Metagenomic comparison of tracheal aspirate and mini-bronchial alveolar lavage for assessment of respiratory microbiota

Abstract

Accurate and informative microbiological testing is essential for guiding diagnosis and management of pneumonia in patients who are critically ill. Sampling of tracheal aspirate (TA) is less invasive compared with mini-bronchoalveolar lavage (mBAL) and is now recommended as a frontline diagnostic approach in patients who are mechanically ventilated, despite the historical belief that TA was suboptimal due to contamination from oral microbes. Advancements in metagenomic next-generation sequencing (mNGS) now permit assessment of airway microbiota without a need for culture and, as such, provide an opportunity to examine differences between mBAL and TA at a resolution previously unachievable. Here, we engaged shotgun mNGS to assess quantitatively the airway microbiome in matched mBAL and TA specimens from a prospective cohort of critically ill adults. We observed moderate differences between sample types across all subjects; however, we found significant compositional similarity in subjects with bacterial pneumonia, whose microbial communities were characterized by dominant pathogens. In contrast, in patients with noninfectious acute respiratory illnesses, significant differences were observed between sample types. Our findings suggest that TA sampling provides a similar assessment of airway microbiota as more invasive testing by mBAL in patients with pneumonia.

Keywords: lower respiratory tract infection; mini-bronchial alveolar lavage; next-generation sequencing; pneumonia; tracheal aspirate.

Fig. 1.

Fraction of total microbial sequencing reads represented by each genus in matched mini-bronchial alveolar lavage (left column) and tracheal aspirate (right column) specimens. Summary of the Spearman correlation ρ-values for pairwise comparisons in each PNA group is listed under each header. Legend colors correspond to the topmost abundant microbes across all samples, with red shading indicating microbes with established respiratory pathogenicity as recently defined (13), blue indicating genera known to be common oropharyngeal microbiota, and yellow/gray indicating other microbial genera. Category Other refers to all other microbes identified and those identified at abundance <1%. IQR, interquartile range.

Fig. 2.

Shannon diversity index in subjects who were pneumonia-positive (PNA-pos; △) vs. subjects who were pneumonia-negative (PNA-neg; ▲) by specimen type [mini-bronchial alveolar lavage (mBAL), left; tracheal aspirate (TA), right]. Subjects who were PNA-pos had lower diversity compared with subjects who were PNA-neg when assessed by either specimen type.