Phylogeny and molecular taxonomy of the Bacillus subtilis species complex and description of Bacillus subtilis subsp. inaquosorum subsp. nov.

Abstract

Phylogeny and molecular taxonomy of the Bacillus subtilis species complex and description of Bacillus subtilis subsp. inaquosorum subsp. nov. The Bacillus subtilis species complex is a tight assemblage of closely related species. For many years, it has been recognized that these species cannot be differentiated on the basis of phenotypic characteristics. Recently, it has been shown that phylogenetic analysis of the 16S rRNA gene also fails to differentiate species within the complex due to the highly conserved nature of the gene, yet DNA-DNA hybridization values fall well below 70 % for the same species comparisons. As a complementary approach, we propose that phylogenetic analysis of multiple protein-coding loci can be used as a means to detect and differentiate novel Bacillus taxa. Indeed, our phylogenetic analyses revealed the existence of a previously unknown group of strains closely related to, but distinct from, Bacillus subtilis subsp. spizizenii. Results of matrix-assisted laser desorption ionization-time of flight mass spectrometry analyses revealed that the group produces a novel surfactin-like lipopeptide with mass m/z 1120.8 that is not produced by the other currently recognized subspecies. In addition, the group displayed differences in the total cellular content of the fatty acids C 16 : 0 and iso-C 17 : 1 v10c that distinguish it from the closely related B. subtilis subsp. spizizenii. Consequently, the correlation of these novel phenotypic traits with the phylogenetic distinctiveness of this previously unknown subspecies group showed that phylogenetic analysis of multiple protein-coding loci can be used as a means to detect and differentiate novel Bacillus taxa. Therefore, we propose that this new group should be recognized as representing a novel taxon, Bacillus subtilis subsp. inaquosorum subsp. nov., with the type strain NRRL B-23052 T (5KCTC 13429 T 5BGSC 3A28 T ). INTRODUCTIONThe genus Bacillus is composed of rod-shaped, endosporeforming bacteria that are members of the phylum Firmicutes. Owing largely to the fact that they are common inhabitants of soil and aquatic sediment, species within the genus are widespread in nature and are found in virtually every environment. While their main roles appear to involve carbon and nitrogen cycling, some species are wellknown human and livestock pathogens (e.g. Bacillus anthracis and Bacillus cereus) and insect pathogens (e.g. Bacillus thuringiensis). However, the overwhelming majority of Bacillus species are non-pathogenic. Because of this fact, many have been exploited for biotechnological andAbbreviations: FAME, fatty acid methyl ester; MALDI-TOF MS, matrixassisted laser desorption ionization-time of flight mass spectrometry; NJ, neighbour-joining; ML, maximum-likelihood; MP, maximumparsimony.The GenBank/EMBL/DDBJ accession numbers for the sequences generated in this study are EU138452-EU138865.A maximum-parsimony cladogram of the Bacillus subtilis species complex and tables showing the mean values of synonymous and non-synonymous substitutions per s...

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