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Infection and Immunity, October 2001, p. 6411-6418, Vol. 69, No. 10
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.10.6411-6418.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Differential Expression of the Aspergillus fumigatus pksP Gene Detected In Vitro and In Vivo with Green Fluorescent Protein

Kim Langfelder,1 Bruno Philippe,2 Bernhard Jahn,3 Jean-Paul Latgé,2 and Axel A. Brakhage1,*

Institut für Mikrobiologie, Universität Hannover, D-30167 Hannover,1 and Institut für Medizinische Mikrobiologie und Hygiene, Universität Mainz, D-55101 Mainz,3 Federal Republic of Germany, and Institut Pasteur, Laboratoire des Aspergillus, F-75015 Paris, France2

Received 7 March 2001/Returned for modification 17 May 2001/Accepted 28 June 2001

Aspergillus fumigatus is an important pathogen of immunocompromised hosts, causing pneumonia and invasive disseminated disease with high mortality. To be able to analyze the expression of putative virulence-associated genes of A. fumigatus, the use of the enhanced green fluorescent protein (EGFP) as a reporter was established. Two 5' sequences, containing the putative promoters of the pyrG gene, encoding orotidine-5'-phosphate decarboxylase, and the pksP gene, encoding a polyketide synthase involved in both pigment biosynthesis and virulence of A. fumigatus, were fused with the egfp gene. The PpksP-egfp construct was integrated via homologous recombination into the genomic pksP locus. EGFP production was analyzed by fluorescence spectrometry, Western blot analysis, and fluorescence microscopy. Differential gene expression in A. fumigatus was observed. Fluorescence derived from the PYRG-EGFP fusion protein was detected during all developmental stages of the fungus, i.e., during germination, during vegetative growth, in conidiophores, and weakly in conidia. In addition, it was also detected in germinating conidia when isolated from the lungs of immunocompromised mice. By contrast, PKSP-EGFP-derived fluorescence was not found in hyphae or stalks of conidiophores but was found in phialides and conidia in vitro when the fungus was grown under standard conditions, indicating a developmentally controlled expression of the gene. Interestingly, pksP-egfp expression was also detected in hyphae of germinating conidia isolated from the lungs of immunocompromised mice. This finding indicates that the pksP gene can also be expressed in hyphae under certain conditions and, furthermore, that the pksP gene might also contribute to invasive growth of the fungus.

* Corresponding author. Mailing address: Institut für Mikrobiologie, Universität Hannover, Am Schneiderberg 50, D-30167 Hannover, Germany. Phone: 49 511 762 5945. Fax: 49 511 762 5287. E-mail: Brakhage{at}mbox.ifmb.uni-hannover.de.

Infection and Immunity, October 2001, p. 6411-6418, Vol. 69, No. 10
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.10.6411-6418.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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Copyright © 2001 by the American Society for Microbiology. All rights reserved.