Application of the Human Virome to Touched Objects and Hair Shafts

This proposal is designed to address the ongoing need to create forensically relevant linkages between persons, places, and objects by developing the heretofore untapped potential of the human viral microbiome (virome). The human virome is a source of rich genetic diversity that needs to be examined to determine if it is stable, transferable, and provides a sufficient power of discrimination to be used as an alternative to traditional human forensic deoxyribonucleic acid (DNA) tests when conditions are not suitable for them. The human bacterial microbiome is already being examined as an alternative method for human identification in forensically relevant cases. The human virome offers some advantages as the viral genomes are even smaller than those of bacteria, and thus are potentially more stable, have a variety of morphologies (double- and single-stranded) increasing the possible number of discriminating markers, and is present throughout the human body, including the skin and body fluids, making it transferrable. Also, the copy number of viral genomes, compared to the copy number of human genomes, in a given volume is substantially higher, increasing the likelihood of isolating a sufficient quantity for successful testing. There is currently a complete lack of empirical data on the human virome with regard to its suitability to forensic applications. The proposed work will address this gap in our knowledge by generating data in the previously mentioned areas of stability, transfer, and discrimination as applied to various types of evidentiary material. The project will recruit 20 adult human subjects, 10 female and 10 male, and collect hair virome samples and transfer skin virome samples from 5 different objects. These samples will be processed for viral DNA extraction and subsequent next generation sequencing (NGS) using a robust sequence preprocessing pipeline including determination of virome composition and abundances. Statistical evaluation and testing will be used to assess the stability, transfer, and discriminatory power of the virome adjusted for subject characteristics. The proposal hypothesizes that the genetic diversity contained in each human being’s particular virome and characterized in our previous work can be transferred to physical objects and detected when standard human DNA samples are not viable. The potential for virome profiles developed from cut and shed hairs will also be examined in detail. The work will be done with protocols, instrumentation, and analysis methods that are either already in forensic DNA laboratories or can be readily assimilated. We anticipate scholarly publications, presentations, and access to data processing and analysis pipelines. This project will further our understanding of the human viral microbiome, address the critical barrier of individual identification when conditions for human DNA testing are poor, and develop new technology for the use of the human virome as a tool for forensic applications. 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