Phenotypic screening, transcriptional profiling, and comparative genomic analysis of an invasive and non-invasive strain of Candida albicans

Abstract

Background: Invasion of host tissue by the human fungal pathogen Candida albicans is an important step during the development of candidosis. However, not all C. albicans strains possess the same invasive and virulence properties. For example, the two clinical isolates SC5314 and ATCC10231 differ in their ability to invade host tissue and cause experimental infections. Strain SC5314 is invasive whereas strain ATCC10231 is non-invasive and strongly attenuated in virulence compared to SC5314. In this study we compare the in vitro phenotypic, transcriptional and genomic profiles of these two widely used laboratory strains in order to determine the principal biological and genetic properties responsible for their differential virulence.

Results: In all media tested, the two strains showed the same metabolic flexibility, stress resistance, adhesion properties and hydrolytic enzyme secretion in vitro. However, differences were observed in response to cell-surface disturbing agents and alkaline pH. Furthermore, reduced hyphal formation in strain ATCC10231 under certain conditions correlated with reduced invasive properties in an in vitro invasion assay and a reduced ability to invade epithelial tissue. Despite these diverse phenotypic properties, no substantial genomic differences were detected by comparative genome hybridisation within the open reading frames. However, in vitro transcriptional profiling displayed major differences in the gene expression of these two strains, even under normal in vitro growth conditions.

Conclusion: Our data suggest that the reason for differential virulence of C. albicans strains is not due to the absence of specific genes, but rather due to differences in the expression, function or activity of common genes.

Figure 1

Growth characteristics of C. albicans strains SC5314 and ATCC10231 in SD medium. Representative figure of n = 4 experiments. Both strains showed similar growth curves in YPD medium (data not shown).

Figure 2

(A) Growth of C. albicans strains SC5314 and ATCC10231 under different stress conditions (addition of itraconazole, ethanol, 5-FOA). Growth on SD plates served as a control. Both strains were tested at 30°C and at 37°C. (B) Invasion of SC5314 and ATCC10231 into carbon-depleted agar surface. After growth of the colonies, plates were washed with water removing the non-invasive cells and photographed. (C) Influence of different pH-values on the growth of SC5314 and ATCC10231 at iron- (150 μM BPS) and phosphate-limiting (0.15 mM P_i) conditions and under cation stress (1 M NaCl). Growth on YPD at pH 5 and pH 8, respectively, served as a control. Numbers under the single colonies represent the number of cells used for inoculation of each spot (also related to A and C).

Figure 3

Adhesion of C. albicans strains SC5314 and ATCC10231 to fibroblasts (NIH/3T3) and epithelial cells (HEp-2). The non-adherent S. cerevisiae strain served as a negative control. * = significantly reduced adhesion compared to SC5314 and ATCC10231 with P < 0.05. Error bars = SD of n = 3 experiments.

Figure 4

(A) Relative LDH activity in the maintenance medium of the RHE after infection with C. albicans strains SC5314 and ATCC10231. * = significantly reduced LDH-activity compared to strain SC5314 with P < 0.05. Error bars = SD of n = 4 experiments (B) Histological sections of infected RHE samples. Arrows indicate invading hyphae of strain SC5314.

Figure 5

Plot of the signal ratio and the sequence homology (compared to strain SC5314) of different genes from strain ATCC10231. Thirty-seven genes gave at least a 2-fold weaker signal with DNA from strain ATCC10231 compared to SC5314 (filled circles). Additionally, 15 genes were included which were already sequenced and published (open circles). Filled triangles indicate 5 genes that gave a stronger signal with DNA from ATCC10231 compared to SC5314. The cut-off value for substantial differences within the sequence homology of the two strains was set at 90%. Only three genes (PHO81, HAL21, HAL22) fulfilled the criteria of both low signal ratio and low sequence homology.