Lung Mycobiota α-Diversity Is Linked to Severity in Critically Ill Patients with Acute Exacerbation of Chronic Obstructive Pulmonary Disease

ABSTRACT Chronic obstructive pulmonary disease (COPD) affects more than 200 million people worldwide. The chronic course of COPD is frequently worsened by acute exacerbations (AECOPD). Mortality in patients hospitalized for severe AECOPD remains dramatically high, and the underlying mechanisms are poorly understood. Lung microbiota is associated with COPD outcomes in nonsevere AECOPD, but no study specifically investigated severe AECOPD patients. The aim of this study is thus to compare lung microbiota composition between severe AECOPD survivors and nonsurvivors. Induced sputum or endotracheal aspirate was collected at admission from every consecutive severe AECOPD patient. After DNA extraction, the V3-V4 and ITS2 regions were amplified by PCR. Deep-sequencing was performed on a MiSeq sequencer (Illumina); the data were analyzed using DADA2 pipeline. Among 47 patients admitted for severe AECOPD, 25 (53%) with samples of sufficient quality were included: 21 of 25 (84%) survivors and 4 of 25 (16%) nonsurvivors. AECOPD nonsurvivors had lower α-diversities indices than survivors for lung mycobiota but not for lung bacteriobiota. Similar results were demonstrated comparing patients receiving invasive mechanical ventilation (n = 13 [52%]) with those receiving only noninvasive ventilation (n = 12 [48%]). Previous systemic antimicrobial therapy and long-term inhaled corticosteroid therapy could alter the lung microbiota composition in severe AECOPD patients. In acidemic AECOPD, lower lung mycobiota α-diversity is linked to the severity of the exacerbation, assessed by mortality and the requirement for invasive mechanical ventilation, whereas lung bacteriobiota α-diversity is not. This study encourages a multicenter cohort study investigating the role of lung microbiota, especially fungal kingdom, in severe AECOPD. IMPORTANCE In AECOPD with acidemia, more severe patients—i.e., nonsurvivors and patients requiring invasive mechanical ventilation—have lower lung mycobiota α-diversity than survivors and patients receiving only noninvasive ventilation, respectively. This study encourages a large multicenter cohort study investigating the role of lung microbiota in severe AECOPD and urges investigation of the role of the fungal kingdom in severe AECOPD.

Supplemental Figure 3. Rarefaction curves for ITS2 sequencing TN: negative control from the extraction step. TN PCR: negative control from the amplification PCR step.
Supplemental Figure 4. Comparison of lung mycobiota between severe AECOPD patients who received antimicrobial therapy within the previous month and those who did not. A. Boxplot of estimated α-diversity by Shannon index. B. Boxplot of estimated α-diversity by Simpson index. C. Boxplot of estimated α-diversity by evenness. D. Non-metric Bray-curtis analysis of β-diversity. Threshold for statistical significance: p=0.05. atb: systemic antibiotics within the previous month.
Supplemental Figure 5. Comparison of lung bacteriobiota between severe AECOPD patients who received antimicrobial therapy within the previous month and those who did not. A. Boxplot of estimated α-diversity by Shannon index. B. Boxplot of estimated α-diversity by Simpson index. C. Boxplot of estimated α-diversity by evenness. D. Non-metric Bray-curtis analysis of β-diversity. Threshold for statistical significance: p=0.05. atb: systemic antibiotics within the previous month.
Supplemental Figure 6. Comparison of lung mycobiota between severe AECOPD patients who receive long-term inhaled corticosteroids therapy and those who do not. A. Boxplot of estimated α-diversity by Shannon index. B. Boxplot of estimated α-diversity by Simpson index. C. Boxplot of estimated α-diversity by evenness. D. Non-metric Bray-curtis analysis of βdiversity. Threshold for statistical significance: p=0.05. icortic: long-term inhaled corticosteroids therapy Supplemental Figure 7. Comparison of lung bacteriobiota between severe AECOPD patients who receive long-term inhaled corticosteroids therapy and those who do not. A. Boxplot of estimated α-diversity by Shannon index. B. Boxplot of estimated α-diversity by Simpson index. C. Boxplot of estimated α-diversity by evenness. D. Non-metric Bray-curtis analysis of βdiversity. Threshold for statistical significance: p=0.05. icortic: long-term inhaled corticosteroids therapy Supplemental Figure 8. Comparison of lung mycobiota between severe AECOPD patients who received systemic corticosteroids therapy within the previous month and those who did not. A. Boxplot of estimated α-diversity by Shannon index. B. Boxplot of estimated α-diversity by Simpson index. C. Boxplot of estimated α-diversity by evenness. D. Non-metric Bray-curtis analysis of β-diversity. Threshold for statistical significance: p=0.05. corticotherapy: corticosteroids therapy within the previous month.
Supplemental Figure 9. Comparison of lung bacteriobiota between severe AECOPD patients who received systemic corticosteroids therapy within the previous month and those who did not. A. Boxplot of estimated α-diversity by Shannon index. B. Boxplot of estimated α-diversity by Simpson index. C. Boxplot of estimated α-diversity by evenness. D. Non-metric Bray-curtis analysis of β-diversity. Threshold for statistical significance: p=0.05. corticotherapy: corticosteroids therapy within the previous month.