Objective: To isolate and characterize novel actinomycetes and to evaluate their antibacterial activity against drug-resistant pathogenic bacteriaMethods: In the present study, 19 soil samples were collected from different localities of Ad-Dawadmi, Saudi Arabia. Actinomycetes were isolated from these samples using serial dilution and plating method on Actinomycetes isolation agar supplemented with nalidixic acid and actidione to inhibit bacteria and fungi. Crude extracts of potential actinomycetes were produced by submerged fermentation. The antimicrobial activity of crude extracts of actinomycetes was tested against different bacteria using the agar well diffusion method. Characterization of the isolates was done by morphological, physiological and biochemical methods.Results: A total of... 9 (47%) isolates of actinomycetes were isolated from 19 different soil samples tested. Among them, 4 (44%) isolates confirmed as Streptomyces sp. showed potential antimicrobial activity against one or more test organisms. Crude extracts were made from these 4 actinomycetes isolates(DOM1, DOM3, DP3, DP4)and tested for their antibacterial activities against 4 different clinical bacterial strains (Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Enterococcus). Crude extract from DP3 isolate showed highest antibacterial activity against all the four test organisms (28 mm, 21 mm, 20 mm and 18 mm) respectively and DP4 showed lowest antibacterial activity against all the four test organisms (14 mm, 12 mm, 0 mm, 6 mm) respectively. The highest zone of inhibition was shown by DP3 against Staphylococcus aureus (28 mm) and Escherichia coli was resistant for DP4. Most of the Inhibition zones produced by crude extracts showed significant differences when compared with control, tested against test organisms (P<0.05). Inhibition zones produced by DP3 and DOM1 against Staphylococcus aureus were 28 mm and 23 mm, respectively which were strong active when compared with control Ciprofloxacin (18 mm).Conclusion: Further studies for purification of bioactive metabolites and molecular characterization analysis of isolated Streptomyces sp. are in progress which would be helpful in discovering novel compounds of commercial value.