Convergent evolution in toxin detection and resistance provides evidence for conserved bacterial–fungal interactions

PA4170 is the most highly differentially expressed P. aeruginosa gene during coculture with A. fumigatus in SCFM2, and expression is activated upon gliotoxin exposure. (A) Schematic illustrating the A. fumigatus and P. aeruginosa coculture setup. A. fumigatus is first inoculated into SCFM2 to facilitate germination and hyphal development, followed by P. aeruginosa inoculation. Coculture was imaged by confocal laser scanning microscopy (CLSM) at 18 h, 37 °C. A. fumigatus was labeled with constitutively expressed mNeonGreen, and P. aeruginosa with mCherry. The data are representative of three independent experiments, each performed in triplicate. (Scale bar, 20 µm.) (B) Volcano plot illustrating P. aeruginosa genes differentially expressed upon coculture with A. fumigatus. Bubble colors are used to indicate genes significantly altered in expression with a log2 fold change greater than 1 (blue), less than −1 (green), above the adjusted P-value threshold (orange), or unchanged (black). Significantly altered clusters of genes (>5) with established functions in P. aeruginosa are shown (zinc acquisition, iron acquisition, denitrification, and copper homeostasis). PA4170 (dotted box) was the highest differentially expressed gene (93.7-fold-change). The experiment was performed using biological triplicates. (C) mCherry-tagged PA4170 reporter assay examining PA4170 expression following P. aeruginosa growth in spent A. fumigatus SCFM culture supernatants (24 h) supplemented with different metals (5 µM FeSO4, ZnSO4, CuSO4, and MnSO4). Fluorescence was read at 6 h and normalized to cell density (OD600). The data are representative of three independent experiments performed in triplicate. Statistical significance was calculated by one-way ANOVA with Dunnett’s multiple comparisons test using “SCFM Af” as the control column (****P < 0.0001). Error bars represent SD from the mean. (D) Untargeted metabolomics of A. fumigatus culture supernatants during growth in SCFM and SCFM + 10 µM ZnSO4 (24 h) identified gliotoxin as significantly lowered upon zinc supplementation. Chromatograms show the peak corresponding to gliotoxin (triplicate), alongside a commercial standard. The experiment was performed using biological triplicates. (E) mCherry-tagged PA4170 reporter fluorescence assay examining PA4170 protein expression in SCFM following gliotoxin (GT) or bisthiomethylgliotoxin (bmGT) supplementation (0 to 60 µM). Fluorescence was read at 8 h and normalized to cell density (OD600). The data are representative of two independent experiments performed in triplicate. Statistical significance was calculated by one-way ANOVA with Dunnett’s multiple comparisons test using “0 µM GT” as the control column (*P = 0.0197 and ****P < 0.0001). Error bars represent SD from the mean.
Stephen Dolan’s latest research, published in PNAS, sheds light on the complex interactions within polymicrobial communities. The study reveals how the human pathogen Pseudomonas aeruginosa has developed a genetic network to detect and neutralize gliotoxin, a potent antimicrobial produced by the fungus Aspergillus fumigatus. Dolan’s findings highlight a fascinating example of convergent evolution, where both the bacterium and fungus have evolved similar mechanisms for toxin resistance despite their evolutionary differences. This discovery enhances our understanding of microbial interactions and could inform future treatments for infections involving P. aeruginosa. Read the article.