Penicillium marneffei: Gene expression profiling of hyphal and yeast cells and during conidiation

Penicillium marneffei (Talaromyces marneffei) is an opportunistic human pathogen that can grow in a multicellular hyphal form at 25°C or a unicellular fission yeast form at 37°C, and can switch between these two forms in response to temperature. The yeast form is found in infected individuals and represents the pathogenic phase. In response to specific environmental cues, the hyphal form can undergo asexual development (conidiation) the produce a differentiated multicellular structure called a conidiophore which produces dormant spores (conidia). Inhaled conidia initiate infection. To identify genes that are ‘phase or cell-state specific’ transcriptional profiling was performed with a custom microarray consisting of ~42% of the predicted P. marneffei genes and RNA from P. marneffei hyphal, yeast and conidiation cell types. A custom microarray consisting of short, random genomic fragments from P. marneffei (~42% of the predicted genes) was generated using PCR products of the inserts from two independent DNA libraries constructed from genomic DNA of the P. marneffei type strain FRR2161. PCR products from previously cloned P. marneffei genes with known expression profiles were included as controls on the microarray. Total RNA from hyphal, yeast or conidiation cell types was prepared and used in pairwise combinations (conidiation versus hyphal cells, conidiation versus yeast cells and hyphal versus yeast cells) on the microarrays. Three biological replicate cultures were prepared for each experiment.