Agaricus species are among the most important commercial mushrooms in the world. Thus far, antibacterial activities of several Agaricus spp. have been reported. However, no study was undertaken to evaluate antibacterial activities of Iranian wild Agaricus spp. In addition, the information about compounds conferring antibacterial activity in these species is scarce. The aim of this study was to determine the antibacterial activity of methanol-dichloromethane (1:1) extracts of several wild and cultivated species of Agaricus through a colorimetric micro-dilution method. In addition, several active fractions obtained from the crude extract of cultivated A. bisporus was assayed for antibacterial activity. The findings revealed that the mushroom crude extracts showed quantifiable inhibition only towards the Gram-positive bacteria. The cultivated A. bisporus significantly exhibited higher antibacterial activity than did the wild strain, even though no complete inhibition was observed. In addition, A. devoniensis and A. gennadii exhibited slight inhibitory effects against only Staphylococcus aureus. Six different fractions were then eluted from the crude extract of cultivated A. bisporus through a step wise gradient elution, of which two fractions exerted quantifiable antibacterial activities. As opposed to the crude extract, these two fractions significantly inhibited both the Gram-positive and the Gram-negative bacteria particularly E. coli at 8 mg/ml. These fractions might promisingly possess bioactive compounds and warrant further investigations to develop new antimicrobials obtained from A. bisporus.