Respiratory and Gut Microbiome Modification during Respiratory Syncytial Virus Infection: A Systematic Review.

BACKGROUND: Respiratory syncytial virus (RSV) infection is a major cause of lower respiratory tract infection, especially in infants, and increases the risk of recurrent wheezing and asthma. Recently, researchers have proposed a possible association between respiratory diseases and microbiome alterations. However, this connection has not been fully established. Herein, we conducted a systematic literature review to evaluate the reported evidence of microbiome alterations in patients with RSV infection. METHODS: The systematic literature review on the association between RSV and microbiome in humans was conducted by searching PubMed, EMBASE, Scopus, and CINAHL from 2012 until February 2022. The results were analyzed qualitatively, focusing on the relationship between microbiome and RSV infection with available key microbiome-related parameters. RESULTS: In the 405 articles identified by searching databases, 12 (Respiratory tract: 9, Gut: 2, Both: 1) articles in line with the research aims were eligible for this qualitative review. The types of samples for the respiratory tract microbiome and the sequencing methods utilized varied from study to study. This review revealed that the overall microbial composition in both the respiratory tract and gut in RSV-infected patients was different from that in healthy controls. Our generated results demonstrated an increase in the abundance of Haemophilus and Streptococcus, which could contribute to the distinctive separation based on the beta diversity in the respiratory tract. CONCLUSIONS: The respiratory tract and gut microbiome changed in patients with RSV infection. Further research with a well-organized longitudinal design is warranted to clarify the impact of microbiome alterations on disease pathogenesis.

Early-life pulmonary viral infection leads to long-term functional and lower airway structural changes in the lungs.

Early-life respiratory virus infections have been correlated with enhanced development of childhood asthma. In particular, significant numbers of respiratory syncytial virus (RSV)-hospitalized infants go on to develop lung disease. It has been suggested that early-life viral infections may lead to altered lung development or repair that negatively impacts lung function later in life. Our data demonstrate that early-life RSV infection modifies lung structure, leading to decreased lung function. At 5 wk postneonatal RSV infection, significant defects are observed in baseline pulmonary function test (PFT) parameters consistent with decreased lung function as well as enlarged alveolar spaces. Lung function changes in the early-life RSV-infected group continue at 3 mo of age. The altered PFT and structural changes induced by early-life RSV were mitigated in TSLPR-/- mice that have previously been shown to have reduced immune cell accumulation associated with a persistent Th2 environment. Importantly, long-term effects were demonstrated using a secondary RSV infection 3 mo following the initial early-life RSV infection and led to significant additional defects in lung function, with severe mucus deposition within the airways, and consolidation of the alveolar spaces. These studies suggest that early-life respiratory viral infection leads to alterations in lung structure/repair that predispose to diminished lung function later in life.NEW & NOTEWORTHY These studies outline a novel finding that early-life respiratory virus infection can alter lung structure and function long-term. Importantly, the data also indicate that there are critical links between inflammatory responses and subsequent events that produce a more severe pathogenic response later in life. The findings provide additional data to support that early-life infections during lung development can alter the trajectory of airway function.

Mucosal IgA immune complex induces immunomodulatory responses in allergic airway and intestinal TH2 disease.

BACKGROUND: IgA is the most abundant immunoglobulin at the mucosal surface and although its role in regulating mucosal immunity is not fully understood, its presence is associated with protection from developing allergic disease. OBJECTIVE: We sought to determine the role of IgA immune complexes for therapeutic application to mucosal allergic responses. METHODS: Trinitrophenol (TNP)-specific IgA immune complexes were applied, using TNP-coupled ovalbumin (OVA), to airway and gut mucosal surfaces in systemically sensitized allergic animals to regulate allergen challenge responses. Animals were assessed for both pathologic and immune-mediated responses in the lung and gut, respectively, using established mouse models. RESULTS: The mucosal application of IgA immune complexes in the lung and gut with TNP-OVA regulated TH2-driven allergic response in the lung and gut, reducing TH2 cytokines and mucus (lung) as well as diarrhea and temperature loss (gut), but increasing IL-10 and the number of regulatory T cells. The IgA-OVA immune complex did not alter peanut-induced anaphylaxis, indicating antigen specificity. Using OVA-specific DO.11-green fluorescent protein IL-4 reporter mouse-derived TH2-skewed cells in a transfer model demonstrated that mucosal IgA immune complex treatment reduced TH2-cell expansion and increased the number of regulatory T cells. To address a potential mechanism of action, TGF-ß and IL-10 were induced in bone marrow-derived dendritic cells when they were exposed to IgA immune complex, suggesting a regulatory phenotype induced in dendritic cells that also led to an altered primary T-cell-mediated response in in vitro OVA-specific assays. CONCLUSIONS: These studies highlight one possible mechanism of how allergen-specific IgA may provide a regulatory signal to reduce the development of allergic responses in the lung and gut.

Stem cell factor inhibition reduces Th2 inflammation and cellular infiltration in a mouse model of eosinophilic esophagitis.

Eosinophilic esophagitis (EoE) is a T helper (Th)2-mediated inflammatory disorder characterized endoscopically by eosinophilic infiltration leading to fibrosis of the esophagus. Stem cell factor (SCF), a multifunctional cytokine, is upregulated in several allergic diseases, including in patients with EoE. Mast cells and eosinophils express c-kit, the cell surface receptor for SCF, and have been found to play an important role in EoE. Therefore, we investigated whether blocking SCF represents a potential therapeutic approach for EoE. Esophageal inflammation was induced in mice using peanut allergen. In mice with experimental EoE, we found that SCF was upregulated in the esophageal tissue. In EoE mice injected with a polyclonal antibody specific for SCF, we observed a decrease in both mast cells and eosinophils by histological and flow cytometric analysis. Furthermore, Th2 cytokines in the esophagus were decreased in anti-SCF treated mice, as were levels of Th2 cytokines from lung-draining and esophageal lymph nodes. Serum levels of peanut-specific immunoglobulin E were reduced following treatment with anti-SCF. In Kitlf/f-Col1-Cre-ERT mice, which have SCF deleted primarily in myofibroblasts that develop in EoE, we observed similar results as the anti-SCF treated animals for inflammatory cell accumulation, cytokines, and histopathology. These results indicate that therapeutic treatments targeting SCF can reduce allergic inflammation in EoE.

A steroid-resistant cockroach allergen model is associated with lung and cecal microbiome changes.

The pathogenesis of asthma has been partially linked to lung and gut microbiome. We utilized a steroid-resistant chronic model of cockroach antigen-induced (CRA) asthma with corticosteroid (fluticasone) treatment to examine lung and gut microbiome during disease. The pathophysiology assessment demonstrated that mucus and airway hyperresponsiveness were increased in the chronic CRA with no alteration in the fluticasone (Flut)-treated group, demonstrating steroid resistance. Analysis of mRNA from lungs showed no decrease of MUC5AC or Gob5 in the Flut-treated group. Furthermore, flow-cytometry in lung tissue showed eosinophils and neutrophils were not significantly reduced in the Flut-treated group compared to the chronic CRA group. When the microbiome profiles were assessed, data showed that only the Flut-treated animals were significantly different in the gut microbiome. Finally, a functional analysis of cecal microbiome metabolites using PiCRUSt showed several biosynthetic pathways were significantly enriched in the Flut-treated group, with tryptophan pathway verified by ELISA with increased kynurenine in homogenized cecum samples. While the implications of these data are unclear, they may suggest a significant impact of steroid treatment on future disease pathogenesis through microbiome and associated metabolite pathway changes.

Transepidermal water loss rises before food anaphylaxis and predicts food challenge outcomes.

BACKGROUNDFood allergy (FA) is a growing health problem requiring physiologic confirmation via the oral food challenge (OFC). Many OFCs result in clinical anaphylaxis, causing discomfort and risk while limiting OFC utility. Transepidermal water loss (TEWL) measurement provides a potential solution to detect food anaphylaxis in real time prior to clinical symptoms. We evaluated whether TEWL changes during an OFC could predict anaphylaxis onset.METHODSPhysicians and nurses blinded to the TEWL results conducted and adjudicated the results of all 209 OFCs in this study. A study coordinator measured TEWL throughout the OFC and had no input on the OFC conduct. TEWL was measured 2 ways in 2 separate groups. First, TEWL was measured using static, discrete measurements. Second, TEWL was measured using continuous monitoring. Participants who consented provided blood samples before and after the OFCs for biomarker analyses.RESULTSTEWL rose significantly (2.93 g/m2/h) during reactions and did not rise during nonreacting OFCs (-1.00 g/m2/h). Systemic increases in tryptase and IL-3 were also detected during reactions, providing supporting biochemical evidence of anaphylaxis. The TEWL rise occurred 48 minutes earlier than clinically evident anaphylaxis. Continuous monitoring detected a significant rise in TEWL that presaged positive OFCs, but no rise was seen in the OFCs that resulted in no reaction, providing high predictive specificity (96%) for anaphylaxis against nonreactions 38 minutes prior to anaphylaxis onset.CONCLUSIONSDuring OFCs, a TEWL rise anticipated a positive clinical challenge. TEWL presents a monitoring modality that may predict food anaphylaxis and facilitate improvements in OFC safety and tolerability.

Sex-associated early-life viral innate immune response is transcriptionally associated with chromatin remodeling of type-I IFN-inducible genes.

This study investigates sex-associated systemic innate immune differences by examining bone marrow-derived dendritic cells (BMDCs). BMDC grown from 7-day-old mice show enhanced type-I interferon (IFN) signaling in female compared to male BMDC. Upon respiratory syncytial virus (RSV) infection of 7-day-old mice, a significantly altered phenotype of BMDC at 4 weeks post-infection is observed in a sex-dependent manner. The alterations include heightened Ifnb/ interleukin (Il12a) and enhanced IFNAR1+ expression in BMDC from early-life RSV-infected female mice that leads to increased IFN-γ production by T cells. Phenotypic differences were verified upon pulmonary sensitization whereby EL-RSV male-derived BMDC promoted enhanced T helper 2/17 responses and exacerbated disease upon RSV infection while EL-RSV/F BMDC sensitization was relatively protective. Assay for transposase-accessible chromatin using sequencing analysis (ATAC-seq) demonstrated that EL-RSV/F BMDC had enhanced chromatin accessibility near type-I immune genes with JUN, STAT1/2, and IRF1/8 transcription factors predicted to have binding sites in accessible regions. Importantly, ATAC-seq of human cord blood-derived monocytes displayed a similar sex-associated chromatin landscape with female-derived monocytes having more accessibility in type-I immune genes. These studies enhance our understanding of sex-associated differences in innate immunity by epigenetically controlled transcriptional programs amplified by early-life infection in females via type-I immunity.

Trained innate immunity, epigenetics, and food allergy.

In recent years the increased incidence of food allergy in Western culture has been associated with environmental factors and an inappropriate immune phenotype. While the adaptive immune changes in food allergy development and progression have been well-characterized, an increase in innate cell frequency and activation status has also recently received greater attention. Early in prenatal and neonatal development of human immunity there is a reliance on epigenetic and metabolic changes that stem from environmental factors, which are critical in training the immune outcomes. In the present review, we discuss how trained immunity is regulated by epigenetic, microbial and metabolic factors, and how these factors and their impact on innate immunity have been linked to the development of food allergy. We further summarize current efforts to use probiotics as a potential therapeutic approach to reverse the epigenetic and metabolic signatures and prevent the development of severe anaphylactic food allergy, as well as the potential use of trained immunity as a diagnostic and management strategy. Finally, trained immunity is presented as one of the mechanisms of action of allergen-specific immunotherapy to promote tolerogenic responses in allergic individuals.

The impact of prenatal dog keeping on infant gut microbiota development.

INTRODUCTION: Prenatal and early-life dog exposure has been linked to reduced childhood allergy and asthma. A potential mechanism includes altered early immune development in response to changes in the gut microbiome among dog-exposed infants. We thus sought to determine whether infants born into homes with indoor dog(s) exhibit altered gut microbiome development. METHODS: Pregnant women living in homes with dogs or in pet-free homes were recruited in southeast Michigan. Infant stool samples were collected at intervals between 1 week and 18 months after birth and microbiome was assessed using 16S ribosomal sequencing. Perinatal maternal vaginal/rectal swabs and stool samples were sequenced from a limited number of mothers. Mixed effect adjusted models were used to assess stool microbial community trajectories comparing infants from dog-keeping versus pet-free homes with adjustment for relevant covariates. RESULTS: Infant gut microbial composition among vaginally born babies became less similar to the maternal vaginal/rectal microbiota and more similar to the maternal gut microbiota with age-related accumulation of bacterial species with advancing age. Stool samples from dog-exposed infants were microbially more diverse (p = .041) through age 18 months with enhanced diversity most apparent between 3 and 6 months of age. Statistically significant effects of dog exposure on ß-diversity metrics were restricted to formula-fed children. Across the sample collection period, dog exposure was associated with Fusobacterium genera enrichment, as well as enrichment of Collinsella, Ruminococcus, Clostridaceae and Lachnospiraceae OTUs. CONCLUSION: Prenatal/early-life dog exposure is associated with an altered gut microbiome during infancy and supports a potential mechanism explaining lessened atopy and asthma risk. Further research directly linking specific dog-attributable changes in the infant gut microbiome to the risk of allergic disorders is needed.

Jagged-1 Reduces Th2 Inflammation and Memory Cell Expansion in Allergic Airway Disease.

Notch ligands present during interactions between T cells and dendritic cells (DCs) dictate cell phenotype through a myriad of effects including the induction of T cell regulation, survival, and cytokine response. The presence of Notch ligands on DCs varies with the context of the inflammatory response; Jagged-1 is constitutively expressed, whereas Delta-like 1 and Delta-like 4 are induced in response to pathogen exposure. Although Delta-like and Jagged ligands send different signals through the same Notch receptor, the role of these two ligands in peripheral T cell immunity is not clear. The goal of our studies was to determine the role of Jagged-1 in the pathogen-free inflammation induced by OVA during allergic airway disease in mice. Our studies show that a deletion in DC-expressed Jagged-1 causes a significant increase in cytokine production, resulting in increased mucus production and increased eosinophilia in the lungs of mice sensitized and challenged with OVA. We also observed that a reduction of Jagged-1 expression is correlated with increased expression of the Notch 1 receptor on the surface of CD4+ T cells in both the lung and lymph node. Through transfer studies using OT-II transgenic T cells, we demonstrate that Jagged-1 represses the expansion of CD44+CD62L+CCR7+ memory cells and promotes the expansion of CD44+CD62L- effector cells, but it has no effect on the expansion of naive cells during allergic airway disease. These data suggest that Jagged-1 may have different roles in Ag-specific T cell responses, depending on the maturity of the stimulated T cell.

Complete Genome Sequence of Lactobacillus johnsonii MR1, Isolated from a BALB/c Mouse Cecum.

Lactobacillus johnsonii strain MR1, which was isolated from the cecum of a BALB/cJ mouse in an airway allergy model, can decrease allergic airway inflammation in the model upon oral administration. Long-read sequencing of this isolate, which was performed using a MinION sequencer, yielded a single, closed genome of 1,953,837 bp, with a GC content of 34.67%.

Early-Life Lung and Gut Microbiota Development and Respiratory Syncytial Virus Infection.

Several environmental factors can influence the development and establishment of the early-life microbiota. For example, exposure to different environmental factors from birth to childhood will shape the lung and gut microbiota and the development of the immune system, which will impact respiratory tract infection and widespread disease occurrence during infancy and later in life. Respiratory syncytial virus (RSV) infects most infants by the age of two and is the primary cause of bronchiolitis in children worldwide. Approximately a third of infants hospitalized with bronchiolitis develop asthma later in life. However, it is unclear what factors increase susceptibility to severe RSV-bronchiolitis and the subsequent asthma development. In recent years, the role of the gut and lung microbiota in airway diseases has received increased interest, and more studies have focused on this field. Different epidemiological studies and experimental animal models have associated early-life gut microbiota dysbiosis with an increased risk of lung disease later in life. This work will review published evidence that correlated environmental factors that affect the early-life microbiota composition and their role in developing severe RSV infection.

ER stress protein PERK promotes inappropriate innate immune responses and pathogenesis during RSV infection.

The activation of dendritic cells (DC) during respiratory viral infections is central to directing the immune response and the pathologic outcome. In these studies, the effect of RSV infection on development of ER stress responses and the impact on innate immunity was examined. The upregulation of ER stress was closely associated with the PERK pathway through the upregulation of CHOP in RSV infected DC. The inhibition of PERK corresponded with decreased EIF2a phosphorylation but had no significant effect on Nrf2 in DC, two primary pathways regulated by PERK. Subsequent studies identified that by blocking PERK activity in infected DC an altered ER stress response and innate cytokine profile was observed with the upregulation of IFNß and IL-12, coincident to the down regulation of IL-1ß. When mitochondria respiration was assessed in PERK deficient DC there were increased dysfunctional mitochondria after RSV infection that resulted in reduced oxygen consumption rates (OCR) and ATP production indicating altered cellular metabolism. Use of a CD11c targeted genetic deleted murine model, RSV infection was characterized by reduced inflammation and diminished mucus staining as well as reduced mucus-associated gene gob5 expression. The assessment of the cytokine responses showed decreased IL-13 and IL-17 along with diminished IL-1ß in the lungs of PERK deficient infected mice. When PERK-deficient animals were assessed in parallel for lung leukocyte numbers, animals displayed significantly reduced myeloid and activated CD4 and CD8 T cell numbers. Thus, the PERK activation pathway may provide a rational target for altering the severe outcome of an RSV infection through modifying immune responses.

Infant gut bacterial community composition and food-related manifestation of atopy in early childhood.

BACKGROUND: Immunoglobulin E-mediated food allergy (IgE-FA) has emerged as a global public health concern. Immune dysregulation is an underlying mechanism for IgE-FA, caused by "dysbiosis" of the early intestinal microbiota. We investigated the association between infant gut bacterial composition and food-related atopy at age 3-5 years using a well-characterized birth cohort. METHODS: The study definition of IgE-FA to egg, milk, or peanut was based on physician panel retrospective review of clinical and questionnaire data collected from birth through age 3-5 years. Using 16S rRNA sequencing, we profiled the bacterial gut microbiota present in stool specimens collected at 1 and 6 months of age. RESULTS: Of 447 infants with data for analysis, 44 (9.8%) met physician panel review criteria for IgE-FA to ≥1 of the three allergens. Among children classified as IgE-FA at 3-5 years, infant stool samples showed significantly less diversity of the gut microbiota compared with the samples of children classified as no IgE-FA at age 3-5 years, especially for milk and peanut (all covariate-adjusted p's for alpha metrics <.007). Testing of individual operational taxonomic units (OTUs) revealed 6-month deficiencies in 31 OTUs for IgE-FA compared with no IgE-FA, mostly in the orders Lactobacillales, Bacteroidales, and Clostridiales. CONCLUSIONS: Variations in gut microbial composition in infant stool were associated with a study definition of IgE-FA at 3-5 years of age. This included evidence of a lack of bacterial diversity, deficiencies in specific OTUs, and delayed microbial maturation. Results support dysbiosis in IgE-FA pathogenesis.

The Lung Elastin Matrix Undergoes Rapid Degradation Upon Adult Loss of Hox5 Function.

Hox genes encode transcription factors that are critical for embryonic skeletal patterning and organogenesis. The Hoxa5, Hoxb5, and Hoxc5 paralogs are expressed in the lung mesenchyme and function redundantly during embryonic lung development. Conditional loss-of-function of these genes during postnatal stages leads to severe defects in alveologenesis, specifically in the generation of the elastin network, and animals display bronchopulmonary dysplasia (BPD) or BPD-like phenotype. Here we show the surprising results that mesenchyme-specific loss of Hox5 function at adult stages leads to rapid disruption of the mature elastin matrix, alveolar enlargement, and an emphysema-like phenotype. As the elastin matrix of the lung is considered highly stable, adult disruption of the matrix was not predicted. Just 2 weeks after deletion, adult Hox5 mutant animals show significant increases in alveolar space and changes in pulmonary function, including reduced elastance and increased compliance. Examination of the extracellular matrix (ECM) of adult Tbx4rtTA; TetOCre; Hox5a f a f bbcc lungs demonstrates a disruption of the elastin network although the underlying fibronectin, interstitial collagen and basement membrane appear unaffected. An influx of macrophages and increased matrix metalloproteinase 12 (MMP12) are observed in the distal lung 3 days after Hox5 deletion. In culture, fibroblasts from Hox5 mutant lungs exhibit reduced adhesion. These findings establish a novel role for Hox5 transcription factors as critical regulators of lung fibroblasts at adult homeostasis.

COVID-19 Modulates Inflammatory and Renal Markers That May Predict Hospital Outcomes among African American Males.

BACKGROUND AND OBJECTIVES: African Americans and males have elevated risks of infection, hospitalization, and death from SARS-CoV-2 in comparison with other populations. We report immune responses and renal injury markers in African American male patients hospitalized for COVID-19. METHODS: This was a single-center, retrospective study of 56 COVID-19 infected hospitalized African American males 50+ years of age selected from among non-intensive care unit (ICU) and ICU status patients. Demographics, hospitalization-related variables, and medical history were collected from electronic medical records. Plasma samples collected close to admission (≤2 days) were evaluated for cytokines and renal markers; results were compared to a control group (n = 31) and related to COVID-19 in-hospital mortality. RESULTS: Among COVID-19 patients, eight (14.2%) suffered in-hospital mortality; seven (23.3%) in the ICU and one (3.8%) among non-ICU patients. Interleukin (IL)-18 and IL-33 were elevated at admission in COVID-19 patients in comparison with controls. IL-6, IL-18, MCP-1/CCL2, MIP-1α/CCL3, IL-33, GST, and osteopontin were upregulated at admission in ICU patients in comparison with controls. In addition to clinical factors, MCP-1 and GST may provide incremental value for risk prediction of COVID-19 in-hospital mortality. CONCLUSIONS: Qualitatively similar inflammatory responses were observed in comparison to other populations reported in the literature, suggesting non-immunologic factors may account for outcome differences. Further, we provide initial evidence for cytokine and renal toxicity markers as prognostic factors for COVID-19 in-hospital mortality among African American males.

Maternal gut microbiome regulates immunity to RSV infection in offspring.

Development of the immune system can be influenced by diverse extrinsic and intrinsic factors that influence the risk of disease. Severe early life respiratory syncytial virus (RSV) infection is associated with persistent immune alterations. Previously, our group had shown that adult mice orally supplemented with Lactobacillus johnsonii exhibited decreased airway immunopathology following RSV infection. Here, we demonstrate that offspring of mice supplemented with L. johnsonii exhibit reduced airway mucus and Th2 cell-mediated response to RSV infection. Maternal supplementation resulted in a consistent gut microbiome in mothers and their offspring. Importantly, supplemented maternal plasma and breastmilk, and offspring plasma, exhibited decreased inflammatory metabolites. Cross-fostering studies showed that prenatal Lactobacillus exposure led to decreased Th2 cytokines and lung inflammation following RSV infection, while postnatal Lactobacillus exposure diminished goblet cell hypertrophy and mucus production in the lung in response to airway infection. These studies demonstrate that Lactobacillus modulation of the maternal microbiome and associated metabolic reprogramming enhance airway protection against RSV in neonates.

The Lung Microbiome during Health and Disease.

Healthy human lungs have traditionally been considered to be a sterile organ. However, culture-independent molecular techniques have reported that large numbers of microbes coexist in the lung and airways. The lungs harbor diverse microbial composition that are undetected by previous approaches. Many studies have found significant differences in microbial composition between during health and respiratory disease. The lung microbiome is likely to not only influence susceptibility or causes of diseases but be affected by disease activities or responses to treatment. Although lung microbiome research has some limitations from study design to reporting, it can add further dimensionality to host-microbe interactions. Moreover, there is a possibility that extending understanding to the lung microbiome with new multiple omics approaches would be useful for developing both diagnostic and prognostic biomarkers for respiratory diseases in clinical settings.