AbstractEach year, Candida albicans causes ~700,000 cases of invasive life-threatening
candidemia with a mortality rate of 40%. The opportunistic pathogen C. albicans is the fourth
most common cause of nosocomial infections. While neutrophils are the first line of defence
against invasive C. albicans, the yeast has developed several strategies to escape neutrophil
killing effectively. The project aims to identify C. albicans genes that are important for the
survival of neutrophil antifungal responses. These could then be exploited as future drug targets
to stop C. albicans immune evasion.
A high-throughput fluorescence-based killing assay was established and used to screen
384 conditional mutants from the GRACETM library, the largest publicly available C. albicans
mutant library. Neutrophil-like cells, PLB-985, were used in the initial screen. Any mutants
that were susceptible to PLB-985 killing were further validated with primary human
neutrophils. Validated mutants were analysed in a series of functional assays to identify
potential pathways and mechanisms involved in immune evasion.
25 out of 384 C. albicans mutants were found to be susceptible to PLB-985 killing.
After four further validations with neutrophils, 5 mutants were found to be vulnerable to
neutrophil killing. These mutants comprise brr2, taf7, noc3, tfb3 and smt3 with approximately
38%, 44%, 27%, 28% and 29% reduction in cell viability relative to the C. albicans WT
control, respectively. Functional assays further revealed that the five identified genes might
play essential roles in C. albicans immune evasion. For example, the noc3 mutant appeared to
be a weak inducer, and the tfb3 mutant appeared to be a strong inducer of reactive oxygen
species and neutrophil extracellular traps production. Therefore, these genes may represent
good therapeutic targets for candidemia.
|Date of Award||24 Sep 2021|
|Supervisor||Borko Amulic (Supervisor) & Stephanie Diezmann (Supervisor)|