Increased Filamentous Growth of Candida albicans in Simulated Microgravity

Knowledge of simulated microgravity (SMG)-induced changes in the pathogenicity of microorganisms is important for success of long-term spaceflight. In a previous study using the high aspect ratio vessel bioreactor, we showed that the yeast species Saccharomyces cerevisiae underwent a significant phenotypic response when grown in modeled microgravity, which was reflected in the analysis of gene expression profiles. In this study, we establish that Candida albicans responds to SMG in a similar fashion, demonstrating that there is a conserved response among yeast to this environmental stress. We also report that the growth of C. albicans in SMG results in a morphogenic switch that is consistent with enhanced pathogenicity. Specifically, we observed an increase in filamentous forms of the organism and accompanying changes in the expression of two genes associated with the yeast-hyphal transition. The morphological response may have significant implications for astronauts’ safety, as the fungal pathogen may become more virulent during spaceflight

Relative gene expression of <i>C. albicans</i> grown in spaceflight versus ground control conditions, as determined by microarray and qRT-PCR analysis.

<p>*P < 0.05,</p><p>**p < 0.01</p><p>Gene expression was normalized using the average of 4 housekeeping genes (<i>ACT1</i>, <i>PMA1</i>, <i>RIP</i>, <i>RPP2B</i>)</p

Hierarchical ranking of the GO Term Finder Process categories that were significantly enriched.

<p>Only categories that are significantly enriched (p<0.05) are presented, except for those labeled grey added for hierarchical purposes. Subcategories with more than 2 higher rank categories that were not significantly enriched are not included in this figure (i.e., dicarboxylic acid transport and copper ion transport). For clarity purposes, categories with more than one connector are not presented, if the connecting category/categories was/were not significantly enriched. Color codes indicate p-values.</p

Light microscopic analyses of fixed <i>C. albicans</i> cultured in spaceflight (A, B) and ground control (a, b) conditions.

<p>Panels A and B: Differential interface contrast (DIC) images at 400× magnification. Panels a, b: DIC images are 630× magnification. Purple circles indicate cell clumps of 4 or more cells.</p

Measurement of cell size and shape of <i>C. albicans</i> spaceflight and ground control cultures.

<p>(A) Surface area of spaceflight and ground cells, organized as percentage of cells per size range (1 µm increments). The percentages for ground and flight cultured <i>C. albicans</i> with a surface area between 0 and 5 µm are indicated. (B) Width-to-length ratio of spaceflight and ground cells, organized as percentage of cells per width-to-length range (0.1 increments). Results were obtained based on surface area and width-to-length determination of 143 ground control cells and 197 spaceflight-cultured cells.</p

Scanning electron microscopy analysis of <i>C. albicans</i> cultured in spaceflight and ground control conditions.

<p>Cell clusters of spaceflight (A, B) and ground control (a, b) conditions are shown. Black arrow points to filament, white arrows indicate aberrant cell shapes, grey arrows indicate normal bipolar budding, and white dotted arrows indicate random budding scars. Magnification  = 5,000× for A and a, and 8,000× for B and b. C and c show images of spaceflight and ground control cells respectively at lower magnification (2,500×) to demonstrate the difference in space occupancy between the test conditions (3D architecture for spaceflight compared to flat structure for ground cultures).</p

Primers used for qRT-PCR analysis.

<p>*<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0080677#pone.0080677-Nailis1" target="_blank">[52]</a>, other primers were designed in this study</p

Biological process categories of <i>C. albicans</i> affected by spaceflight conditions as compared to ground control, based on GO Slim Mapper analysis.

<p>*Based on 454 genes differentially regulated in response to spaceflight</p