Transcriptional profiling reveals the expression of novel genes in response to various stimuli in the human dermatophyte Trichophyton rubrum.

BACKGROUND: Cutaneous mycoses are common human infections among healthy and immunocompromised hosts, and the anthropophilic fungus Trichophyton rubrum is the most prevalent microorganism isolated from such clinical cases worldwide. The aim of this study was to determine the transcriptional profile of T. rubrum exposed to various stimuli in order to obtain insights into the responses of this pathogen to different environmental challenges. Therefore, we generated an expressed sequence tag (EST) collection by constructing one cDNA library and nine suppression subtractive hybridization libraries. RESULTS: The 1388 unigenes identified in this study were functionally classified based on the Munich Information Center for Protein Sequences (MIPS) categories. The identified proteins were involved in transcriptional regulation, cellular defense and stress, protein degradation, signaling, transport, and secretion, among other functions. Analysis of these unigenes revealed 575 T. rubrum sequences that had not been previously deposited in public databases. CONCLUSION: In this study, we identified novel T. rubrum genes that will be useful for ORF prediction in genome sequencing and facilitating functional genome analysis. Annotation of these expressed genes revealed metabolic adaptations of T. rubrum to carbon sources, ambient pH shifts, and various antifungal drugs used in medical practice. Furthermore, challenging T. rubrum with cytotoxic drugs and ambient pH shifts extended our understanding of the molecular events possibly involved in the infectious process and resistance to antifungal drugs.

In vitro antifungal drug susceptibilities of dermatophytes microconidia and arthroconidia.

OBJECTIVES: Arthroconidia have been considered as the primary cause of infection by dermatophytes. However, the in vitro antifungal testing evaluates the responses mainly of microconidia or hyphae, and dermatophytes in vivo often produce arthroconidia, a cellular structure presumably more resistant to antifungals. The aim of this study was to compare the in vitro susceptibility of microconidia and arthroconidia of Trichophyton rubrum, Trichophyton tonsurans and Trichophyton equinum to griseofulvin, itraconazole, terbinafine, fluconazole, amphotericin B and hygromycin B. METHODS: Microconidia and arthroconidia were produced in vitro, and their susceptibility to each drug was evaluated by assessing the CLSI M38-A broth microdilution method. RESULTS: Arthroconidia of all strains analysed appeared to be more resistant to fluconazole, griseofulvin and itraconazole than microconidia. The MIC of terbinafine was the same for microconidia and arthroconidia for all strains, and the MIC of amphotericin B for microconidia and arthroconidia was the same for isolates of T. equinum and T. tonsurans, but differed for T. rubrum. Finally, the level of resistance of microconidia for all strains towards the antibiotic hygromycin B was from 25 to 400 mg/L. CONCLUSIONS: The difference in the susceptibility between microconidia and arthroconidia depends on the drug and on the strain, and may be one of the causes of therapeutic failure. Also, the level of resistance to the antibiotic hygromycin B presented by microconidia of these isolates will allow the use of hygromycin resistance as a dominant marker in fungal transformation procedures in future studies of gene function.

Electrophoretic molecular karyotype of the dermatophyte Trichophyton rubrum

The electrophoretic karyotype of the dermatophyte Trichophyton rubrum was established using contour-clamped homogeneous electric field (CHEF) gel electrophoresis. Five chromosomal bands of approximately 3.0 to 5.8 megabase pairs (Mbp) each were observed and together indicated that 22.05 Mbp of the total genome are organized as chromosomal macromolecules. In addition to establishing the number and size of T. rubrum chromosomes, these results open perspectives for the construction of chromosome-specific libraries and for the physical mapping of genes of interest, thus permitting future gene linkage studies in this pathogen. A detailed understanding of the karyotype and genomic organization of T. rubrum should contribute to further genetic, taxonomic and epidemiological studies of this dermatophyte.