A polyphasic study was performed on 10 soil isolates of thermophilic denitrifying Bacillus strains from different geographical areas. The presence of two main characteristic bands following amplification of the internal transcribed spacer (ITS) region of rrn operons suggests a close relatedness to 'Bacillus thermodenitrificans'. The isolates cluster around two strains of 'B. thermodenitrificans' in riboprint and fatty acid analyses, though differences occur at the strain level. Subsequent DNA-DNA reassociation studies including the 10 isolates, 'B. thermodenitrificans' DSM 465T and DSM 466, and Bacillus stearothermophilus ATCC 12980T and Bacillus thermoleovorans ATCC 43513T revealed such a high level of genomic relatedness between the isolates and the DSM strains (> 73% similarity) that they must be considered strains of the same taxon. The degree of DNA-DNA similarity between the 12 strains of 'B. thermodenitrificans' and the type strains of the other two phylogenetically neighbouring Bacillus species was significantly lower (21-43% similarity). Based upon phylogenetic, chemotaxonomic and phenotypic evidence, the designation of B. thermodenitrificans sp. nov., nom. rev. is proposed. The type strain of B. thermodenitrificans is DSM 465T.