Polymicrobial airway bacterial communities in adult bronchiectasis patients

Abstract

Background: Chronic airway infection contributes to the underlying pathogenesis of non-cystic fibrosis bronchiectasis (NCFBr). In contrast to other chronic airway infections, associated with COPD and CF bronchiectasis, where polymicrobial communities have been implicated in lung damage due to the vicious circle of recurrent bacterial infections and inflammation, there is sparse information on the composition of bacterial communities in NCFBr. Seventy consecutive patients were recruited from an outpatient adult NCFBr clinic. Bacterial communities in sputum samples were analysed by culture and pyrosequencing approaches. Bacterial sequences were analysed using partial least square discrimination analyses to investigate trends in community composition and identify those taxa that contribute most to community variation.

Results: The lower airway in NCFBr is dominated by three bacterial taxa Pasteurellaceae, Streptococcaceae and Pseudomonadaceae. Moreover, the bacterial community is much more diverse than indicated by culture and contains significant numbers of other genera including anaerobic Prevotellaceae, Veillonellaceae and Actinomycetaceae. We found particular taxa are correlated with different clinical states, 27 taxa were associated with acute exacerbations, whereas 11 taxa correlated with stable clinical states. We were unable to demonstrate a significant effect of antibiotic therapy, gender, or lung function on the diversity of the bacterial community. However, presence of clinically significant culturable taxa; particularly Pseudomonas aeruginosa and Haemophilus influenzae correlated with a significant change in the diversity of the bacterial community in the lung.

Conclusions: We have demonstrated that acute exacerbations, the frequency of exacerbation and episodes of clinical stability are correlated, in some patients, with a significantly different bacterial community structure, that are associated with a presence of particular taxa in the NCFBr lung. Moreover, there appears to be an inverse relationship between the abundance of P. aeruginosa and that of of H. influenzae within the NCFBr lung bacterial community. This interaction requires further exploration.

Figure 1

Canonical correspondence analysis of (A) total cohort showing that sputum samples that were culture positive (P = 0.016); had culturable P. aeruginosa (P = 0.002) and culturable H. influenzae (P = 0.002) were associated with distinct bacterial community assemblies, (B) Frequent exacerbators also showing that samples that were culture positive (P = 0.05); had culturable P. aeruginosa (P = 0.002) and culturable H. influenzae (P = 0.002) were associated with distinct bacterial community assemblies. Discrete variables, indicated by ▲, are; Culture positive sputum; H.i, H. influenzae detected by culture and P.a, P. aeruginosa detected by culture. Other variables analysed; the presence of an exacerbation at time of sampling; 12 month history of persistent; intermittent or absence of culturable P. aeruginosa, current azithromycin treatment; current nebulised colistin treatment; gender, FEV1% predicted; frequent exacerbation and age. None were found to significantly affect the community structure and for clarity are not shown. Percentage values show variance within data explained by that axis.

Figure 2

Partial least squares discriminant analysis (PLS-DA) loading plot based on the relative abundance of bacterial taxa determined by 454 sequence analysis of the microbiota of sputum from patients reporting current stability (green circle) and sputum from patients reporting a current exacerbation (blue circle). PLS1 (R²X = 0.169, R²Y = 0.232, Q² = 0.0287) and PLS 2 (R²X = 0.107, R²Y = 0.124, Q² = 0.0601) are given. The solid ellipse indicates Hotellings T² range, at 95% confidence. Patient samples derived from current exacerbators contained within the dashed ellipse, and including BX6 are deemed to be the major outliers, having a microbial community composition which is dissimilar to the stable and a small proportion of exacerbating patients. Some sample labels have been removed for ease of interpretation.

Figure 3

Partial least squares discriminant analysis (PLS-DA) loading plot showing the contributing microbial community members towards the separation of the PLS-DA scores between patients reporting current stability (▲) and sputum from patients reporting a current exacerbation (▼). PLS1 (R2X = 0.169, R2Y = 0.232, Q2 = 0.0287) and PLS 2 (R2X = 0.107, R2Y = 0.124, Q2 = 0.0601) are given. Taxa deemed clinically relevant (based on those screened during standard culture) are highlighted in blue. Some sample labels have been removed for ease of interpretation.

Figure 4

Partial least squares discriminant analysis (PLS-DA) loading plot based on the relative abundance of bacterial taxa determined by 454 sequence analysis of the microbiota of sputum from patients with greater than 3 exacerbation event per annum (green circle) and sputum from patients with 3 or fewer exacerbation events per annum (blue circle). PLS1 (R²X = 0.0701, R²Y = 0.232, Q² = 0.0467) and PLS 2 (R²X = 0.0477, R²Y = 0.124, Q² = 0.0601) are given. The solid ellipse indicates Hotellings T² range, at 95% confidence. Patient samples falling outside of the ellipse are deemed to be the major outliers. Some sample labels have been removed for ease of interpretation.

Figure 5

Partial least squares discriminant analysis (PLS-DA) loading plot showing the contributing microbial community members towards the separation of the PLS-DA scores between patients are frequent exacerbators (>3 exacerbation events per annum) and sputum from patients who are stable (≤3 exacerbation events per annum). Taxa deemed clinically relevant (based on those screened during standard culture) are highlighted in blue. Some sample labels have been removed for ease of interpretation.