Urosepsis by multidrug-resistant Nakaseomyces glabratus with non-functional Erg3 and Erg11—do collateral sensitivity and a unique mode of action make nitroxoline a viable UTI antifungal?

The emergence of multidrug-resistant fungal pathogens from urinary tract infections (UTIs) poses a growing challenge in clinical settings. Here, we report a case of a complicated UTI caused by Nakaseomyces glabratus ( Candida glabrata ) that progressed to urosepsis, leading to the emergence of an isolate carrying simultaneous loss-of-function mutations in ERG3 and ERG11, and abrogated ergosterol biosynthesis. Together with a missense mutation in FUR1 —likely responsible for 5-fluorocytosine resistance—this constellation confers resistance to all viable UTI antifungals: azoles, amphotericin B, and flucytosine. Engineered ERG3 Δ + ERG11 Δ strains recapitulated this multidrug resistance and revealed profound fitness costs that come with it, challenging the assumption that high-cost mutations are unlikely to persist during infection. Among fitness trade-offs, we detected collateral sensitivity to nitroxoline, a commonly used urinary tract antibiotic with potent antifungal activity and a unique mechanism of action. This study provides the first clinical evidence of an elusive mechanism of hyper-multidrug resistance in N. glabratus and highlights nitroxoline as a promising repurposing agent for treating multidrug-resistant fungal infections of the urinary tract. IMPORTANCE Evolutionary theory states that fitness determines survival. In a drug-treatment environment, resistance increases fitness, but it often comes at a cost, such as slower growth or reduced stress tolerance. If these costs are too severe, they can undermine virulence, making resistance unlikely to persist. Our study challenges this assumption. We describe the first clinical case of Nakaseomyces glabratus evolving multidrug resistance through loss-of-function mutations in ERG3 and ERG11 , despite severe fitness trade-offs. This case suggests that certain infection niches, such as the urinary tract, can provide conditions where even highly impaired yet resistant strains persist under strong antifungal pressure. Importantly, we show that this extreme resistance induces collateral sensitivity to nitroxoline, a urinary tract infection antibiotic with potent antifungal activity and a unique mechanism of action. These findings open promising therapeutic avenues to counter multidrug-resistant fungal infections of the urinary tract.