BACKGROUND:Platelets can recognize and destroy microbial pathogens through the release of antimicrobial proteins, suggesting that they may contribute to innate immune defense against infection. OBJECTIVES:This study aimed to determine whether platelets contribute to pulmonary innate immunity during bacterial pneumonia and to distinguish their antimicrobial functions from their hemostatic role. METHODS:Mice were exposed to aerosolized Gram-negative Pseudomonas aeruginosa or Gram-positive Streptococcus pneumoniae. Platelet number or function was reduced by antibody-mediated platelet depletion, thrombopoietin gene deletion, or aspirin treatment. Clinical outcomes, mortality, pulmonary pathogen burden, and lung histopathology were assessed. RESULTS:Thrombocytopenic and aspirin-treated mice had increased mortality following bacterial pneumonia. Lung congestion and pathogen burden increased in proportion to the severity of thrombocytopenia. In contrast, pulmonary hemorrhage was observed only in severely thrombocytopenic mice, indicating that impaired antimicrobial defense occurred at platelet levels sufficient to maintain hemostasis. CONCLUSION:Platelets actively protect against both Gram-negative and Gram-positive bacterial pneumonia. This protective effect is distinct from their hemostatic function: platelets contribute directly to antimicrobial host defense, whereas their hemostatic activity maintains a physical barrier that limits local pathogen spread.

BACKGROUND:Acute respiratory distress syndrome (ARDS) causes significant morbidity and mortality during viral pneumonia, including SARS-CoV-2 infections. Nevertheless, most patients with SARS-CoV-2 infections recover seamlessly without developing ARDS, suggesting the existence of endogenous pathways to protect the lungs. Since microRNAs (miRNAs) can regulate endogenous molecular pathways involved in lung protection, we hypothesized that alveolar miRNAs could function to dampen SARS-CoV-2-associated lung injury. METHODS:Screening studies in human alveolar epithelial cells and SARS-CoV-2-infected mice were performed to identify miRNAs induced during infection. Candidate miRNAs were confirmed via RT-qPCR. The role of hypoxia-inducible factor 1A (HIF1A) in regulating miRNA expression was examined in molecular studies. Loss- and gain-of-function approaches in a murine SARS-CoV-2 ARDS model were used to assess the physiological relevance of miRNA. Viral sequence analyses and site-directed mutagenesis were used to determine direct miRNA - viral RNA interactions. RESULTS:Screening studies identified miR-147b (hsa-miR-147b-3p or mmu-miR-147-3p) as the leading candidate during infection of human alveolar epithelia or mice with SARS-CoV-2. Functional and molecular studies implicate HIF1A in miR-147b induction during alveolar injury or SARS-CoV-2 infection. Studies in mice with induced deletion of miR-147b in alveolar epithelia (miR147fl/fl SPC-CreER mice) or nano-particle-mediated miR-147b overexpression revealed a protective role of alveolar-expressed miR-147b during murine SARS-CoV-2-associated ARDS. Moreover, we identified the ORF8 region within the SARS-CoV-2 template strand as a direct target of miR-147b, with an ORF8 silent mutation of the SARS-CoV-2 miR-147b-binding site abolishing the observed protection in vitro and in vivo. CONCLUSIONS:Alveolar epithelial cell-derived miR-147b serves as an endogenous lung protective miRNA against SARS-CoV-2-associated ARDS by directly targeting virus-encoded RNA, revealing a previously unrecognized antiviral and lung-protective mechanism.

Facultative stem cells including lung alveolar type 2 (AT2) cells must toggle between unipotent specialization at baseline and multipotent plasticity during injury-repair. The underlying molecular switches and epigenetic changes remain unclear yet dictate regenerative versus pathological outcomes. Using multiomics and mouse genetics, we show that AP-1 members FOS, FOSB, and JUNB promote an injury-induced transitional AT2 cell state and associated chromatin landscape. Joint transcriptomic-epigenomic profiling and immunostaining distinguish CLDN4+ AT2 cells as a KRT8high subset with open chromatin highly enriched for AP-1 motifs. JUNB and FOSB accumulate in these cells upon viral injury and, along with constitutive FOS, are required for CLDN4 induction, AT2 cell dispersion, senescence signaling, and fibroblast activation, while impacting region-specific alveolar type 1 (AT1) differentiation. AP-1 activation also occurs in mouse AT2 cells expressing oncogenic Kras and transitional cells in human lung tissues with premalignant or adenocarcinoma lesions. Our work refines AT2 transitional states and reveals a gene regulatory logic shared by tissue repair and tumorigenesis.

Infectious pneumonias affect millions of individuals annually worldwide, accounting for unacceptably high mortalities in many populations. Following pathogen introduction to the lower respiratory tract by inhalation or aspiration, complex interactions between the host and the microbe determine whether the microbe is cleared or establishes an infectious pneumonia. Even modest variations in the host immune response to pathogens may have potentially profound impacts on pathogenesis, severity, and eventual survival of infectious pneumonias. In this article, the authors summarize the most common conditions that alter host defenses in a manner that predisposes to development of pneumonia.

Coronaviruses can cause serious disease in humans and animals. They are often difficult to study due to the variability of the disease as well as safety concerns related to biosafety requirements. Here, we use a biological safety level (BSL) 2-compatible murine betacoronavirus (MHV-A59), which infects multiple organs, to study the respiratory phase of disease and subsequent systemic dissemination. We asked whether inducible epithelial resistance, activated by the combination of pattern recognition receptor agonists Pam2CSK4 (Pam2), synthetic diacylated lipopeptide, + oligodeoxynucleotide (ODN) M362, can limit both pulmonary infection and systemic spread of disease. Prophylactic exposure with Pam2 + ODN improves survival and reduces extrapulmonary viral burden, including in the liver, consistent with limited systemic spread. Stopping the infection at the level of the lungs is associated with improved physiological outcomes and increased protection for the mice. We further characterize the response of the epithelial cells to infection and treatment and demonstrate modulation of epithelial gene expression, including attenuation of virus-induced responses, to better understand the mechanisms of protection and how these responses may be leveraged against future infectious agents.NEW & NOTEWORTHY This study establishes a BSL 2-compatible murine betacoronavirus model to investigate respiratory and systemic disease. We show that inducible epithelial resistance via Pam2 + ODN improves survival, preserves lung function, and reduces extrapulmonary viral burden. Mechanistically, Pam2 + ODN reprograms lung epithelial gene expression, reversing virus-induced transcriptional responses. These findings support epithelial-targeted, pathogen-agnostic strategies to limit both pulmonary and systemic consequences of respiratory viral infection.

Invasive pulmonary mucormycosis (IPM) is a severe opportunistic mold infection whose outcome is predominantly host driven. Preclinical proof-of-concept studies and clinical case reports in salvage therapy settings suggested a benefit of immune checkpoint inhibitors (ICIs) in IPM management. However, the kinetics of infection-induced immune paralysis and optimal timing of ICI therapy remain poorly understood. Here, we performed sequential nCounter-based transcriptomics on lung tissue of cyclophosphamide-immunosuppressed mice with IPM (Rhizopus arrhizus infection) to dynamically study the pulmonary immune environment. Within 7 d after infection, lungs of mice with IPM showed reversal of early proinflammatory signaling, impaired T cell signaling, and upregulation of exhaustion markers. Similar immune paralysis signatures were seen in mice with invasive pulmonary aspergillosis and fusariosis. For therapeutic studies, Mucorales-active antifungal therapy with isavuconazonium sulfate (ISAV) was initiated on day 3 after IPM infection, along with anti-PD-L1 or a nontargeting isotype antibody (control) given either on days 3 + 5 (early) or days 6 + 8 (late). Both early and late adjunct anti-PD-L1 therapy were well tolerated and significantly improved morbidity/mortality outcomes compared to ISAV + isotype. Notably, early adjunct anti-PD-L1 therapy promoted significantly stronger innate immune cell activation, upregulation of key cytokine pathways, reinvigoration of T-helper-cell signaling, and reversal of IPM-induced exhaustion signals than both late adjunct anti-PD-L1 and isotype control. These findings indicate that combined antifungal and early immunomodulatory therapy may be an important strategy to intercept immune paralysis and improve outcomes in immunocompromised hosts with IPM, inviting further preclinical and clinical exploration of early host-directed interventions to treat deadly mold pneumonias.

Pneumonias remain a leading cause of death worldwide. Seeking novel strategies to protect susceptible patients, we have reported that inhaled delivery of a diacylated lipopeptide and a synthetic CpG oligodeoxynucleotide (ODN) protects animals against a broad range of infectious pneumonias by stimulating antimicrobial responses from the lung epithelium. Toll-like receptor 9 (TLR9) is well-established as the primary cellular receptor for CpG ODNs. However, we recently reported that ODNs also stimulate TLR9-independent generation of antimicrobial mitochondrial reactive oxygen species. By testing a variety of synthetic ODN molecules, we found that ODNs containing a phosphorothioate backbone, but not those with a phosphodiester backbone, induce TLR9-independent pathogen killing in lungs and improve mouse survival. Phosphorothioate-backboned ODN binds mitochondrial protein voltage-dependent anion channel 1 (VDAC1) at its N-terminus, initiating pneumonia-protective metabolic reprogramming in lung epithelial cells that yield the protective antimicrobial effect. Thus, the phosphorothioate backbone of ODN is a critical structural pattern that activates TLR9-independent, metabolically modulated innate immune protection that may be harnessed to protect vulnerable patients against pneumonia.

Prophylactic inhalation of the synergistic agents ODN M362 and Pam2CSK4 ("Pam2ODN") protects mice against allergic lung disease, including allergic inflammation caused by house dust mite (HDM). By preventing sensitization, Pam2ODN reduces HDM-induced eosinophilic and lymphocytic inflammation. How Pam2ODN affects interactions among lung epithelial cells, dendritic cells, and T cells to prevent eosinophilic lung inflammation remains unclear. In the present study, we show that a single inhaled dose of Pam2ODN before HDM sensitization reduces airway Th2 polarization without affecting Th1 or Treg responses. Furthermore, Pam2ODN pretreatment inhibits the recruitment of lung monocyte-derived dendritic cells (moDCs) and conventional Type 2 dendritic cells (DC2s), while preventing the HDM-induced decrease in conventional Type 1 dendritic cells (DC1s). Bulk RNA-seq of the whole lung reveals that Pam2ODN pretreatment restricts the expression of proinflammatory transcripts induced by HDM sensitization. This tolerogenic effect is also reflected at the single-cell level in lung epithelial cells, where proinflammatory transcripts, pathways, and chromatin accessibility are inhibited. These results indicate that Pam2ODN reprograms lung epithelial cells to attenuate allergen-induced Th2-promoting cytokines and DCs while maintaining the population of protective DC1s. These findings suggest a strategy to mitigate chronic allergic lung diseases.

Lower respiratory tract infections remain a leading cause of death worldwide, highlighting the need for host-directed therapies. We previously identified an inhaled immunostimulatory combination of Pam2CSK4 (Pam2) and CpG oligodeoxynucleotide ODN M362 (ODN) that synergistically protects against bacterial, viral, and fungal pneumonias by reprogramming lung epithelial defenses. Using RNA-seq, dual transcription factor ChIP-seq, and reverse phase protein array, we define the transcriptional basis of this synergy. Cooperative regulation by the AP-1 component cJun and the NF-κB subunit RelA is central to the Pam2+ODN response. cJun occupancy distinguishes synergistic genes, whereas RelA occupancy aligns with expression directionality. Promoter-proximal cJun occupancy facilitates enhanced RelA loading, enabling rapid transcriptional reprogramming that strengthens epithelial survival pathways and amplifies microbicidal effector programs. This AP-1-assisted NF-κB cooperation establishes inducible resistance in lung epithelium and provides a mechanistic foundation for host-directed therapies against respiratory infection.

The epithelial tree of the lung is shaped proximo-distally by airway smooth muscle cells (ASMCs), ductal myofibroblasts (DMFs), and, transiently, alveolar myofibroblasts (AMFs). Lineage tracing and snapshot imaging suggest the clearance of AMFs via apoptosis post-alveologenesis, although definitive evidence is lacking. Here, we generate an inducible BCL2 overexpression mouse allele to inhibit AMF apoptosis. Using three independent Cre drivers and single-cell RNA sequencing, we show that BCL2-rescued AMFs persist around distal alveolar ducts and alveoli and, unexpectedly, mature toward DMFs. Both normal DMFs and rescued DMF-like cells upregulate contractile proteins in a house dust mite-induced asthma model. Our findings demonstrate apoptosis as the chief mechanism of AMF clearance, as well as fate plasticity and pathophysiological convergence of lung mesenchymal cells of the epithelial axis.

Pneumonia is one of the deadliest and most common infections worldwide. Traditional management has focused on pathogen-directed therapy with antimicrobial drugs. However, continued increase of antimicrobial resistance represents a growing health care crisis and heightens the urgency of shifting the treatment focus outside of the traditional paradigm. Adjunctive and host-directed therapies (HDT) target the response to infection by either enhancing the host immune response or mitigating immunopathology. They encompass a broad range of treatments, repurposed medications, and interventions that alter how the host responds to infections. This review summarizes adjunctive and HDTs for bacterial, tuberculous, fungal, and viral pneumonia.

Rationale: Respiratory tract infections are a major cause of worldwide morbidity, and increased antimicrobial resistance requires development of novel anti-infective therapeutics. Host-based immunomodulatory therapeutics are a promising strategy to mitigate the risk of emerging pathogens. We have reported that the drug complex Pam2ODN, composed of the immunostimulatory pattern recognition receptor ligands Pam2CSK4 and ODNM362, stimulates synergistic innate immune responses from lung epithelial cells that protect against respiratory infections. In this study, we seek to integrate multi-Omics data from murine and human models to understand the mechanisms of these responses. Methods: Using imaging flow cytometry (IFC), we tested the efficacy of Pam2ODN in increasing the binding of transcription factors within H3k27ac accessible sites in lung epithelium. In vitro, we assessed the signaling response of mouse lung epithelial cells (MLE-15) and human lung epithelial cells (HBEC3kt) over 24h at 4-hour intervals. In vivo, we treated wildtype mice with Pam2ODN or PBS by nebulization, then analyzed single cell lung suspensions 24h after treatment. IFC allowed single-cell nuclear evaluation of expression (mean fluorescence intensity, MFI) and colocalization (bright detail similarity, BDS) of RelA, cJun and H3K27ac. RNAseq, RelA and cJun ChIPseq were performed in HBEC3kt cells 4h after treatment, aligned to GRCh38 human genome. Differential analysis with R was performed to corroborate gene expression and genome accessibility changes. Results: Both murine and human models showed RelA and cJun increased expression after Pam2ODN stimulation. Consistently, IFC showed Pam2ODN-induced increases in RelA and cJun co-expression and colocalization in vitro and in vivo within epithelial cells. At 24h, in vitro nuclear expression of RelA (+24.05% MFI) and cJun (+23.48% MFI) increased, with nuclear colocalization increasing for Rela/H3K27ac (+72.23% BDS) and RelA/cJun (+11.05% BDS). At 24h, in vivo lung epithelial populations (CD45-/CD31-/EpCAM+) had increased nuclear expression of RelA (+67.9% MFI) and increased nuclear colocalization of cJun/H3K27ac (+8.8% BDS) and RelA/cJun (+9.6% BDS). Multi-Omics integration confirmed that >70% Pam2ODN differential genes were associated with RelA and/or cJun binding sites. Conclusions: Pam2ODN enhances genomic binding of Rela and cJun to H3K27ac sites in human and mouse epithelium. RelA/cJun genomic binding correlates with epithelial gene expression changes suggesting a highly cooperative transcriptional signaling mechanism with Pam2ODN. Host-directed modulation of lung epithelial immune responses with Pam2-ODN can be beneficial against known and emerging pathogens, especially in immunocompromised patients.

[This corrects the article DOI: 10.3389/fmed.2025.1582714.].

The global incidence of respiratory infectious diseases caused by bacteria continues to increase, with acute lower respiratory tract infections contributing to significant morbidity and mortality. Preclinical models designed to investigate such respiratory bacterial diseases are of utmost importance to decipher their pathogenesis and develop novel targets for intervention and treatment. Animal models offer the powerful ability to investigate different pneumonia types at varying stages of infection and disease. However, the same models can promote important variations in outcome, potentially confounding scientific understanding in the field. Therefore, an expert panel was convened to deliberate best practices in animal models of bacterial pneumonia to identify validated methodologies and acknowledge limitations in the use of animal and non-animal models in this field of study. Herein, we summarize this American Thoracic Society workshop on animal models of bacterial pneumonia. This workshop further includes review of non-animal complementary or alternative models for studying bacterial pneumonia. Emphasis was placed on discussion of bacterial pathogens that frequently cause community- and hospital-acquired pneumonia, highlighting key aspects in modeling infection. Animal models discussed included small and large animals, based on their strengths. Finally and most importantly, the ethical considerations in the use of animal modeling for the study of bacterial lung infections was discussed. This workshop report is intended to provide insights to investigators in the field and may serve as a starting point for formal recommendations in the future.

Rationale: Transcription factor (TF) co-activation shared over multiple species often regulates conserved physiological or pathological responses. We have reported a novel host-directed aerosolized therapeutic of immunostimulatory ligands, Pam2CSK4 and ODN M362 (“Pam2+ODN”), that provides supra-additive pathogen reduction and host survival benefits against a broad range of respiratory pathogens. Here, we perform multi-Omics interrogation of anti-infectious responses in mouse and human lung epithelium models stimulated with Pam2+ODN to discover transcriptional regulatory mechanisms against respiratory pathogens. Methods: Multi-Omics responses from mouse (microarrays: GSE26864 – MLE-15; GSE289984 – Lungs) and human (HBEC3-KT: RNA-seq, RelA & cJun ChIP-seq) lung epithelial cells were analyzed to discover differential genomic-accessibility and transcriptional landscapes that lead to protection with Pam2+ODN against respiratory infections. Next-generation sequencing reads were aligned to GRCh38 for feature calling and gene/peak annotation. Differential analysis, functional gene enrichment and motif discovery were performed independently per platform using R, QIAGEN IPA, and HOMER software respectively. Biological validation of TF mechanisms related to RelA and cJun were performed in vitro (HBEC3-KT, MLE-15) and in vivo (C57BL/6J mice) after ligand exposure (PBS, Pam2, ODN, Pam2+ODN) using imaging flow cytometry (IFC), co-immunoprecipitation (Co-IP), and immunofluorescence (IF). Influenza A H3N2 (IAV) infection and viral burden was assessed by qPCR, IF or hemagglutination (HA) assays. Loss of transcriptional regulatory mechanisms was studied with single or dual RelA/cJun TF inhibitors. Results: Both mouse and human analysis of lung epithelial differential genes with Pam2+ODN revealed NF-kB and AP-1 as top transcriptional regulators, within both in vitro and in vivo microarrays/RNAseq. IFC revealed increased nuclear expression and colocalization of RelA, cJun, and H3K27Ac in isolated epithelial cells (MLE-15, HBEC3-KT) and lung epithelium populations (CD45-/CD31-/EpCam+). IF kinetic analysis (28HR) found increased RelA-H3K27Ac and cJun-H3K27Ac after Pam2+ODN stimulation of isolated human and mouse lung epithelial cells. Co-IP discarded that increases in RelA/cJun expression and colocalization resulted from formation of RelA:cJun heterodimers. Dual TF ChIPseq analysis showed that Pam2+ODN induced differentially accessible regions for RelA, with high enrichment for AP-1 motifs (including cJun). RelA and cJun also showed increased occupancy in genomic regions shared by both TFs. IF and HA confirmed that loss of single or dual RelA/cJun activity in vitro precluded Pam2-ODN-induced protection against IAV. Conclusion: We found that Pam2+ODN modulates NF-kB and AP-1 signaling in a conserved fashion across mouse and human lung epithelia. Upon infection, disruption of either RelA or cJun signaling abrogated Pam2+ODN-induced epithelial protection against viral infection.

Influenza-associated pulmonary aspergillosis (IAPA) is a potentially deadly superinfection in patients with influenza pneumonia, especially those with severe disease, underlying immunosuppression, corticosteroid therapy, or requiring intensive care support. Given the high mortality of IAPA, adjunct immunomodulatory strategies remain a critical unmet need. Previously, the desensitization of pattern recognition pathways has been described as a hallmark of IAPA pathogenesis and a predictor of mortality in IAPA patients. Therefore, we studied the impact of nebulized Toll-like receptor 2/6/9 agonists Pam2 CSK4 (Pam2) and CpG oligodeoxynucleotides (ODNs) on infection outcomes and pulmonary immunopathology in a corticosteroid-immunosuppressed murine IAPA model. Mice with IAPA receiving mock therapy showed rapidly progressing disease and a paralyzed immune response to secondary Aspergillus fumigatus infection. Nebulized Pam2ODN was well tolerated and significantly prolonged event-free survival. Specifically, dual-dose Pam2ODN therapy before and after A. fumigatus infection led to 81% survival and full recovery of all survivors. Additionally, transcriptional analysis of lung tissue homogenates revealed induction of pattern recognition receptor signaling and several key effector cytokine pathways after Pam2ODN therapy. Moreover, transcriptional and flow cytometric analyses suggested increased frequencies of macrophages, natural killer cells, and T cells in the lungs of Pam2ODN-treated mice. Collectively, immunomodulatory treatment with nebulized Pam2ODN strongly improved morbidity and mortality outcomes and alleviated paralyzed antifungal immunity in an otherwise lethal IAPA model. These findings suggest that Pam2ODN might be a promising candidate for locally delivered immunomodulatory therapy to improve outcomes of virus-associated mold infections such as IAPA.IMPORTANCEThe COVID-19 pandemic has highlighted the significant healthcare burden, morbidity, and mortality caused by secondary fungal pneumonias. Given the heightened prevalence of severe viral pneumonias, such as influenza, and poor outcomes of secondary mold pneumonias, adjunct immunotherapies are needed to prevent and treat secondary infections. We herein demonstrate severely paralyzed immunity to secondary Aspergillus fumigatus infection in a corticosteroid-immunosuppressed mouse model of influenza-associated pulmonary aspergillosis (IAPA), partially due to dysregulated pathogen-sensing pathways. To overcome immune paralysis and IAPA progression, we used a dyad of nebulized immunomodulators (Toll-like receptor agonists). Nebulized immunotherapy significantly improved morbidity and mortality compared to mock therapy, increased frequencies of mature mononuclear phagocytes and natural killer cells in the lung, and stimulated antimicrobial signaling. Collectively, this proof-of-concept study demonstrates the feasibility and efficacy of locally delivered immunomodulatory therapy to alleviate virus-induced immune dysregulation in the lung and improve outcomes of post-viral mold pneumonias such as IAPA.

Due to the high global morbidity caused by viral pneumonias, new approaches to treating and preventing these potentially fatal infections are required. Aerosolized innate immune ligands can be used to activate local antimicrobial defenses because the lung epithelium is accessible to inhaled therapies. Through the inhalation of synergistic agents Pam2CSK4, a TLR2/6 ligand, and ODN M362, a TLR9 ligand (collectively, “Pam2ODN”), we have demonstrated that therapeutic immunomodulation to stimulate lung intrinsic defenses protects mice against models of bacterial and viral infections by generating pathogen killing reactive oxygen species (ROS) from multiple lung epithelium sources, including dual oxidase 2 (DUOX2). Our lab has previously shown that lung epithelial cell treated with Pam2ODN exhibit increased expression of numerous transcription factor genes, including the antiviral transcription factor interferon regulatory factor 9 (IRF9) gene Irf9. Although Irf9 and Duox2, an antiviral IRF9-dependent gene, are both driven by Pam2ODN, it is unclear if Pam2ODN-induced Duox2 results from Pam2ODN-induced IRF9 activation. To elucidate the function of Pam2ODN-induced IRF9 in the Pam2ODN-dependent DUOX2-induced reactive oxygen species generation and pathogen killing, we employed the western blot, qPCR analysis, siRNA, and shRNA techniques in conjunction with an in vitro influenza infection model of lung epithelial cells. Here, we show that the IRF9 protein expression quickly rises after 4 hours of Pam2ODN treatment of human bronchial epithelial cells (HBEC3kt). IRF9 siRNA knockdown cells treated with Pam2ODN showed decreased DUOX2 expression compared to HBEC3kt cells transfected with scrambled siRNA, indicating that Pam2ODN-induced IRF9 drives the Pam2ODN-induced DUOX2 expression. Additionally, compared to scrambled shRNA epithelial cells treated under the same conditions, IRF9 shRNA knockdown cells treated with Pam2ODN and infected with the influenza virus had a higher viral burden. Altogether these results suggest that Pam2ODN-dependent DUOX2-induced reactive oxygen species and pathogen killing requires the induction of the transcription factor IRF9 by Pam2ODN treatment. These findings provide mechanistic insights into the function of pattern recognition receptors (PRRs) agonist-induced IRF9 in the ROS-mediated pathogen killing mediated by lung epithelial antiviral DUOX2.