Medicinal plants are potential of antimicrobial compounds. The present study deals with the antibacterial activity of methanolic extract of Thymus daenensis. Aerial parts of the plant were collected from Alvand mountainside (Hamadan, Iran) in May 2005, air-dried and extracted by methanol. The dried extract was redissolved in methanol to make a 100 mg/ml solution and then filtered. Antibacterial activity of the extract was evaluated against various Gram-positive and Gram-negatives bacteria using disk diffusion technique. Blank paper disks were loaded with 40 microl of the methanol solution and then dried up. The impregnated disks were placed on Mueller-Hinton agar inoculated with bacterial suspension equal to 0.5 McFarland. The extract inhibited the growth Gram-positive bacteria, i.e., Staphylococcus aureus, Micrococcus luteus, Entrococcus faecalis, Streptococcus pyogenes, but it showed no activity against Gram-negative bacteria. The most significant effect was seen against S.aureus including MRSA, which are important nosocomial pathogens. MIC90 of the extract was determined against Gram-positive bacteria (3.12 mg/ml) and 11 MRSA strain (1.56 mg/ml).