Phenotype and molecular phylogenetic analysis of Bacillus subtilis MJ01 using different methods based on comparative genomics

A gram-positive bacterium Bacillus subtilis MJ01 strain was isolated from crude oil contaminated soil, Pazanan oil field in Khuzestan province, in order to produced surfactin and applied as microorganism enhanced oil recovery(MEOR). The primary research on this strain revealed that chemical and characteristics its surfactin was different among other surfactin variants. In the next step, whole genome sequencing of MJ01 strain was designed due to better understanding about variation of this strain and other close relative bacteria. The 16SrRNA genes analysis categorized MJ01 strain in Bacillus subtilis cluster close to spizizenii TU-B-10 strain in the same group. Multi-locus sequence typing(MLST) results was confirmed four of seven housekeeping genes in B. subtilis family were new mutant loci and one of them was inverted-competently in MJ01 genome. These results were shown that MJ01 MLST profile template have high similarity with spizizenii BGSC3A17 strain. ANIb (Average Nucleotide Identity based on BLAST) analysis with 12 close related genomes demonstrated that this strain has high identity to three spizizenii subspecies strains include TU-B-10 (99.13), NRS 231(96.52) and W23(96.52). Also based on ANI analysis revealed that it was closely related with Jeotglibacillus marinus DSM 1297. Non-ribosomal lipoprotein prediction online tools were used for located surfactin operon and flanking regions in MJ01 genome. Comparison of this operon and flanking region with other close related B. subtilis complete genome indicated that MJ01 placed in the spizizenii subspecies group. In summary, integrative phylogeny analysis and DNA homology using comparative genomics approaches supported that MJ01 to be included as a member of spizizenii subspecies group, although MJ01 have some specific strain differences.