Abstract: The characteristic of average nucleotide identity (ANI) based on the whole genomes from Bacillus species in Bacillus-like genus was analyzed using the soft of Jspecies.The results showed that there have a species specificity based on the ANI distribution and which correlate with the relationship of taxonomic and evolutionary affinities.The ANI values computed from different Bacillus-like genus were below a 65% identity or above a 50% different in orthologous genes identity.The ANI values from different Bacillus species were below 94% with a wide range from 63.77 to 93.81and the most of them ranging from 64.77% to 68.4% with the weighted average of 70.12%.The ANI value between subspecies was mainly ranged from 90% to 96%.Our results suggest that the ANI criterion of the identification for genus, species and subspecies within Bacillus are 50%-65%, 65%-90% and 90%-96%, respectively.Furthermore, Tetra regression index was correlated with the ANI value and displayed the species specificity.Regression fitting analysis showed that the equation of y=271-573.56x+399.65x2 with the correlation coefficient of 0.981 2occurred between the Tetra and the ANI values above a identity of 70 were positive correlated with Tetra regression index with the equation of y=178.58x-81.521and the correlation coefficient of 0.876 7.