SRI International (SRI) has screened a large number of compounds to discover lead compounds with broad spectrum activity against biodefense organisms. Many of these compounds were nitro-heterocycles that contain sulfur or oxygen. The compounds were tested in a broth microdilution method against BSL-3 level biodefense organisms Bacillus anthracis, Yersinia pestis, and Francisella tularensis (Category A strains), and Brucella abortus, Burkholderia mallei, and Burkholderia pseudomallei (Category B strains). SRI has found 36 compounds eliciting broad spectrum activity against all six strains. Furthermore, 316 compounds have revealed stronger or similar activity to those of well-established antibiotics such as ciprofloxacin, gentamicin, and tetracycline. SRI also tested a number of compounds against the intracellular bacterium, Coxiella burnetii (Category B strain). Only one of these compounds exhibited some antimicrobial activity at 17 µg/ml, the highest dose level used. However, this compound was slightly toxic to the host Vero cells. The results from the assays contribute to information about the structure-activity relationships (SAR) of the various classes of compounds. This information provides biological and chemical profiles for these compound classes, and is therefore vital to the optimization of producing an effective antibiotic to advance in the Investigational New Drug (IND) process. Financial support was provided by the National Institute of Allergy and Infectious Diseases (Contract Number: N01-AI-40005).