Aspergillus fumigatus, One Uninucleate Species with Disparate Offspring

Abstract

Establishment of a fungal infection due to Aspergillus fumigatus relies on the efficient germination of the airborne conidia once they penetrate the respiratory tract. However, the features of conidial germination have been poorly explored and understood in this fungal species as well as in other species of filamentous fungi. We show here that the germination of A. fumigatus is asynchronous. If the nutritional environment and extensive gene deletions can modify the germination parameters for A. fumigatus, the asynchrony is maintained in all germinative conditions tested. Even though the causes for this asynchrony of conidial germination remain unknown, asynchrony is essential for the completion of the biological cycle of this filamentous fungus.

Keywords: Aspergillus fumigatus; aspergillosis; asynchrony; conidium; germination; transcriptome.

Figure 1

Asynchrony of conidial germination after 6 h (a) and 9.30 h (b) of incubation in the MM medium. See the resting (arrow heads), swollen (arrows) and germinated conidia

Figure 2

Asynchrony of conidial germination estimated by the measurement of the conidial volume of 100 conidia after incubation in the MM medium; (a) scatter plot of the volume of each conidia measured every 4 min; (b) box plot of the volume of all conidia measured every 60 min. Volume (estimated as pixels) was measured only on conidia without germ tubes.

Figure 3

Morphology of the conidia germinating in G (glucose), GA (glucose + ammonium tartrate), GAP (glucose + ammonium tartrate + phosphate) and complex MM for 24 h.

Figure 4

Quantification of the conidium germination in G, GA, GAP and MM media. (a) Average size in the different media until the emission of the first germ tube (computed as conidial surface estimated by pixels); (b) heterogeneity of germination with 100 swollen conidia (until the emission of the first germ tubes) randomly selected; (c) percentage of germinated conidia with germ tubes.

Figure 5

Heterogeneity in the population of germinated conidia estimated by the size of the germ tubes (in µm) in the different media. Incubation time was 24 h in G, GA, GAP and 11 h in MM. Median values and variance are shown in the upper right corner of each plot.

Figure 6

Nuclei in germinating conidia. Germinated conidia in G and GA media only displayed one nucleus per conidium after 24 h of germination. Active nuclear division in MM and GAP media after respectively 11 h and 24 h germination. Staining with Hoechst of germinating conidia; Nucleus staining by Hoechst.

Figure 7

Septa shown by CalcoFluor staining of germinating conidia in different media. (a) G; (b) GA; (c) GAP; (d) MM.

Figure 8

Heat maps showing significative examples of differentially regulated A. fumigatus genes during germination in GAP and MM media. Level of expression of genes for a ratio GAP/MM: (a) down regulation of ergosterol synthesis and upregulation of trehalose synthesis suggesting the entrance in quiescence of conidia germinating in GAP medium; (b) ribosome and protein synthesis pathways down regulated in absence of growth in GAP medium; (c) Down regulation of cell wall synthases and up regulation of autolytic cell wall glycosyl hydrolases indicating that the germ tubes are starved in GAP medium; (d) high expression of genes coding for plant material degrading glycosylhydrolases indicating the starvation status of GAP cells and the attemps to its quest for food.

Figure 8

Heat maps showing significative examples of differentially regulated A. fumigatus genes during germination in GAP and MM media. Level of expression of genes for a ratio GAP/MM: (a) down regulation of ergosterol synthesis and upregulation of trehalose synthesis suggesting the entrance in quiescence of conidia germinating in GAP medium; (b) ribosome and protein synthesis pathways down regulated in absence of growth in GAP medium; (c) Down regulation of cell wall synthases and up regulation of autolytic cell wall glycosyl hydrolases indicating that the germ tubes are starved in GAP medium; (d) high expression of genes coding for plant material degrading glycosylhydrolases indicating the starvation status of GAP cells and the attemps to its quest for food.

Figure 8

Heat maps showing significative examples of differentially regulated A. fumigatus genes during germination in GAP and MM media. Level of expression of genes for a ratio GAP/MM: (a) down regulation of ergosterol synthesis and upregulation of trehalose synthesis suggesting the entrance in quiescence of conidia germinating in GAP medium; (b) ribosome and protein synthesis pathways down regulated in absence of growth in GAP medium; (c) Down regulation of cell wall synthases and up regulation of autolytic cell wall glycosyl hydrolases indicating that the germ tubes are starved in GAP medium; (d) high expression of genes coding for plant material degrading glycosylhydrolases indicating the starvation status of GAP cells and the attemps to its quest for food.

Figure 8

Heat maps showing significative examples of differentially regulated A. fumigatus genes during germination in GAP and MM media. Level of expression of genes for a ratio GAP/MM: (a) down regulation of ergosterol synthesis and upregulation of trehalose synthesis suggesting the entrance in quiescence of conidia germinating in GAP medium; (b) ribosome and protein synthesis pathways down regulated in absence of growth in GAP medium; (c) Down regulation of cell wall synthases and up regulation of autolytic cell wall glycosyl hydrolases indicating that the germ tubes are starved in GAP medium; (d) high expression of genes coding for plant material degrading glycosylhydrolases indicating the starvation status of GAP cells and the attemps to its quest for food.

Figure 9

Asynchrony of conidial germination in a conidial chain of a glucanase mutant characterized by a defect in conidial separation. Note that the presence of resting, swollen or germinating conidia is totally independent of the conidium age.

Figure 10

Stimulation of the germ tube growth by the GSK1324726A compound in the G or GA medium. The elongation of the germ tube was associated to an increase in nuclear divisions. Note the high number of nuclei per germinated conidia while in absence of drug the growth of the germ tube was stopped and contained only one nucleus per germ tube.