The use of flow cytometry as a tool for monitoring filament formation of fungi

Abstract

Flow cytometry (FC) has the ability to discriminate a variety of cell parameters including cell size and complexity, and fluorescence intensity. As yeast cells or fungal spores germinate they undergo a morphological transformation from round oval shaped cells to elongate filamentous forms. To date, monitoring these events has been performed using microscopic examination. Microscopic examination is a labor intensive process that examines a very small percentage of the total cell population. We have developed a method using FC that is rapid, simple to perform, and reproducible. The major advantages of FC include analysis of a larger number of cells, increased objectivity due to nonselective measurements of all cells in the population studied, and the computer related data analysis capability of the flow cytometer.

Antifungal Susceptibility Testing: Practical Aspects and Current Challenges

“…Staining (or lack of staining) with suitable dyes permits the rapid detection of damaged fungi. Recent work has further explored this possibility (80,88,97,151,175,176,223) and shown the potential for correlation of flow cytometry-based techniques with the reference method. One study suggested that flow cytometry might be especially useful for detection of amphotericin B resistance (69).…”

Role of flow cytometry for the improvement of bioprocessing of oleaginous microorganisms

“…This may be due to the fact that the filamentous nature of fungi makes it complex to screen or sort using flow cytometry. However, some studies reported the use of flow cytometry for the improvement of non‐oleaginous fungal strains . Successful isolation of fluorescent dye stained and/or fluorescent protein/tag expressed fungal spores and conidia was demonstrated in different non‐oleaginous fungal strains including, Aspergillus fumigatus, Coniothyrium minitans, Trichoderma reesei, Trichoderma viridae, Penicillium expansum and Phoma herbarum .…”

“…The study analysed metabolic activities in the developing hyphae of non‐oleaginous filamentous fungi using permanent fluorescent dyes DHE and HI. Another study by Hopfer et al . used flow cytometry to monitor filament formation in EtBr stained A. fumigatus .…”

“…However, some studies reported the use of flow cytometry for the improvement of non-oleaginous fungal strains. [98][99][100][101][102] Successful isolation of fluorescent dye stained and/or fluorescent protein/tag expressed fungal spores and conidia was demonstrated in different non-oleaginous fungal strains including, Aspergillus fumigatus, Coniothyrium minitans, Trichoderma reesei, Trichoderma viridae, Penicillium expansum and Phoma herbarum. Green fluorescent protein (GFP) was the most frequently used fluorescent protein for the improvement of non-oleaginous fungal strains.…”

“…The study analysed metabolic activities in the developing hyphae of non-oleaginous filamentous fungi using permanent fluorescent dyes DHE and HI. Another study by Hopfer et al 102 used flow cytometry to monitor filament formation in EtBr stained A. fumigatus. A study by Pang et al 103 reported BODIPY 505/515 staining of intracellular lipid bodies of marine oomycete Halophytophthora species, and lipid bodies distribution within mycelia of Halophytophthora was demonstrated.…”