%0 Figure %A Altamirano, Sophie %A Simmons, Charles %A Kozubowski, Lukasz %D 2018 %T Image_3_Colony and Single Cell Level Analysis of the Heterogeneous Response of Cryptococcus neoformans to Fluconazole.JPEG %U https://frontiersin.figshare.com/articles/figure/Image_3_Colony_and_Single_Cell_Level_Analysis_of_the_Heterogeneous_Response_of_Cryptococcus_neoformans_to_Fluconazole_JPEG/6598175 %R 10.3389/fcimb.2018.00203.s003 %2 https://frontiersin.figshare.com/ndownloader/files/12086846 %K Cryptococcus neoformans %K fluconazole %K drug susceptibility %K drug resistance %K phenotypic heterogeneity %X

Cryptococcus neoformans is a human fungal pathogen that can cause fatal meningitis in immunocompromised individuals. Fluconazole (FLC) is a fungistatic drug administered to treat cryptococcosis. When exposed to the inhibitory concentration of FLC, C. neoformans exhibits heteroresistance where a small subpopulation of cells develops into FLC-resistant colonies. FLC-resistant cells are aneuploids with regard to specific beneficial chromosomal regions. Factors underlying the potential for only certain C. neoformans cells in a genetically isogenic population to become FLC-resistant are unknown. In this study, we systematically examine the heterogeneous response of C. neoformans to FLC at a colony and individual cell level. We find that the heterogeneity in response to FLC is reflected by variable diminishment of the ergosterol at the plasma membrane. A population of C. neoformans spread on a semi-solid medium displays two types of outcomes following FLC exposure. The first outcome is colonies consisting of non-resistant cells (survivors). The size of colonies consisting of survivors ranges from a few cells to visible colonies, which reflects intrinsic phenotypic heterogeneity of the C. neoformans population. The second outcome is FLC-resistant cells forming colonies of sizes significantly larger as compared to colonies made of survivors. We propose a model that describes how a distribution of these types of cellular responses within a population changes depending on FLC concentration and factors that influence the rate of cellular growth including temperature, media type, growth phase, and the age of cells. Our findings highlight a complex nature of the response to a fungistatic drug and provide insights that may help to optimize FLC therapy.

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