Antifungal Susceptibility Testing and Drug Discovery in the Dimorphic Fungus Histoplasma Capsulatum.

Abstract

The thermal dimorphism of the fungal pathogen Histoplasma is linked to its virulence in mammalian hosts. Mammalian body temperature triggers differentiation of the fungus into virulent yeasts which successfully infect host phagocytes. Accurate determination of antifungal susceptibility with relevance to infection requires that the tests be performed specifically using the yeast form, not the filamentous environmental form. However, traditional CLSI methodology for antifungal susceptibility testing of yeasts with Histoplasma is in adequate. We present optimized methodology for performing antifungal susceptibility assays on Histoplasma yeasts with an emphasis on quantitative yeast growth determination. Colorimetric and fluorometric assays for Histoplasma growth overcome challenges associated with quantifying some Histoplasma strains which grow as aggregates of yeasts. We also describe antifungal susceptibility testing of Histoplasma yeasts within macrophages to provide improved accuracy and better physiological relevance of antifungal susceptibility profiles.