Two thermophilic strains, designated 607T and 606b, were isolated from a compost pile in Japan. The novel strains were Gram-positive, aerobic, spore-forming rods. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strains 607T and 606b were closely related to Bacillus naganoensis (94.0-94.1% similarity) and separated from clusters of the related genera Bacillus (<91.9%) and Sporolactobacillus (91.0-92.5%). In addition, some chemotaxonomic and physiological characteristics of strains 607T and 606b differed from those of B. naganoensis and the two related genera. Several differences in physiological characteristics and 16S-23S rRNA gene internal transcribed spacer region nucleotide sequences were observed between strains 607T and 606b; however, DNA-DNA hybridization indicated that these two strains belonged to the same species. From these results, it is proposed that strains 607T and 606b represent the type species of a new genus, Tuberibacillus calidus gen. nov., sp. nov., with strain 607T (=JCM 13397T=DSM 17572T) as the type strain. In addition, the results of phylogenetic analyses, as well as chemotaxonomic and physiological characterization, indicated that B. naganoensis and Bacillus laevolacticus did not belong to the genus Bacillus. Based on these results, it is proposed that B. naganoensis and B. laevolacticus should be transferred to Pullulanibacillus naganoensis gen. nov., comb. nov. and Sporolactobacillus laevolacticus comb. nov., respectively.