Volatile Compounds Emitted by Pseudomonas aeruginosa Stimulate Growth of the Fungal Pathogen Aspergillus fumigatus

Abstract

Chronic lung infections with opportunistic bacterial and fungal pathogens are a major cause of morbidity and mortality especially in patients with cystic fibrosis. Pseudomonas aeruginosa is the most frequently colonizing bacterium in these patients, and it is often found in association with the filamentous fungus Aspergillus fumigatus. P. aeruginosa is known to inhibit the growth of A. fumigatus in situations of direct contact, suggesting the existence of interspecies communication that may influence disease outcome. Our study shows that the lung pathogens P. aeruginosa and A. fumigatus can interact at a distance via volatile-mediated communication and expands our understanding of interspecific signaling in microbial communities.

Importance: Microbiota studies have shown that pathogens cannot be studied individually anymore and that the establishment and progression of a specific disease are due not to a single microbial species but are the result of the activity of many species living together. To date, the interaction between members of the human microbiota has been analyzed in situations of direct contact or liquid-mediated contact between organisms. This study showed unexpectedly that human opportunistic pathogens can interact at a distance after sensing volatiles emitted by another microbial species. This finding will open a new research avenue for the understanding of microbial communities.

FIG 1

Growth characteristics of A. fumigatus in a plate-in-plate (PIP) coculture assay. (A) The A. fumigatus (Af) fungus is isolated from a separate compartment containing either medium alone (top) or medium inoculated with P. aeruginosa (Pa) (bottom). The presence of P. aeruginosa increases mycelial growth, particularly in the vicinity of the bacterium. The medium used was Brian medium (BM). (B) The growth-promoting effects of P. aeruginosa on A. fumigatus were also evident on minimal medium (MM), although asymmetric growth of the colony was not observed. (C) Quantitation of the surface area of A. fumigatus colonies growing on MM or BM in sealed PIP cultures in the presence and absence of P. aeruginosa. Values are averages plus standard errors of the means (SEM) (error bars) of at least three replicates. Values that are significantly different (P ≤ 0.01) are indicated by a bar and two asterisks.

FIG 2

Effects of VOCs on the growth of A. fumigatus. (A) Quantitation of A. fumigatus (Af) biomass in the presence of 100 ppm of pure VOCs identified in the P. aeruginosa culture headspace. Only DMS significantly increased growth, and only 2-nonanone had a significant inhibitory effect. (B) Comparison of the growth-promoting effects of 1 ppm DMS (DMS1) and 10 ppm DMS (DMS10) on sulfur-replete MM and on sulfur-deficient MM (MM-S). CTRL, control. (C) Residual concentration of DMS in the culture headspace of the double-plate setup. DMS produced by P. aeruginosa (grown on MM) or the pure compound (1 and 10 ppm) is partially consumed by the A. fumigatus fungus when grown on MM (Af) but is fully consumed on sulfur-deficient MM [Af(-S)]. (D) Agar plates showing the growth stimulation of A. fumigatus by DMS. As shown in panel B, the effect is increased when the fungus is grown under sulfur starvation. All error bars show the SEM of the mean values. Values that are significantly different are indicated as follows: *, P ≤ 0.05; †, P ≤ 0.05; **, P ≤ 0.01.