In vivo analysis of Aspergillus fumigatus developmental gene expression determined by real-time reverse transcription-PCR


Gravelat FN, Doedt T, Chiang LY, Liu H, Filler SG, Patterson TF, Sheppard DC

Date: 4 July 2008


Very little is known about the developmental stages of A. fumigatus during invasive aspergillosis. We performed real-time RT-PCR analysis on lung samples of 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 pre-competent 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. Expression of gliP mRNA in vitro was found to be highly dependent on culture conditions. Further, 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 evaluating of putative virulence factors expressed by competent hyphae and analyzing gene expression levels during invasive infection rather than in vitro alone.

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