Microbiome in Asthma and COPD

Introduction

The microbiome, defined as the total collection of microbiota (microorganisms specific to an anatomic site, such as lung or skin) that inhabits the human body, consists of microorganisms such as bacteria, viruses, and fungi.¹ There is a long-held belief that the normal human lung is sterile, but an increasing body of evidence suggests this is not the case.² Composition of the lung microbiome (defined as the complete collection of microbiota residing in the airways and parenchymal tissues) is a central contributor to obstructive lung disease.^1,2 Although much remains to be learned about the specific pathogenic role of microbiota in the diseased lung, recent findings have linked changes in lung bacterial communities to disease progression and exacerbation in patients with either chronic obstructive pulmonary disease (COPD) or asthma.¹ In addition, some studies have shown differences in the bronchial microbiome between patients with asthma, patients with COPD, and healthy individuals, suggesting an association between microbial communities and bronchial health.³ The aim of this newsletter is to provide an introduction to recent advances in the understanding of the impact of microorganisms on the normal and diseased lung for patients with asthma and COPD.

Methodologies for Studying the Microbial Environment of the Lungs

It has been suggested that current culture techniques are unable to grow an estimated 60% to 90% of bacterial species that inhabit bodily surfaces due to lack of knowledge regarding the exact conditions needed for growth (eg, osmotic pressure, temperature, nutrients).^4-6 Additionally, lung tissue samples are not easy to obtain from the upper airways without contamination.⁷ However, a study of lung tissues removed during lung volume reduction surgery confirmed that the lungs are not a sterile environment.⁸ Technological advances, especially those that use polymerase chain reaction (PCR) amplification of the 16S ribosomal RNA (rRNA) sequences for identification of bacteria, have allowed us to understand the vast number of microbes present in the lung. This technique has been crucial in identifying specific bacteria and determining bacterial populations in the lungs of patients with asthma or COPD²; its use in identifying viruses and fungi is also of great importance.⁹ Most analyses are carried out on bronchoalveolar lavage (BAL) fluid using culture-independent techniques.⁹ Here we will briefly describe some of the seminal studies evaluating the role of the microbiome in asthma and COPD.

In a landmark study showing that the lungs are not sterile and that various bacterial species are found in the lower airways, Hilty and colleagues used molecular analysis of the bacterial 16S rRNA gene to compare the bacterial communities in the airways of adult patients with asthma (n=11) or COPD (n=5) with those of healthy individuals (n=8); results were replicated in children with asthma attending clinics for therapy resistance.⁵ Strong similarities were observed in airway microbiota between adult patients with asthma and children with asthma, including the potential pathogens Haemophilus, Moraxella, and Neisseria spp.⁵ The investigators also found important differences in the airway microbial flora between patients with asthma or COPD and control subjects.⁵ Bacteria from the phylum Proteobacteria, particularly the potential pathogen Haemophilus, were more common in patients with asthma or COPD than in controls.⁵ Notably, prevalence of asthma at >5 years was higher in children who as neonates had colonization of the hypopharyngeal region by Haemophilus and other species of Proteobacteria than in children who were not colonized (33% vs 10%, respectively; odds ratio: 4.6 [95% CI, 2.2-9.6]).¹⁰

In an important follow-up study, Erb-Downward et al used pyrosequencing of 16S amplicons to analyze the lung microbiome using BAL samples from healthy nonsmokers (n=3), healthy smokers with normal spirometry (n=7), and COPD patients (n=4 [2 mild, 1 moderate, 1 severe]), and explant samples from 6 patients undergoing lung transplantation for advanced COPD.¹¹ Patients with moderate and severe COPD were found to have limited bacterial community diversity, whereas healthy subjects and patients with mild COPD generally had more heterogeneous bacterial communities (ie, multiple phyla and genera represented).¹¹ The predominant bacterial species found in the specimens of the patients with advanced COPD in this study were Pseudomonas and Haemophilus spp.¹¹ Interestingly, lung explants from patients with advanced COPD also had significant microanatomic differences in bacterial communities within the same lung.¹¹ The results of this study highlight the potential for a single site containing a pathogenic bacterial community to interact with host immunity, providing a mechanism for localized disease progression.¹¹

Other reports have shown distinct but related microbiota between the upper and lower respiratory tract.^9,11-14 Park and colleagues used pyrosequencing of 16S rRNA genes to characterize the bacterial microbiota in the upper airways within the oropharynx in patients with asthma (n=18) or COPD (n=17) compared with healthy controls (n=12).¹⁵ Diverse microbiota were observed in the upper respiratory tract of patients with asthma and COPD, including abundant levels of both Pseudomonas spp and Lactobacillus spp, both of which were observed in low frequencies in healthy controls.¹⁵ No significant differences in microbiota were observed between patients with asthma or COPD.¹⁵ In contrast, Streptococcus spp, Neisseria spp, Prevotella spp, and Veillonella spp were dominant in the oropharynx of healthy controls.¹⁵ These studies highlight the importance of the culture-independent molecular approach, but they also show that additional studies are required to fully understand the bacterial colonization of the lungs in patients with asthma or COPD.

Interactions Between Microbiome and Mucosal Immunity

The best-studied mucosal microbiome for humans is in the gut, which currently informs our thinking about what occurs in the lung.² In the gut, interactions between the microbiome and mucosal immunity are established and have an essential role in shaping host immunity.² First, the microorganisms control the activation state of gut dendritic cells and regulate the balance of T-helper type 17 cells and T-regulatory cells for pathogenic infections.² Second, the microbiome of the gut has an impact on the development of inflammatory bowel diseases, specifically through gut inflammation that is caused by abnormal host response to bacterial antigens.² Although studies have not shown a direct correlation between the lung microbiome and obstructive lung diseases, studies in patients with asthma or COPD have shown results similar to those observed in the gut.^2,5,11 One mouse study showed that altering gut microflora by administering oral antibiotics decreased CD4⁺ and CD8⁺ T cells as well as antibody immunity in the lungs (demonstrated by intranasal challenge with influenza virus).¹⁶ In addition, chronic azithromycin therapy has shown benefits for both asthma and COPD, and it has been proposed that the antibacterial, anti-inflammatory, and immunomodulatory effects of azithromycin may be due to alterations it produces in the microbiome of the lungs.²

One model that emerged from the studies discussed above is the “common mucosal immune system,” which states that there is crosstalk among mucosal compartments and that mucosal immune function may be shared among different sites, which can alter the response to infection.³ Additional research is required to confirm the mechanisms by which the microbiome affects lung immunity, but its potential role in asthma and COPD is very intriguing and is described further below.

Microbiome in Asthma

The connection between asthma and microbial colonization is not thoroughly understood, but recent research is attempting to explain the role of the microbiome in asthma. One explanation—discussed for years but still incompletely understood—is the hygiene hypothesis, which states that decreased infectious exposures early in life can lead to defective mucosal tolerance and increased autoimmune pathology (Figure 1).^2,9,17,18 This hypothesis could account for the association between decreasing frequency of childhood infections and increasing development of asthma.⁹ Also in support of this theory, exposure to specific environmental stimuli, especially pet dogs and farm animals, appears to protect against asthma.³ It has been postulated that the protection may be due to an alteration of the gut and lung microbiomes during early childhood when the adult microbiome is developing.³

Figure 1. Interactions between genetic background and exposure to environment   determine mucosal response — resident microbiome status2,9,17,18

Adapted with permission of the American Thoracic Society Copyright © 2016 American Thoracic Society. Martinez FD. The human microbiome. Early life determinant of health outcomes. Ann Am Thorac Soc. 2014;11(suppl 1):S7-S12. The Annals of the American Thoracic Society is an official journal of the American Thoracic Society.

Authors of a retrospective, population-based, case-control study of 174 cases of serious pneumococcal disease (SPD) described an association between SPD and a prior history of asthma in adults, suggesting that asthma may increase the risk for SPD.¹⁹ Asthma has also been associated with increased risk of non–airway-related infections. A retrospective, population-based, case-control study of 518 patients found that patients with active or current asthma were at higher risk of community-acquired Escherichia coli bloodstream infection.²⁰

Changes in the airway microbiome have also been linked to treatment response to inhaled corticosteroids. A prospective study comparing 16S rRNA from BAL samples found alterations in the lung microbiome in patients with asthma (n=39) compared with healthy controls (n=12).²¹ At the phylum level, there were no differences between patients with corticosteroid-resistant asthma and those who were corticosteroid-sensitive, but their BAL microbiomes differed at the genus level, suggesting a mechanism for influencing treatment responses in asthma patients.²¹ For example, BAL macrophages from corticosteroid-resistant patients showed an enhanced inflammatory response to Haemophilus parainfluenzae but not to Prevotella melaninogenica.²¹ In addition, other studies have reported that patients with asthma have a greater abundance of members of the phylum Proteobacteria, specifically Haemophilus, in their airways.^5,22 While the specific mechanisms by which the microbiome alters asthma development remain unclear, evidence suggests that the lung microbiome is altered in patients with asthma.³

Microbiome in COPD

Current research suggests that COPD is caused in part by inflammation resulting from chronic and recurrent airway infections. Furthermore, evidence suggests that the lung microbiome is less diverse in patients with COPD, which may contribute to disease progression.^8,23,24 In a study evaluating DNA from lung tissue samples using 16S rRNA PCR amplification, patients with very severe COPD (Global Initiative for COPD [GOLD] 4; n=8) had a different bacterial community than nonsmokers (n=8) and smokers without COPD (n=8). In particular, a significant association with the Lactobacillus genus was observed in patients with very severe COPD.²⁴

In addition, persistent and recurrent infections can alter the microbiome of the lung and cause inflammation.² Findings from a 6-month study in 43 patients with moderate to very severe COPD who had >1 exacerbations requiring antibiotics in the previous year showed that 10 patients were chronically colonized with Haemophilus influenza, and these patients showed increased airway inflammation (specifically neutrophil activity) and reduced lung volumes during stable phase compared with patients who were not chronically colonized.²⁵ One proposed hypothesis suggests that persistent infection with pathogenic microorganisms coupled with ongoing inflammation may push patients with COPD toward more severe disease progression.¹ More research is required to test this hypothesis and to better understand the role of bacterial microbiota in the lung.

Microbiome and COPD Exacerbations

Exacerbations are a major source of morbidity and mortality for patients with COPD, and it is thought that these exacerbations are triggered by an acute bacterial or viral infection (Figure 2).^1,27 Consistent with this idea, antibiotic treatment for 7 to 10 days during an acute exacerbation of COPD has been shown to reduce the risk of treatment failure and mortality, suggesting the importance of microbial infection during exacerbation.²⁸ It has also been hypothesized that patients with COPD persistently contain pathogenic bacteria in the lung and that when there is an overgrowth beyond a particular (unknown) threshold, an exacerbation occurs.¹ In support of this hypothesis, a randomized trial of 1142 COPD patients treated with either 250 mg azithromycin daily or placebo for 1 year in addition to usual care found that azithromycin treatment decreased exacerbation frequency by 27% compared with placebo (1.48 vs 1.83 exacerbations per patient-year, respectively; hazard ratio: 0.73 [95% CI, 0.6-0.8]; P<.001).²⁶ In addition, azithromycin treatment improved quality of life in 43% of patients vs 36% of placebo-treated patients (clinically meaningful improvement in the St. George’s Respiratory Questionnaire; P = .03).²⁶

Recent research also suggests that the lung microbiome changes during exacerbations. A longitudinal study that evaluated sputum samples in 12 patients with COPD found a significant shift in the microbiome taxa during and after treatment of an exacerbation.²⁹ Interestingly, distinct shifts in taxa abundance were observed depending on the type of treatment (antibiotics only, oral corticosteroids only, or antibiotics and corticosteroids), mostly within the Proteobacteria phylum.²⁹ Although not yet fully defined, current data suggest an important role of lung microbiota in COPD development and disease progression.

Figure 2. Potential role of the bacterial microbiome in exacerbations and the   pathogenesis of COPD1,27

Conclusions

Although the lung was once thought to be sterile, there is clear evidence that microbial communities are found in the lung in healthy patients. Recent advances in culture-independent technology, particularly using PCR amplification of 16S rRNA sequences, allow identification of species of microbiota present in the lung of healthy individuals and patients with asthma and COPD. It is believed that the microbiome affects immunity in the lungs with a mechanism similar to that in the gut and likely contributes to the development and pathogenesis of asthma and COPD. Research has just begun to elucidate the role of the microbiome in asthma, but the species and diversity of microbes in the lung are clearly altered in patients with asthma. In COPD, it is believed that changes in the lung microbiome lead to chronic inflammation and recurrent airway infection, which subsequently leads to an increased likelihood of exacerbations and disease progression. So far, the most convincing evidence of the importance of the lung microbiome in obstructive airways disease is that chronic azithromycin therapy, which has antibacterial, anti-inflammatory, and immunomodulatory effects, has shown benefits for both asthma and COPD patients.² Although much remains to be learned about the microbiome in the healthy and diseased lung, it will undoubtedly affect how we treat our patients in the near future.

References

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