Interactions between Aspergillus fumigatus and Pulmonary Bacteria: Current State of the Field, New Data, and Future Perspective

Abstract

Aspergillus fumigatus and Pseudomonas aeruginosa are central fungal and bacterial members of the pulmonary microbiota. The interactions between A. fumigatus and P. aeruginosa have only just begun to be explored. A balance between inhibitory and stimulatory effects on fungal growth was observed in mixed A. fumigatus-P. aeruginosa cultures. Negative interactions have been seen for homoserine-lactones, pyoverdine and pyochelin resulting from iron starvation and intracellular inhibitory reactive oxidant production. In contrast, several types of positive interactions were recognized. Dirhamnolipids resulted in the production of a thick fungal cell wall, allowing the fungus to resist stress. Phenazines and pyochelin favor iron uptake for the fungus. A. fumigatus is able to use bacterial volatiles to promote its growth. The immune response is also differentially regulated by co-infections.

Keywords: Aspergillus; Pseudomonas; cell wall; cystic fibrosis; interaction; microbiota; phenazine; pyochelin; rhamnolipid; volatile.

Figure 1

Diagram illustrating the mode of action of P. aeruginosa dirhamnolipids on A. fumigatus growth. Dirhamnolipids inhibit β1,3 glucan synthase (GS) at the hyphal tip (1). This inhibition stimulates the formation of new apices (2), containing active GS which will be further inhibited by the dirhamnolipids, giving the multibranched phenotype with short apical cells. The inhibition of the β1,3 glucan synthesis is compensated by an increase in chitin synthesis (3). Dirhamnolipids also induce melanin and galactosaminogalactan production in the extracellular matrix (4).

Figure 2

Pyochelin (PCH) antifungal activity on A. fumigatus. Growth was measured by the absorbance of the crystal violet-binding hyphae at 560 nm. (a) In minimal medium (MM); (b) In MM depleted in iron (-Fe), Zinc (-Zn) or copper (-Cu); (c) In MM depleted in iron, zinc and copper (-Fe,-Zn,-Cu). Medium composition and methodology are described in Supplementary Materials.

Figure 3

A. fumigatus growth stimulation (arrow) by pyochelin (PCH) sub-inhibitory concentrations. Effect of pyochelin on ΔsidD and ΔsidF in MM (a) and on WT in MM(-Fe) (b). Note the absence of pyochelin stimulation in the TAFC siderophore minus mutant’s ΔsidD and ΔsidF, showing the essentiality of the presence of TAFC. Medium composition and methodology are described in Supplementary Materials.

Figure 4

Pyoverdine (PVD) (a) and pyochelin (PCH) (b) activities on A. fumigatus growth in MM in presence of iron excess, showing that the antifungal effect of pyoverdine on A. fumigatus was abolished in presence of iron excess, whereas pyochelin antifungal activity was not abolished. Methodology is described in Supplementary Materials.

Figure 5

Structure (a) and penetration (b) of pyochelin-4-nitrobenzo[1,2,5]oxadiazole (PCH-NBD) in A. fumigatus swollen conidia. Scale bar represents 5 mm. Methodology is described in Supplementary Materials.

Figure 6

Pyochelin (PCH) induces the production of reactive oxidant (ROS) (a) and reactive nitrogen (RNS) (b) in swollen conidia using 2′,7′-dichlorodihydrofluorescein diacetate (H₂DCFDA) and dihydrorhodamine 123 (DHR123), which are ROS- and RNS-specific fluorescent probes, respectively. Scale bar represents 5 mm. Methodology is described in Supplementary Materials.

Figure 7

Previous bacterial challenge synergistically increases the production of IL1β in response to A. fumigatus. Monocytes were stimulated by RPMI medium (in black), bacterial lipopolysaccharide (LPS, 10ng/mL, in orange), P. aeruginosa alone (in green), A. fumigatus alone (in blue), P. aeruginosa–A. fumigatus sequential infection (in red) according to protocol described in Supplementary Materials. Stars represent statistical difference (* p < 0.05; ** p < 0.01).