Colony and Single Cell Level Analysis of the Heterogeneous Response of Cryptococcus neoformans to Fluconazole
Open accessAbstract
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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For C. neoformans, heteroresistance can be reduced or inhibited by several environmental factors, including temperature, media type, growth phase, and age of cells.
“For C. neoformans, heteroresistance can be reduced or inhibited by several environmental factors, including temperature, media type, growth phase, and the age of cells [147].”
From Insight into the Mechanisms and Clinical Relevance of Antifungal Heteroresistance (2025) · cites this paper as [147]
The Results (‘Rate of growth influences the response to FLC’; page unavailable) show that temperature, medium, and growth phase change the response to fluconazole, while the Results on cell age show that younger cells have a less heterogeneous response. But the paper identifies cell age as an intrinsic, not environmental, factor; it also reports that some medium and growth-phase conditions increase colony survival rather than inhibit heteroresistance. (LLM, conf=0.84)
Experimental — accuracy not yet measuredFor C. neoformans, heteroresistance can be reduced or inhibited by environmental factors including temperature, media type, growth phase, and cell age.
“For C. neoformans, heteroresistance can be reduced or inhibited by several environmental factors, including temperature, media type, growth phase, and cell age [147].”
From Insight into the Mechanisms and Clinical Relevance of Antifungal Heteroresistance (2025) · cites this paper as [147]
The Results (‘Rate of growth influences the response to FLC,’ page unavailable) show that temperature, medium and growth phase change colony formation or fluconazole susceptibility; the Results on cell age show that younger cells have a less heterogeneous response. But these findings do not establish that all four factors reduce or inhibit heteroresistance specifically: the paper distinguishes non-resistant survivors from resistant colonies and reports that higher temperature maintained, or potentially increased, the number of resistant colonies. (LLM, conf=0.84)
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Insight into the Mechanisms and Clinical Relevance of Antifungal Heteroresistance
Su, Y., Li, Y., Yi, Q., Xu, Y., Sun, T., & Li, Y. (2025) · Journal of Fungi · doi:10.3390/jof11020143 · Cited by 18 · see citing works
“For C. neoformans , heteroresistance can be reduced or inhibited by several environmental factors, including temperature, media type, growth phase, and the age of cells [147].”
Influential
- Authors
- Altamirano, S., Simmons, C., & Kozubowski, L.
- Affiliations
- Clemson University (US)
- Corresponding author
- Kozubowski, Lukasz
- ORCID
- Kozubowski, Lukasz
- Venue
- Frontiers in Cellular and Infection Microbiology
- Publisher
- Frontiers Media SA
- Published
- 2018-06-19
- Volume
- 8
- Pages
- 203-203
- ISSN
- 2235-2988 (ISSN-L)
- PMC
- PMC6018158
- OpenAlex
- W2806665854
- Cited by
- 19
- References
- 36
- Citations by year
- 2026: 4 · 2025: 3 · 2024: 2 · 2022: 2 · 2021: 2 · 2020: 1 · 2019: 4 · 2018: 1
- FWCI
- 1.56
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Locations
- Frontiers in Cellular and Infection Microbiology (publishedVersion, cc-by)
- Europe PMC (PubMed Central) (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- Funders
- National Institutes of Health (1R15 AI119801-01)
- Topics
- Health Sciences › Medicine › Epidemiology › Fungal Infections and Studies
- Health Sciences › Medicine › Infectious Diseases › Antifungal resistance and susceptibility
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Fungal and yeast genetics research
- Keywords
- Cryptococcus neoformans, fluconazole, heteroresistance, drug resistance, phenotypic heterogeneity, colony size, single-cell responses, ergosterol, plasma membrane, aneuploidy, cryptococcosis, antifungal therapy
- MeSH terms
- Biological Variation, Population, Antifungal Agents, Cell Membrane, Cryptococcosis, Cryptococcus neoformans, Ergosterol, Humans, Microbial Sensitivity Tests, Fluconazole, Drug Resistance, Fungal, Single-Cell Analysis, Optical Imaging
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