The safety of rebaudioside A, produced fermentatively by Yarrowia lipolytica encoding the Stevia rebaudiana metabolic pathway (fermentative Reb A), is based on several elements: first, the safety of steviol glycosides has been extensively evaluated and an acceptable daily intake has been defined; second, the use of Y. lipolytica, an avirulent yeast naturally found in foods and used for multiple applications; and third the high purity of fermentative Reb A and its compliance with internationally defined specifications. A bacterial reverse mutation assay and an in vitro micronucleus test conducted with fermentative Reb A provide evidence for its absence of mutagenicity, clastogenicity and aneugenicity. The oral administration of fermentative Reb A to Sprague-Dawley rats for at least 91 days did not lead to any adverse effects at consumption levels up to 2057 mg/kg bw/day for males and 2023 mg/kg bw/day for females, which were concluded to be the No Observed Adverse Effect Levels. The results were consistent with outcomes of previous studies conducted with plant-derived rebaudioside A, suggesting similar safety profiles for fermentative and plant-derived rebaudioside A. The results of the toxicity studies reported here support the safety of rebaudioside A produced fermentatively from Y. lipolytica, as a general purpose sweetener.