BIOFILMS IN THE CLINICAL SETTING FUNCTIONAL GENOMIC STUDIES

Frank-Michael Mueller, MD

Author address: 

University of Heidelberg, Germany

Abstract: 

A biofilm is a barrier against external aggressions like desiccation or penetration of toxic molecules, host defences and antimicrobial drugs. Among fungi, biofilm formation has so far most intensively been studied in the pathogenic yeast C. albicans. Recently, the capacity of A. fumigatus to form a biofilm in vitro was investigated (1,2,4,5). The mycelium of A. fumigatus growing on an agar surface is surrounded by an extracellular matrix (ECM) which glues the hyphae together. This ECM is not produced when the fungus grows as separate hyphae like under shaken submerged conditions in vitro (1). A. fumigatus can produce a parallel packed hyphal network on plastic surfaces or on human bronchial epithelia cells (16HBE) and F508del/F508del (CFBE41o-) embedded in an ECM (5). This in vitro ECM, seen under transmission electron microscopy, as an electron dense outer layer, was found to contain polysaccharides (galactomannan, α -1,3-glucan), melanin, proteins (major antigens, hydrophobins) and monosaccharides (1). Just recently characteristic biofilm structures were observed in vivo in a murine model of invasive aspergillosis, and in surgical excisions of lung and sinus aspergilloma from patients (3). Functional genomic studies were initiated for further characterization of regulated genes and proteins during biofilm formation. A. fumigatus ATCC #9197 was left to produce biofilm for 24h and 48h. For proteome analysis proteins were isolated and a 2D DIGE gel was performed following MALDI-TOF. In parallel, the transcriptome was assessed by c-DNA microarrays. The regulation of selected proteins was confirmed by RT-PCR and by detection in culture supernatants using HPLC analysis. The results indicate that biofilm production of A. fumigatus is multifactorial. The most striking result was the significant upregulation of proteins and genes of the gliotoxin secondary metabolite cluster. The glutathione Stransferase GliG showed a 1.5 fold increased protein level in biofilm in comparison to planktonic growth after 48h. The thioredoxin reductase GliT, showed a 2.1 fold increased level over time. Among the genes of the gliotoxin cluster in 48h biofilm, three were slightly up-regulated in a time dependant manner. Only GliJ and GliO were up-regulated more than 2-fold. A. fumigatus can produce biofilms under various in vitro conditions and in the clinical setting in patients suffering from lung and sinus aspergilloma. The production of secondary metabolites such as gliotoxin in vitro may allow A. fumigatus to survive in vivo. Gliotoxin has immunmodulatory effects on the host's immune system. Growing in a multicellular community, A. fumigatus survival may alleviate especially in chronic aspergillosis in cystic fibrosis patients, and patients suffering from aspergilloma, asthma and ABPA. References Beauvais, A. et al. (2007) Cell Microbiol. 9: 1588-1600. Beauvais, A. and Müller, F.M. (2009) In Aspergillus fumigatus and Aspergillosis, Washington, DC: ASM Press, pp. 149-158. Loussert, C. et al. (2009) Cell Microbiol (in press). Mowat, E. et al. (2007) J Med Microbiol 56. 1205-1212. Seidler, M. J. et al. (2008) Antimicrob Agents Chemother 52: 4130-4136.
2010

abstract No: 

none

Full conference title: 

4th Advances Against Aspergillosis
    • AAA 4th (2010)