Analysis of gene expression in filamentous cells of Candida albicans grown on agar plates.

Paul E Creger,Jill R Blankenship

doi:10.14440/jbm.2018.211

Abstract

Candida albicans (C. albicans) is a commensal organism of the human gastrointestinal and genitourinary tracts. C. albicans is also a major human pathogen, causing disease ranging from cutaneous infections to lethal systemic disease. The ability of this fungus to switch between yeast and filamentous forms of growth has long been linked to its pathogenesis. Filamentation can be induced by a variety of distinct environmental cues and can occur in either liquid or solid media. While some evidence suggests that there are differences between filamentation in solid and liquid media, gene expression analysis of filamentation in C. albicans has focused strictly on cells grown in liquid media. We have developed a method for analyzing gene expression of filamentous cells grown on solid induction media at early stages of filamentation, establishing cell plating densities, ideal collection times, and collection techniques. We have also demonstrated the utility of the approach not only in qRT-PCR assays, but high-throughput RNAseq assays as well. These assays will allow for comparison studies of C. albicans filamentation initiation in solid and liquid media.

Highlights

The pathogenic fungus Candida albicans (C. albicans) is a common commensal organism in the gastrointestinal and genitourinary tracts of humans
We have developed a method for harvesting C. albicans filamentous and yeast cells from solid media for RNA extraction that can be used in qRT-PCR and RNAseq studies
Plating density impacts filamentation Overnight cultures of C. albicans wild type SC5314 cells grown at 30°C in YPD liquid media were washed twice with phosphate buffered saline (PBS) to remove YPD and quorum sensing molecules, such as farnesol and tyrosol, that might impact filament development [7,8]

Summary

Introduction

The pathogenic fungus Candida albicans (C. albicans) is a common commensal organism in the gastrointestinal and genitourinary tracts of humans. It can be pathogenic, with infections ranging from cutaneous infections of the oral and genital mucosa to lethal systemic disease. C. albicans and other Candida spp. infections are a leading cause of systemic infections in hospital settings and mortality rates are high (~40%) [1]. C. albicans is a dimorphic fungus that can grow in a variety of forms from yeast-like cells to true filaments. The ability to transition between the yeast and filamentous forms is vital to pathogenesis [2,3,4] and is the most heavily studied aspect of C. albicans biology. Phenotypic assays are generally performed in a variety of media conditions, but very little effort has been focused on comparisons of gene expression between the conditions

Methods

Results

Conclusion