The taxonomic position of one soil isolate 13-12(50)^T was clarified by a polyphasic study. The organism showed a combination of chemotaxonomic and morphological properties typical of the genus Actinomadura. It formed a distinct phyletic line in the Actinomadura 16S rRNA gene tree and was closely associated with Actinomadura mexicana (sequence similarity 99.5%), Actinomadura glauciflava, Actinomadura citrea (sequence similarity 99.4%) and Actinomadura madurae (sequence similarity 99.2%). The result of DNA-DNA hybridizations between 13-12(50)^T and Actinomadura mexicana was 49.9%. On the basis of the phenotypic, chemotaxonomic and genotypic properties, the isolate was differentiated from its closest phylogenetic relatives. It is proposed that the organism be classified as a novel species of the genus Actinomadura. The name proposed for the new taxa are Actinomadura maheshkhaliensis sp. nov. [(13-12(50)^T=JCM 13934^T =MTCC 8055^T].