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In the present study, 134 morphologically distinct actinobacteria isolates were obtained from soil samples from 10 different localities in the Saudi Arabian desert. The preliminary screening revealed that 16 of these isolates possessed antimicrobial activity. One isolate, which was identified as Streptomyces sp. DA3-7, possessed broadspectrum antimicrobial activity against both gram-positive and gram-negative bacteria, as well as against fungi, and modified nutrient glucose medium was suitable for Streptomyces sp. DA3-7 to produce extracellular metabolites. The ethyl acetate extract of Streptomyces sp. DA3-7 exhibited antimicrobial activity against Enterococcus faecalis and Salmonella typhlmurium, with minimum inhibitory concentrations of 78 and 156 mu g/mL, respectively, as well as strong cytotoxicity (24 h IC50 85 mu g/mL) against MCF-7 human breast adenocarcinoma cells. The active compound was separated, purified, and identified as Pyridine-2,5-diacetamide (C9H11N3O2 + H+, 194.21), which possessed a lowest minimum inhibitory concentration (31.25 mu g/mL) against both Escherichia coli and Cryptococcus neoformans. The antimicrobial activities of this novel compound are reported here for the first time.