Fonsecaea pedrosoi, the main agent that causes chromoblastomycosis (CBM) in humans, is a filamentous fungus with a worldwide distribution. CBM is prevalent in tropical and subtropical regions, with high incidence in Africa and Latin America. The biology, virulence factors and mechanisms of pathogenesis of F. pedrosoi are still poor understood and for this reason studies involving the mechanisms this fungal pathogen uses to stablish the infection are essential for the improving the knowledge about the disease and  potentially develop new antifungal therapies. Recently our group described two genetic transformation methods to facilitate the gene function studies in this fungus. We use biolistic transformation to knock out F. pedrosoi ureA gene (coding for Urease) by homologous recombination and gene replacement. The main objective of this work was to characterize F. pedrosoi urease and evaluate its potential role on virulence of this fungal pathogen.For deletion of ureA, the cassette for homologous recombination was construct by Double Joint PCR technique and genetically transformed by biolistic transformation on conidia of F. pedrosoi (CBS 271.37 strain - ATCC 18658). Transfomants were selected by Hygromycin B, checked for mitotically stability and the insertion of the selection marker was confirmed by PCR. The ability of ΔureA to produce urease was evaluated on Christensen's Urea Agar and by in vitro spectrophotometric assay. We also checked if the mutant could use different nitrogen sources on YNB agar. Other phenotypes were tested as the growth at high temperature, at acid and alkali- pH, in osmotic, oxidative, nitrosative and cell wall stress conditions. The ΔureA tests for virulence in vitro are under evaluation. This work describes, for the first time, the literature to date, the construction and obtaining of a F. pedrosoi mutant by homologous recombination and gene deletion. The gene encoding the enzyme urease is well characterized in other fungi such as Cryptococcus sp. and Coccidioides posadasii as a factor of fungal virulence. In F. pedrosoi the absence of the ureA gene was confirmed by the inability of the mutant ΔureA to produce urease in Christensen's Urea Agar. The mutant was able to grow in culture medium with different nitrogen sources but showed a growth defect when urea was the only source of nitrogen. No growth defects were observed in the stress conditions evaluated. In vitro virulence experiments are underway to date. For the first time in literature to date, a knock out mutant was constructed in F. pedrosoi, which is one of the main causative agent of CBM. The ΔureA is under phenotypic characterization and the data generated by this work will potential identify the role of urease on F. pedrosoi virulence traits. This work opens new perspectives for fungal pathogenesis studies by gene deletion in this organism. Financial support: Fundação de Apoio à Pesquisa do Distrito Federal – FAP-DF, Brazil.

Functional characterization of Urease (ureA) in Fonsecaea pedrosoi, the causative agent of Chromoblastomycosis