Extracellular DNA release acts as an antifungal resistance mechanism in mature Aspergillus fumigatus biofilms

Ranjith Rajendran, Craig Williams, David F. Lappin, Owain Millington, Margarida Martins, Gordon Ramage

Research output: Contribution to journalArticle

Abstract

Aspergillus fumigatus has been shown to form biofilms that are associated with adaptive antifungal resistance mechanisms. These include multidrug efflux pumps, heat shock proteins, and extracellular matrix (ECM). ECM is a key structural and protective component of microbial biofilms and in bacteria has been shown to contain extracellular DNA (eDNA). We therefore hypothesized that A. fumigatus biofilms also possess eDNA as part of the ECM, conferring a functional role. Fluorescence microscopy and quantitative PCR analyses demonstrated the presence of eDNA, which was released phase dependently (8 <12 <24 <48 h). Random amplification of polymorphic DNA (RAPD) PCR showed that eDNA was identical to genomic DNA. Biofilm architectural integrity was destabilized by DNase treatment. Biochemical and transcriptional analyses showed that chitinase activity and mRNA levels of chitinase, a marker of autolysis, were significantly upregulated as the biofilm matured and that inhibition of chitinases affected biofilm growth and stability, indicating mechanistically that autolysis was possibly involved. Finally, using checkerboard assays, it was shown that combinational treatment of biofilms with DNase plus amphotericin B and caspofungin significantly improved antifungal susceptibility. Collectively, these data show that eDNA is an important structural component of A. fumigatus ECM that is released through autolysis, which is important for protection from environmental stresses, including antifungal therapy.
Original languageEnglish
Pages (from-to)420-429
Number of pages10
JournalEukaryotic Cell
Volume12
Issue number3
DOIs
Publication statusE-pub ahead of print - 13 Jan 2013
Externally publishedYes

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Aspergillus fumigatus
Biofilms
DNA
Autolysis
Chitinases
Extracellular Matrix
Deoxyribonucleases
caspofungin
Polymerase Chain Reaction
Conservation of Natural Resources
Amphotericin B
Heat-Shock Proteins
Fluorescence Microscopy
Bacteria
Messenger RNA
Growth

Cite this

Rajendran, Ranjith ; Williams, Craig ; Lappin, David F. ; Millington, Owain ; Martins, Margarida ; Ramage, Gordon. / Extracellular DNA release acts as an antifungal resistance mechanism in mature Aspergillus fumigatus biofilms. In: Eukaryotic Cell. 2013 ; Vol. 12, No. 3. pp. 420-429.
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Extracellular DNA release acts as an antifungal resistance mechanism in mature Aspergillus fumigatus biofilms. / Rajendran, Ranjith; Williams, Craig; Lappin, David F.; Millington, Owain; Martins, Margarida; Ramage, Gordon.

In: Eukaryotic Cell, Vol. 12, No. 3, 13.01.2013, p. 420-429.

Research output: Contribution to journalArticle

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