The Respiratory Microbiome in Chronic Hypersensitivity Pneumonitis Is Distinct from That of Idiopathic Pulmonary Fibrosis

Abstract

Rationale: Chronic hypersensitivity pneumonitis (CHP) is a condition that arises after repeated exposure and sensitization to inhaled antigens. The lung microbiome is increasingly implicated in respiratory disease, but, to date, no study has investigated the composition of microbial communities in the lower airways in CHP.Objectives: To characterize and compare the airway microbiome in subjects with CHP, subjects with idiopathic pulmonary fibrosis (IPF), and control subjects.Methods: We prospectively recruited individuals with a CHP diagnosis (n = 110), individuals with an IPF diagnosis (n = 45), and control subjects (n = 28). Subjects underwent BAL and bacterial DNA was isolated, quantified by quantitative PCR and the 16S ribosomal RNA gene was sequenced to characterize the bacterial communities in the lower airways.Measurements and Main Results: Distinct differences in the microbial profiles were evident in the lower airways of subjects with CHP and IPF. At the phylum level, the prevailing microbiota of both subjects with IPF and subjects with CHP included Firmicutes, Bacteroidetes, Proteobacteria, and Actinobacteria. However, in IPF, Firmicutes dominated, whereas the percentage of reads assigned to Proteobacteria in the same group was significantly lower than the percentage found in subjects with CHP. At the genus level, the Staphylococcus burden was increased in CHP, and Actinomyces and Veillonella burdens were increased in IPF. The lower airway bacterial burden in subjects with CHP was higher than that in control subjects but lower than that of those with IPF. In contrast to IPF, there was no association between bacterial burden and survival in CHP.Conclusions: The microbial profile of the lower airways in subjects with CHP is distinct from that of IPF, and, notably, the bacterial burden in individuals with CHP fails to predict survival.

Keywords: 16S; fibrosis; lung microbiota.

Figure 1.

Bacterial burden in BAL fluid of control subjects, subjects with CHP, and subjects with IPF. The Kruskal-Wallis test with the Dunn test for multiple comparisons was used (P < 2.2 × 10⁻¹⁶). Data are presented as the median and interquartile range (***P < 0.0001 and ****P < 0.00001). The bacterial burden was calculated by using quantitative PCR and was expressed in log₁₀ values (16S ribosomal RNA gene copies/ml of BAL fluid). CHP = chronic hypersensitivity pneumonitis; IPF = idiopathic pulmonary fibrosis.

Figure 2.

Survival probability of subjects with chronic hypersensitivity pneumonitis (CHP) compared with individuals with idiopathic pulmonary fibrosis (IPF). A Kaplan-Meier curve generated by using a Cox proportional hazards model displaying the survival probability (percentage) of subjects with CHP compared with subjects with IPF is shown. Log-rank P test values are reported.

Figure 3.

Taxonomic composition of bacteria in BAL of healthy control subjects and subjects with chronic hypersensitivity pneumonitis (CHP) and idiopathic pulmonary fibrosis (IPF). (A) Relative abundance of bacteria at the phylum (Bacteroidetes, Firmicutes, Proteobacteria, Actinobacteria, and Fusobacteria) and genus (Prevotella, Streptococcus, Veillonella, Neisseria, Haemophilus, Actinomyces, Rothia, Fusobacterium, Actinobacillus, and Staphylococcus) level. Statistical significance was tested by using the Kruskal-Wallis test with the Dunn multiple-comparison test adjusted with the Bonferroni method. Statistical differences between CHP and IPF are indicated by black asterisks, and those between control subjects and IPF are indicated by red asterisks (*P < 0.05, **P < 0.01, and ***P < 0.001). Data are presented as the mean ± SD. (B) Shannon and Chao1 α-diversity measures comparing healthy control subjects and individuals with CHP and IPF diagnoses. Statistical significance was tested by using the Kruskal-Wallis test with the Dunn multiple-comparison test adjusted with the Bonferroni method. Data are presented as the median and interquartile range. (C) PC analysis on Euclidean distance comparing healthy control subjects, subjects with CHP, and subjects with IPF (permutational multivariate ANOVA [PERMANOVA], P < 0.001). PERMANOVA was adjusted for age, sex, smoking status, FVC (% predicted), and Dl_CO (% predicted). PC = principal component.