A novel actinomycete, designated as strain OS1-43T, was isolated from the rhizosphere soil of a cactus (Opuntia ficus-indica) in Mara Island, Jeju, Republic of Korea and the taxonomic position was unraveled by a polyphasic approach. A 16S rRNA gene-based phylogenetic analysis showed that strain OS1-43T belonged to the genus Actinomadura and was closely associated with Actinomadura xylanilytica BK147T. Morphological and chemotaxonomic characteristics supported the affiliation of strain OS1-43T to the genus Actinomadura. The calculation of 16S rRNA gene sequence similarity showed that strain OS1-43T revealed the highest sequence identity with Actinomadura violacea LCR2-06T (99.1%), followed by Actinomadura xylanilytica BK147T (98.9%). Core gene-based phylogenomic tree revealed that strain OS1-43T formed an independent lineage within the genus Actinomadura and well separated from representatives of this genus. The genome of strain OS1-43T revealed the highest OrthoANIu and dDDH values with Actinomadura montaniterrae CYP1-1BT (90.1 and 40%, respectively), followed by Actinomadura physcomitrii LD22T (89.8 and 38.7%, respectively), Actinomadura nitritigenes L46T (89.8 and 38.6%, respectively), and Actinomadura violacea LCR2-06T (89.7 and 38.1%, respectively), albeit that the former three type strains showed low 16S rRNA gene sequence similarities (97.5-98.0%) with strain OS1-43T. Based on the results of phenotypic and phylogenetic analyses, and the values of overall genomic relatedness indices, strain OS1-43T (= KACC 19750T = NBRC 114686T) represents a new species of the genus Actinomadura, for which the name Actinomadura opuntiae sp. nov. is proposed. Also, the results of genome analysis in addition to similar phenotypic features support the reclassification of Actinomadura namibiensis Wink et al. 2003 as a later heterotypic synonym of Actinomadura kijaniata Horan and Brodsky 1982.
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