Background: Microemulsions (MEs), which consist of oil, water, surfactants, and cosurfactants, have recently generated considerable interest as antimicrobial agents. Objectives: To determine the antifungal and antiviral activities of three ME formulations (MEa, MEb, and MEc) that differ in their hydrophilicity. Methods:The ME formulas were produced by mixing different fractions of Tween 80, Span 20, ethanol, oil, isopropyl myristate, and distilled water. The antifungal activity of the ME formulas against Aspergillus niger, A. flavus, Bacillus, Candida albicans, and C. glabrata were determined by the solid medium diffusion cytotoxicity test against the mitochondria, measuring the minimum inhibitory concentration, dry biomass, and leakage of potassium, and characterizing the cell morphology. The antiviral activities of the ME formulas against the herpes simplex virus type 2 (HSV-2) were determined using the cytopathic effect assay. Results: Significant antimicrobial activities were recorded against A. niger and herpes simplex virus type 2 (HSV-2) when treated