Utilizing eDNA from Four Biological Taxa Associated with Geologic Evidence for Sample-to-Sample Comparisons and Study Site Separation

Geologic materials, such as soil and dust, are often submitted as trace evidence to crime laboratories, and can provide valuable information to link a suspect to a crime scene, identify the origin of evidence, and for intelligence gathering. Forensic geologists routinely examine and characterize inorganic components of geologic materials (e.g., mineral content, pH, color), but the organic components (e.g., pollen, plant/insect fragments) are rarely analyzed primarily due to a lack of available expertise. To harness the full evidentiary value, analyzing biological taxa associated with geologic materials could be used to supplement traditional inorganic examinations; associations between biological taxa and their environment are possible, and thus could provide valuable information for sample-to-sample comparisons and sample origin. Research to date has primarily focused on using bacteria to characterize soils; however, before the analysis of biological taxa can be implemented into casework, the following gaps need to be addressed: 1) utility of other biological taxa, not only bacteria, 2) applicability to diverse geologic evidence, and 3) impact of temporal and spatial variables. This proposed research project will explore whether the same species of four different biological taxa are recovered from soil and dust regardless of collection season, in order to determine which taxa(on) is most appropriate to separate geologic materials. To answer these research questions, mock geologic evidence (n, 6) will be collected in triplicate monthly for one year, from an urban and agricultural site in Raleigh, NC. DNA metabarcoding will be used to characterize bacteria, fungi, arthropods, and plants associated with geologic material isolated from mock evidence. With this method, short yet informative regions of the genome will be amplified in multiple species simultaneously, with amplicons subsequently sequenced on the Illumina (R)MiniSeq. Sequence data will be processed through the DADA2 pipeline, and only high-quality unique sequences will be retained for searching against taxa-specific databases for identification. Taxonomic abundance charts will be created to visualize the difference between replicates, across mock evidence types, study sites, and collection times. Canonical analysis of principal coordinates will be used to assess which taxa(on) can be used for sample-to-sample comparisons and spatial separation. Bray-Curtis dissimilarity will be employed to measure the dissimilarity between all 432 samples. This study will provide the forensics community with an assessment of the utility of various biological taxa for characterizing mock geologic evidence. This is necessary before this approach can be considered for use in routine casework.
Note: This project contains a research and/or development component, as defined in applicable law, and complies with Part 200 Uniform Requirements - 2 CFR 200.210(a)(14). CA/NCF