Evolution of strategies for biodegradation of synthetic organic pollutants relies on recruitment of genes from catabolic pathways for natural compounds. Investigation of metabolic diversity in nature can provide insight into biochemical strategies that could be recruited for bioremediation of pollutants. As part of a search for novel metabolic diversity we isolated soil bacteria able to degrade 2-nitroimidazole (2NI) and 5-nitroanthranilic acid (5NAA), and determined the biochemistry and molecular biology of their biodegradation pathways. 2NI and its analogs are increasingly used as prodrugs for the treatment of both tuberculosis and cancer. The biodegradation of 2NI by a soil Mycobacterium sp. is initiated by an unusual hydrolytic denitration. The reaction is catalyzed by a novel nitrohydrolase with a divergent sequence and represents the discovery of a previously unreported drug resistance mechanism in soil prior to its identification in clinical situations. 5NAA is the starting material for various nitroaromatic compounds and dyes. The biodegradation pathway of 5NAA is initiated by an unusual hydrolytic deamination. The corresponding gene is very distantly related to biochemically characterized genes in the NCBI database. The nitro group of 5NAA is eliminated as nitrite during the spontaneous formation of lactones from a ring fission product, a previously unreported mechanism. Degradation pathways of 5NAA and 2NI serve as precedents for those of nitroaniline and nitroimidazole pollutants. The work supports the hypothesis that the study of the metabolism of natural organic compounds selected on the basis of unusual structural features and ecological roles can reveal new metabolic diversity.