Anthrax molecular epidemiology and forensics: using the appropriate marker for different evolutionary scales

Area of Science:

• Genetics
• Microbiology
• Bioforensics

Background:

• Bacillus anthracis strain identification is critical for forensic and epidemiological analyses.
• Limited molecular variation among B. anthracis isolates necessitates the use of rare genetic variations for precise strain typing.
• Mutation is the primary driver of diversity in clonal pathogens like B. anthracis.

Purpose of the Study:

• To develop a precise and efficient method for subtyping Bacillus anthracis isolates.
• To leverage single nucleotide polymorphisms (SNPs) as stable phylogenetic markers for bacterial identification.
• To establish a hierarchical approach for discriminating closely related B. anthracis strains.

Main Methods:

• Utilized whole genome discovery to identify rare single nucleotide polymorphisms (SNPs).
• Selected diagnostic "Canonical SNPs" (canSNPs) at key phylogenetic positions.
• Developed a nested hierarchical strategy called Progressive Hierarchical Resolving Assays using Nucleic Acids (PHRANA) for tiered strain resolution.

Main Results:

• Identified rare SNPs as effective markers for B. anthracis subtyping.
• Demonstrated the utility of canSNPs for efficient and defining diagnostic assays.
• PHRANA successfully provided both deep phylogenetic insights and high-resolution discrimination among closely related isolates.

Conclusions:

• The PHRANA strategy offers a robust method for B. anthracis strain identification, applicable to various pathogens.
• This nested hierarchical approach mitigates data weighting issues and enhances phylogenetic accuracy.
• canSNPs and the PHRANA method significantly improve the precision of forensic and epidemiological analyses of anthrax cases.