ZIRCONIA nanoparticles (ZrNPs) were efficiently biosynthesized by Fusarium oxysporum. The formation of ZrNPs was characterized by UV-visible spectral analysis, transmission electron microscopy, selected area diffraction pattern analysis, scanning electron microscopy, energy-dispersive X-ray analysis, and FT-IR spectroscopy. The results identified the generation of ZrNPs by the transformation of zirconia cations in the solution into zirconium dioxide in the nanosized particles with narrow size distribution. ZrNPs had moderate cytotoxic effects on HePG-2 (IC50= 32.38μg/mL), MCF-7 (IC50= 47.19μg/mL), PC3 (IC50= 43.50μg/mL), HeP2 (IC50= 38.23μg/mL), and HeLa (IC50= 33.62μg/mL) cell lines, complemented by weak cytotoxicity in HCT-116 (IC50= 58.13μg/mL) and normal WI-38 cell lines (IC50= 58.51μg/mL). Finally, the antimicrobial activity of ZrNPs was evaluated, in which the results indicated a minimum inhibitory concentration value at 50 μg/mL in the tests against K. pneumoniae, E. coli, and C. albicans, whereas the MIC value was recorded at 25μg/mL against S. aureus.