Select methods for microbial forensic nucleic acid analysis of trace and uncultivable specimens

Genetic characterization of microorganisms is facilitated by their inherent amplification process. Microbial growth in vitro is an enrichment process that can be exploited to obtain sufficient, if not substantial, amounts of nucleic acids for molecular characterization. However, the vast majority of microorganisms cannot be cultured in the laboratory, and, even of those forensically relevant pathogens than can be cultured, there will be microbial forensic investigations where residual nucleic acids may be of low quantity and/or highly damaged. Genetic analysis of uncultivable microorganisms and/or highly compromised DNA samples is one of the challenges facing analysts. However, advances in molecular biology have provided tools for the characterization of a variety of molecular targets. A number of high throughput methods have been developed that employ next-generation sequencing or microarrays to genetically characterize biological evidence. All of these methods require some form of target enrichment to reliably identify the pathogen and to attempt some degree of attribution. In this chapter, capture and whole genome amplification strategies are described to provide insight into how poor quality and limited quantity microbial nucleic acid samples may be analyzed. With such strategies, the analytical repertoire of the microbial forensic scientist is augmented allowing for more successful investigative leads to support attribution.