The taxonomic positions of three novel strains isolated from soil, driftwood and sediment samples collected in Japan were investigated based on the results of chemotaxonomic, phenotypic and genotypic characteristics. The strains that we examined were Gram-positive, catalase-positive bacteria with L-ornithine as a diagnostic diamino acid of peptidoglycan. The acyl type of peptidoglycan was N-glycolyl. The major menaquinones were MK-11, -12, -13 and/or -14. Mycolic acids were not detected. The G+C content of the DNA was 68 to 70 mol%. These morphological and chemotaxonomical characters and comparative 16S rDNA analysis of the three isolated strains revealed that they belong to the genus Microbacterium. DNA-DNA relatedness data revealed that the three isolates are three new species of the genus Microbacterium. On the basis of the polyphasic evidence, the isolates should be classified as novel species of the genus Microbacterium: Microbacterium awajiense sp. nov., Microbacterium fluvii sp. nov. and Microbacterium pygmaeum sp. nov. with the type strains YM13-414^T (=MBIC08276^T, DSM 18907^T), YSL3-15^T (=MBIC08277^T, DSM 18908^T) and KV-490^T (=NRRL B-24469^T, NBRC 101800^T), respectively.

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Partial nucleotide sequences of the gyrB gene were determined for members of the genus Nocardiopsis and subjected to phylogenetic analysis. Interspecies DNA similarity values of partial gyrB gene sequences ranged from 79.9 to 99.7% among the Nocardiopsis species. The average similarity of the gyrB gene (87.7%) was significantly less than that of the 16S rRNA gene (96.65%), indicating a high discriminatory power of the gyrB gene. The topology of neighbor-joining, maximum likelihood and maximum parsimony phylogenetic trees based on nucleic acid sequences and protein sequences of gyrB gene were reconstructed. Compared with the 16S rRNA gene dendrogram, the gyrB gene provided more information for understanding the genetic relationships among Nocardiopsis species. Using the information derived from 16S rRNA and gyrB genes, a more comprehensive understanding of the evolutionary history of Nocardiopsis species was obtained.

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