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Infection and Immunity, August 2008, p. 3632-3639, Vol. 76, No. 8
0019-9567/08/$08.00+0     doi:10.1128/IAI.01483-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

In Vivo Analysis of Aspergillus fumigatus Developmental Gene Expression Determined by Real-Time Reverse Transcription-PCR

Fabrice N. Gravelat,^1, Thomas Doedt,^2, Lisa Y. Chiang,² Hong Liu,² Scott G. Filler,^2,3 Thomas F. Patterson,⁴ and Donald C. Sheppard^1*

Departments of Microbiology and Immunology, Medicine, McGill University, Montreal, Quebec, Canada,¹ Division of Infectious Diseases, Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, California,² David Geffen School of Medicine at UCLA, Los Angeles, California,³ University of Texas Health Science Center at San Antonio, San Antonio, Texas⁴

Received 6 November 2007/ Returned for modification 13 December 2007/ Accepted 4 May 2008

Very little is known about the developmental stages of Aspergillus fumigatus during invasive aspergillosis. We performed real-time reverse transcription-PCR analysis on lung samples from mice with invasive pulmonary aspergillosis to determine the expression of A. fumigatus genes that are expressed at specific stages of development. In established infection, A. fumigatus exhibited mRNA expression of genes specific to developmentally competent hyphae, such as stuA. In contrast, mRNA of genes expressed by conidia and precompetent hyphae was not detected. Many genes required for mycotoxin synthesis, including aspHS, gliP, mitF, and metAP, are known to be expressed by developmentally competent hyphae in vitro. Interestingly, each of these genes was expressed at significantly higher levels during invasive infection than in vitro. The expression of gliP mRNA in vitro was found to be highly dependent on culture conditions. Furthermore, gliP expression was found to be dependent on the transcription factor StuA both in vitro and in vivo. Therefore, developmentally competent hyphae predominate during established invasive infection, and many mycotoxin genes are expressed at high levels in vivo. These results highlight the importance of the evaluation of putative virulence factors expressed by competent hyphae and analysis of gene expression levels during invasive infection rather than in vitro alone.

Published ahead of print on 19 May 2008.

Editor: A. Casadevall

F.N.G. and T.D. contributed equally to this work.