Systematic Genetic Analysis of C. albicans CNS Infection

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PROJECT SUMMARY The central nervous system (CNS) is an important target organ for nearly every major human fungal pathogen including Cryptococcus, Coccidioides, Candida, Aspergillus, Histoplasma and others. In almost all examples, the fungus either colonizes or infects one organ system and then disseminates to the CNS. The most common initial site of infection is the lung as exemplified by the pathogenesis of Cryptococcus, Histoplasma, Coccidioides, and Aspergillus while Candida albicans is unique in that it is a commensal of the human GI tract that infects in the brain during invasive disease. Premature infants have long been recognized as the patient cohort most at-risk for C. albicans CNS disease, although recently it has been recognized that adults with mutations in the CARD9 innate immune pathway are also at high risk. Importantly, C. albicans meningoencephalitis contributes to the fact that, among infants who survive sepsis, those infected with C. albicans have much poorer neurodevelopmental outcomes than infants infected with bacteria. Thus, CNS disease is a clinically important sequelae of candidiasis. However, we know very little about the mechanisms that underlie the ability of C. albicans to infect the CNS. To address this gap in knowledge, we propose to utilize a new model of C. albicans CNS infection to screen collections of transcription factor (TF) and protein kinase (PK) mutants to identify key regulators of C. albicans neurotropism. We discovered that mice lacking the IL-1 receptor (IL1-R) develop dramatically increased brain burden during disseminated candidiasis relative to congenic WT mice. The increased CNS fungal burden in il1r-/- mice provides sufficiently robust dynamic range to allow the identification of C. albicans mutants with decreased CNS infectivity whereas standard WT mouse strains are only transiently infected by C. albicans at relatively low fungal burden. We, therefore, propose to use this novel model to perform a genetic screen to identify TFs and PKs that are required for C. albicans CNS infectivity (Aim 1). We will generate competitive fitness profiles of barcoded pools of TF mutants isolated from mouse brain tissue after both intravenous (IV) and intracranial (IC) inoculation routes. In Aim 2, we will characterize the effects of a focused set of TF and PK mutants by: 1) analyzing their interactions with on brain endothelial cells; 2) determining their effect on the transcription of virulence and environmentally responsive genes in vivo during CNS infection; and 3) determining their genetic interaction profiles with the other TFs and PKs. These experiments will provide the first characterization of the genetic and transcriptional requirements for C. albicans CNS infection and will set the stage for future, detailed mechanistic studies of C. albicans neurotropism.