Patterns in the longitudinal oropharyngeal microbiome evolution related to ventilator-associated pneumonia

Abstract

Background: The aim of the study was to evaluate the composition and the temporal evolution of the oropharyngeal microbiome in antibiotic-naïve patients requiring mechanical ventilation and to gain new insights into the pathogenesis of ventilator-associated pneumonia (VAP).

Methods: Prospective, observational single-center nested case-control study. Patients with acute critical illness and anticipated duration of mechanical ventilation > 4 days were eligible. We took oropharyngeal swabs (and if available, tracheal secretions) daily, starting at the day of intubation. The microbiota was characterized by 16S rRNA high-throughput sequencing and compared between patients developing VAP versus controls.

Results: Five patients developed VAP. In three patient the causative pathogens were Enterobacteriaceae and in two Haemophilus influenzae. Locally weighted polynomial regression suggested that the within diversity (=alpha) was lower in Enterobacteriaceae VAP patients between days two to five of mechanical ventilation when compared to controls. Detection of Enterobacteriaceae in the oropharynx occurred on day two of follow-up and consisted of a single operational taxonomic unit in 2/3 patients with enterobacterial VAP.

Conclusions: In acutely-ill patients who developed enterobacterial VAP the causative pathogen gained access to the oropharynx early after starting mechanical ventilation and outgrew the commensal members of the microbiome. Whether a specific pattern of the oropharyngeal microbiome between days three to five of mechanical ventilation may predict VAP enterobacterial VAP has to be evaluated in further studies.

Keywords: Infection prevention; Intensive care; Nosocomial pneumonia; Oropharyngeal and tracheal microbiome; Ventilator-associated pneumonia.

Conflict of interest statement

Competing interestsThe authors declare that they have no competing interests.

Figures

Fig. 1

Flowchart. The process of subject screening, inclusion, and selection for microbiome evaluation is shown in the flow chart. Subjects were selected to fulfill possible VAP criteria according to clinical criteria (new infiltrate plus two of the following three: temperature greater than 38 °C, leukocyte count greater than 12 G/L or lower than 4 G/L, or purulent respiratory secretions)

Fig. 2

Timeline. For each study patient who underwent microbiome evaluation, a timeline indicates the moment of intubation, study start, endpoint (VAP or extubation), and if performed tracheostomy. Intermittent extubations and reintubations

Fig. 3

Heatmap of oropharyngeal and tracheobronchial samples. Relative abundances of bacterial communities of the subjects are shown. The left main column are oropharyngeal samples, the middle tracheobronchial and the right shows the relative dissimilarity between the two corresponding samples. Each subcolumn for oropharyngeal (oro) and tracheobronchial (tbs) samples represents family-level taxonomic assignment of the sample’s 16S rRNA gene sequences. The color indicates proportional abundance of the sequences assigned to each bacterial family within the sample. The rows represent individual time points of sampling for the subjects. IO and IT represent the initial oropharyngeal and tracheobronchial samples, respectively. Missing samples are indicated with grey fill

Fig. 4

Early access of VAP causative Enterobacteriaceae to the oropharynx. Relative abundance of causative (red) and any other (green) Enterobacteriaceae operational taxonomic units is shown for ten individual study subjects. Results of paired oropharyngeal (upper panels) and tracheobronchial (lower panels) samples are shown. Enterobacteriaceae VAP patients are subjects 3, 16 & 20. The x-axis denotes days post intubation

Fig. 5

Decrease of oropharyngeal longitudinal alpha diversity in enterobacterial VAP. Longitudinal changes in alpha diversity of the oropharyngeal samples of the individual patients (dots) and by VAP diagnosis. Lines are based on a local polynomial regression fitting (loess function in R, grey bands indicate 95% CI). a Richness, b Shannon diversity index. Note: Summary lines for H. influenzae VAP patients were omitted due to low number of cases (n = 2)