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The traditional 16S rRNA sequence analysis and DNA-DNA hybridization experiment lack resolution power at the species level and below.However, in clinic practice and environmental studies there is an urgent need to distinguish strains at species level and below.The whole-genome-based and alignment-free CVTree approach provides the required resolution.We use the examples of the Shigella-Escherichia, Yersinia pestis-Yersinia pseudotuberculosis problems to demonstrate the potential application of the CVTree method .