathway.

A large-scale genome-wide association analysis of lung function in the Chinese population identifies novel loci and highlights shared genetic aetiology with obesity.

BACKGROUND: Lung function is a heritable complex phenotype with obesity being one of its important risk factors. However, knowledge of their shared genetic basis is limited. Most genome-wide association studies (GWASs) for lung function have been based on European populations, limiting the generalisability across populations. Large-scale lung function GWASs in other populations are lacking. METHODS: We included 100 285 subjects from the China Kadoorie Biobank (CKB). To identify novel loci for lung function, single-trait GWAS analyses were performed on forced expiratory volume in 1 s (FEV1), forced vital capacity (FVC) and FEV1/FVC in the CKB. We then performed genome-wide cross-trait analysis between lung function and obesity traits (body mass index (BMI), BMI-adjusted waist-to-hip ratio and BMI-adjusted waist circumference) to investigate the shared genetic effects in the CKB. Finally, polygenic risk scores (PRSs) of lung function were developed in the CKB and their interaction with BMI's association on lung function were examined. We also conducted cross-trait analysis in parallel with the CKB using up to 457 756 subjects from the UK Biobank (UKB) for replication and investigation of ancestry-specific effects. RESULTS: We identified nine genome-wide significant novel loci for FEV1, six for FVC and three for FEV1/FVC in the CKB. FEV1 and FVC showed significant negative genetic correlation with obesity traits in both the CKB and UKB. Genetic loci shared between lung function and obesity traits highlighted important biological pathways, including cell proliferation, embryo, skeletal and tissue development, and regulation of gene expression. Mendelian randomisation analysis suggested significant negative causal effects of BMI on FEV1 and on FVC in both the CKB and UKB. Lung function PRSs significantly modified the effect of change in BMI on change in lung function during an average follow-up of 8 years. CONCLUSION: This large-scale GWAS of lung function identified novel loci and shared genetic aetiology between lung function and obesity. Change in BMI might affect change in lung function differently according to a subject's polygenic background. These findings may open new avenues for the development of molecular-targeted therapies for obesity and lung function improvement.

The fungal airway microbiome in cystic fibrosis and non-cystic fibrosis bronchiectasis.

BACKGROUND: The prevalence of fungal disease in cystic fibrosis (CF) and non-CF bronchiectasis is increasing and the clinical spectrum is widening. Poor sensitivity and a lack of standard diagnostic criteria renders interpretation of culture results challenging. In order to develop effective management strategies, a more accurate and comprehensive understanding of the airways fungal microbiome is required. The study aimed to use DNA sequences from sputum to assess the load and diversity of fungi in adults with CF and non-CF bronchiectasis. METHODS: Next generation sequencing of the ITS2 region was used to examine fungal community composition (n = 176) by disease and underlying clinical subgroups including allergic bronchopulmonary aspergillosis, chronic necrotizing pulmonary aspergillosis, non-tuberculous mycobacteria, and fungal bronchitis. Patients with no known active fungal disease were included as disease controls. RESULTS: ITS2 sequencing greatly increased the detection of fungi from sputum. In patients with CF fungal diversity was lower, while burden was higher than those with non-CF bronchiectasis. The most common operational taxonomic unit (OTU) in patients with CF was Candida parapsilosis (20.4%), whereas in non-CF bronchiectasis sputum Candida albicans (21.8%) was most common. CF patients with overt fungal bronchitis were dominated by Aspergillus spp., Exophiala spp., Candida parapsilosis or Scedosporium spp. CONCLUSION: This study provides a framework to more accurately characterize the extended spectrum of fungal airways diseases in adult suppurative lung diseases.

Estimating cell-type-specific DNA methylation effects in heterogeneous cellular populations.

Aim: To develop a method for estimating cell-specific effects in epigenomic association studies in the presence of cell type heterogeneity. Materials & methods: We utilized Monte Carlo Expectation-Maximization algorithm with Metropolis-Hastings sampler to reconstruct the 'missing' cell-specific methylations and to estimate their associations with phenotypes free of confounding by cell type proportions. Results: Simulations showed reliable performance of the method under various settings including when the cell type is rare. Application to a real dataset recapitulated the directly measured cell-specific methylation pattern in whole blood. Conclusion: This work provides a framework to identify important cell groups and account for cell type composition useful for studying the role of epigenetic changes in human traits and diseases.

Neutrophilic inflammation in the respiratory mucosa predisposes to RSV infection.

The variable outcome of viral exposure is only partially explained by known factors. We administered respiratory syncytial virus (RSV) to 58 volunteers, of whom 57% became infected. Mucosal neutrophil activation before exposure was highly predictive of symptomatic RSV disease. This was associated with a rapid, presymptomatic decline in mucosal interleukin-17A (IL-17A) and other mediators. Conversely, those who resisted infection showed presymptomatic activation of IL-17- and tumor necrosis factor-related pathways. Vulnerability to infection was not associated with baseline microbiome but was reproduced in mice by preinfection chemokine-driven airway recruitment of neutrophils, which caused enhanced disease mediated by pulmonary CD8+ T cell infiltration. Thus, mucosal neutrophilic inflammation at the time of RSV exposure enhances susceptibility, revealing dynamic, time-dependent local immune responses before symptom onset and explaining the as-yet unpredictable outcomes of pathogen exposure.