Selenium (Se) is a nutritional component necessary for animal and plant development and reproduction. Selenium nanoparticles (SeNPs) have a high absorption rate during routine supplementation. In the current study, a cell-free extract of Aspergillus terreus was used as a reducing and stabilizing agent in the synthesis of SeNPs using a green and eco-friendly method. The mycosynthesized SeNPs were characterized by UV-visible spectrophotometry, Fourier transform infrared spectroscopy, X-ray diffraction spectroscopy, dynamic light scattering, transmission electron microscopy, and scanning electron microscopy. The results of the characterization process showed that the mycosynthesized SeNPs had spherical shapes and sizes less than 100 nm. Results showed that mycosynthesized SeNPs exhibited promising antibacterial activity against both Gram-positive and Gram-negative bacteria where inhibition zones were 14, 20, 16, and 13 mm toward S. haemolyticus, S. aureus, E. coli, and K. pneumoniae, respectively. However, it had weak antifungal activity against C. albicans, where the inhibition zone was 12 mm. The efficacy of mycosynthesized SeNPs for the decolorization of malachite green dye was investigated. Results illustrated that SeNPs exhibited rapid biodegradation of malachite green dye, reaching up to 89% after 240 min. In conclusion, SeNPs were successfully biosynthesized using A. terreus and demonstrated both antimicrobial and photocatalytic activities.