The antimicrobial effect of three Achillea sp. (A. fragrantissima, A. biebersteinni and A. santolina) on five bacteria (Staphylococcus aureus,Pseudomonas aeruginosa, Klebsiella sp. Escherichia coli and Enterobacter sp.) was investigated. Also, the genetic variation among the Achillea sp. was determined. The three Achillea sp. showed different antimicrobial activity in different extract amounts (50, 100, 150 and 200 mg mL-1) against the five bacteria. A. fragrantissima inhibited the growth of S. aureus at all used extract amounts. A. biebersteinni inhibited the growth of all tested bacteria except P. aeruginosa. The extract of A. santolina inhibited the growth of S. aureus at 150 and 200 mg mL-1, the growth of Klebsiella sp. at extract amounts 150 and 200 mg mL-1 and the growth of Enterobacter sp. at the four extract amounts. The three Achillea sp. revealed no activity against P. aeruginosa. Based on variation of RAPD patterns obtained with four primers (B-1, B-10, B-15 and B-18), the similarity between A. fragrantissima and A. santolina was 35%. The UPGMA-based (unweighted pair group method of arithmetic means) dendogram was constructed between the two species by using the similarity matrix results.