Nosocomial infections are the major concern in the world. Today, with the emergence of antibiotic resistance among pathogenic bacteria, treatment of this category of infectious diseases is facing numerous problems. The present study investigation of antibacterial activities selenium and selenium nano-particles (synthesized by Lactobacillus acidophilus). Lactobacillus acidophilus and Lactobacillus plantarum were cultured in (MRS) broth medium. The 3-5% of inoculums culture and 10 ml of stock solution of selenium (NaHSeO3) separately were added into two Erlenmeyer flasks containing (MRS) broth medium. The nano-particles were analyzed using scanning electron microscopy (SEM). In this study, the urine, blood, sputum and wound specimens of patients were collected. The isolates were identified and confirmed using the standard microbiological method and 50 antibiotic resistance strains from 436 samples were selected (80% Escherichia coli and 20% Acinetobacter). Antibacterial activities selenium and selenium nanoparticles on isolated bacterial strains were evaluated by disk diffusion and serial broth dilution methods to determine (MIC) and (MBC). The SEM results showed that L.acidophilus nano-particles are smaller than of L. plantarum, therefore it would be preferred for further investigation. The antibacterial activities of the selenium and selenium nano-particles demonstrated to be effective on 32% and 56% of the strains studied, respectively.