Patients with chromoblastomycosis (CBM) suffer chronic tissue lesions that are hard to treat. Considering that biofilm is the main growth lifestyle of several pathogens and it is involved with both virulence and resistance to antimicrobial drugs, we have investigated the ability of CBM fungi to produce this complex, organized and multicellular structure. Fonsecaea pedrosoi and Phialophora verrucosa conidial cells were able to adhere on a polystyrene abiotic substrate, differentiate into hyphae and produce a robust viable biomass containing extracellular matrix. Confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM) showed the tridimensional architecture of the mature biofilms, revealing a dense network of interconnected hyphae, inner channels and amorphous extracellular polymeric material. Interestingly, the co-culture of each fungus with THP-1 macrophage cells, used as a biotic substrate, induced the formation of a mycelial trap covering and damaging the macrophages. In addition, the biofilm-forming cells of F. pedrosoi and P. verrucosa were more resistant to the conventional antifungal drugs than the planktonic-growing conidial cells. The efflux pump activities of P. verrucosa and F. pedrosoi biofilms were significantly higher than those measured in conidia. Taken together, the data pointed out the biofilm formation by CBM fungi and brought up a discussion of the relevance of studies about their antifungal resistance mechanisms.