Application of RE-SNPs genotyping in phylogenic studies of Bacillus anthracis in Iran

Although the single nucleotide polymorphisms (SNPs) genotyping is now a standard method in the evolutionary and epidemiological studies of anthrax, but application of this technique is not possible in low-budget laboratories due to its sequence-based nature. Molecular epidemiologic studies of Bacillus anthracis play an important role in identifying and differentiating different strains during the bioterrorism-related outbreaks. The aim of this study was to investigate the application of restriction enzymes in Van Ert SNPs typing. This applied study was conducted in research & development laboratory of aerobic veterinary bacterial vaccines department in Razi Vaccine and Serum Research Institute during 2014. The corresponding genomic segments of the B. anthracis Sterne 34F2 strain were searched in order to identify appropriate restriction enzymes to digest SNPs loci. The accuracy of the results was confirmed by sequencing of the PCR products. Of 13 studied loci, 3 were found to be available for restriction enzymes due to their target SNPs. Feasibility of this technique was examined through enzymatic treatment of a 560 bp-long PCR product including the A.Br004 SNP of the standard strain by BsmI. The PCR product was digested and was divided into 224 bp and 336 bp long fragments. With regards to the results, it seems that the RE-SNPs typing as a simple method is appropriate for the van Ert genotyping of B. anthracis and resolves the need for expensive sequencing.