Stevia rebaudiana Bertoni, is an ancient perennial shrub of South America, with great potential as a crop for the production of high-power natural sweetener. Due to its chemical composition and content of the phytochemical compounds, it is suitable as a raw material for the extraction and production of functional foods. Diterpene glycosides component of low-calorie sweetener is approximately 300 times sweeter than sucrose. In this study, the antimicrobial properties of Stevia rebaudiana Bertoni extract on pathogenic and food spoiling bacteria were tested by the disk diffusion method. The effect of Stevia rebaudiana extract on gram-positive and gram-negative bacteria were examined and on Bacillus cereus, Lactobacillus Plantarum, Leuconostoc mesenteroides, Salmonella typhimurium and Salmonella enterica exhibited inhibitory and germicidal effect, but the effects on Listeria monocytogenes was lower than the rest of organisms. The Minimum Inhibitory Concentrations (MIC) and Minimum Bacterial Concentrations (MBC) of Stevia extract was 1250 and 10000-1250 mg /ml respectively. The results showed that Stevia extract can prevent the growth of food pathogens and spoilage bacteria. Therefor we can use it as a natural sweetener and preservative in food products.