Clearing the Fungal FoG: Perseverance, a property distinct from resistance, is associated with clinical persistence

Drug resistance, defined by the minimal inhibitory concentration (MIC), often does not predict whether fungal infections will respond to drug treatment in the clinic. Here we define and quantify an antifungal response, termed perseverance, that correlates with clinical outcomes in patients treated with fluconazole, the most widely used antifungal drug. Perseverance is defined by the ability of fungal cells to grow at drug concentrations above the MIC, and is measured either as the fraction of growth (FoG) in drug disk diffusion assays, or as the degree of supra-MIC growth (SMG) in broth microdilution assays. Perseverance is distinct from resistance and is related to the proportion of cells that form colonies at supra-MIC drug concentrations. Higher perseverance correlates with shorter lag times at high drug concentrations, highlighting a critical distinction from tolerance in bacterial populations. Importantly, several adjuvant drugs, including inhibitors of Hsp90 and calcineurin, eliminate perseverance without altering the MIC. We also find that perseverance and resistance are sensitive to mutations in different pathways, underscoring the distinct nature of the two phenomena. Analysis of isolates recovered from clinically persistent or non-persistent infections in immunocompetent patients reveals that persistent isolates show higher levels of perseverance than isolates cleared by antifungals. Thus, perseverance is an intrinsic property of fungal isolates that correlates with the success or failure of treatment in the clinic, and may provide a useful parameter for predicting clinical persistence and choosing appropriate antifungal therapies.