Characterization and microbial growth activity of polyvinyl alcohol (PVA) nanofibers (NFs) produced by an electrospinning technique containing different amounts of silver nanoparticles (AgNPs) was investigated. AgNPs were synthesized by chemical reduction of silver nitrate (AgNO3) (in different weight ratios) in PVA solution followed by electrospinning to produce PVA/AgNPs NFs. The properties of PVA/AgNPs NFs were studied as a function of the silver content. The morphology and distribution of NFs and AgNPs were characterized by transmission and scanning electron microscopy. The size of NFs was 135-160 nm, and the size of AgNPs was 15-27 nm. TEM images clearly showed that spherical AgNPs were homogeneously integrated in PVA NFs. The microbial activity of NFs containing different amounts of AgNPs was examined against both Gram-negative (Pseudomonas aeruginosa and Escherichia coli) and Gram-positive (Bacillus megaterium and Staphylococcus aureus) bacteria and against Candida albicans. PVA NFs containing AgNPs exhibited increasing inhibitory effects against microbial pathogenic strains with increasing AgNPs amounts. The results suggest that PVA/AgNPs NFs could be used for wound dressing components, protective coatings, biomedical devices, and water purification.