Genome-Wide Single Nucleotide Polymorphism Typing Method for Identification of Bacillus anthracis Species and Strains among B. cereus Group Species

• Published in: Journal of Clinical Microbiology Vol. 48; no. 8; pp. 2821 - 2829
• Main Authors: Kuroda, Makoto, Serizawa, Masakuni, Okutani, Akiko, Sekizuka, Tsuyoshi, Banno, Satomi, Inoue, Satoshi
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.08.2010; American Society for Microbiology (ASM)
• Subjects: Bacillus anthracis; Bacillus anthracis - classification; Bacillus anthracis - genetics; Bacillus anthracis - isolation & purification; Bacillus cereus; Bacillus cereus - classification; Bacillus cereus - genetics; Bacillus cereus - isolation & purification; Bacterial Typing Techniques; Bacterial Typing Techniques - methods; Bacteriology; Biological and medical sciences; Chromosomes, Bacterial; DNA, Bacterial; DNA, Bacterial - genetics; Fundamental and applied biological sciences. Psychology; Genome, Bacterial; Genotype; Humans; Japan; Microbiology; Miscellaneous; Phylogeny; Plasmids; Polymerase Chain Reaction; Polymerase Chain Reaction - methods; Polymorphism, Single Nucleotide; Japan; Bacillus anthracis; Typing; Bacillales; Bacteria; Identification; Method; Single nucleotide polymorphism; Bacillaceae; Genome; Strain
• ISSN: 0095-1137; 1098-660X; 1098-660X
• DOI: 10.1128/jcm.00137-10

As an issue of biosecurity, species-specific genetic markers have been well characterized. However, Bacillus anthracis strain-specific information is currently not sufficient for traceability to identify the origin of the strain. By using genome-wide screening using short read mapping, we identified strain-specific single nucleotide polymorphisms (SNPs) among B. anthracis strains including Japanese isolates, and we further developed a simplified 80-tag SNP typing method for the primary investigation of traceability. These 80-tag SNPs were selected from 2,965 SNPs on the chromosome and the pXO1 and pXO2 plasmids from a total of 19 B. anthracis strains, including the available genome sequences of 17 strains in the GenBank database and 2 Japanese isolates that were sequenced in this study. Phylogenetic analysis based on 80-tag SNP typing showed a higher resolution power to discriminate 12 Japanese isolates rather than the 25 loci identified by multiple-locus variable-number tandem-repeat analysis (MLVA). In addition, the 80-tag PCR testing enabled the discrimination of B. anthracis from other B. cereus group species, helping to identify whether a suspected sample originates from the intentional release of a bioterrorism agent or environmental contamination with a virulent agent. In conclusion, 80-tag SNP typing can be a rapid and sufficient test for the primary investigation of strain origin. Subsequent whole-genome sequencing will reveal apparent strain-specific genetic markers for traceability of strains following an anthrax outbreak.