Ludmila Gouveia-eufrasio, Gustavo José Cota de Freitas, Danielle L. da Silva, Iara Rinco Silva, Daniel Santana de Carvalho, Milton T. Drott, Bruna Carolina Teixeira Almeida, César da Silva Santana Moura, Beatriz Murta Rezende Moraes Ribeiro, Maria Fernanda Liphaus Almeida Negreli, Lucas Cecílio Vilar, Silvia Maria Cordeiro Werneck, Grace S. Tavares, Álan Natanael Pereira Gomes, Isabela Lima de Miranda, Mariana Guerra de Aguilar, Rossimiriam Pereira de Freitas, Luana Rossato, Rafael Wesley Bastos, Livia Kmetzsch, Júlia C.V. Reuwsaat, Isabela Costa César, Rafaela Salgado Ferreira, Nalu Teixeira de Aguiar Peres, Daniel Assis Santos
Antifungal resistance is considered a global health threat. However, enzymatic inactivation of antifungals, a common mechanism seen in antibacterial resistance, has not yet been described in fungi. From a One Health perspective, this study demonstrates that Cryptococcus deuterogattii and C. neoformans, the leading agents of cryptococcosis, enzymatically inactivate fluconazole. Agrochemicals induce the overexpression of genes that code for acetyltransferases, specifically GCN5 and NAT10. These enzymes catalyze the acetylation of fluconazole into O-acetyl-fluconazole. This metabolite is unable to properly bind to 14-α-demethylase, the azole target, abolishing the antifungal activity. GCN5 and NAT10 inhibitors constrained acetylation and restored fluconazole activity, highlighting their potential as therapeutic adjuvants. The same phenotype was observed in other fungal species, suggesting broader relevance. Furthermore, O-acetyl-fluconazole was also detected in cerebrospinal fluid from cryptococcal meningitis patients undergoing fluconazole treatment. These findings reveal a previously unrecognized antifungal resistance mechanism and suggest that environmental traits shape clinically relevant resistance through conserved enzymatic pathways.