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. 2019 Jan 18;9:218. doi: 10.1038/s41598-018-36524-8

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Induction of cell wall stress and upregulation of cell wall-related genes in A. fumigatus mediated by K. pneumoniae. (A) Schematic view of flow sorting of A. fumigatus and K. pneumoniae (ATCC 700603). Sorted cells were plated onto 12-well agar plates containing kanamycin (1000 µg/mL) and colistin (100 µg/mL) for selection of fungal growth and prevention of any residual bacterial contamination. Plates also contained different concentration ranges of cell wall stressors and antifungal drugs to facilitate the observation of the sensitivity of fungi previously grown alone or exposed to K. pneumoniae. (B) Sensitivity of A. fumigatus to oxidative (H₂O₂), osmotic (sorbitol, SDS) and cell wall (CFW) stresses, with and without previous exposure to K. pneumoniae. (C) Sensitivity of A. fumigatus to antifungal drugs (amphothericin B, voriconazole, micafungin) with and without previous exposure to K. pneumoniae. (D) Upregulation of A. fumigatus cell wall-related genes in the presence of K. pneumoniae. (E) Downregulation of hyphae-related genes of A. fumigatus in the presence of K. pneumoniae determined by quantification of mRNA expression of hyphae-related genes of A. fumigatus upon exposure to K. pneumoniae (ATCC 700603). Fungal spores were germinated for up to 12 h followed by addition of bacteria for a total of 24 h. Transcript levels of the A. fumigatus single culture were set to one, and the transcript levels in the co-culture were normalized to the single culture. Additionally, normalization was also performed against the reference gene TUBA. Data are presented as mean + SE of four independent experiments.