NIJ 2014-DN-BX-K022 and 2016-DN-BX-0171

Forensic laboratories have not taken advantage of intrinsic levels of mitochondrial (mt) DNA heteroplasmy that exists in humans.  A next generation DNA sequencing (NGS) approach using massively parallel sequencing (MPS) methods will allow the community to achieve this goal.  Prior to implementation, validation studies must be conducted, and a better understanding of the rates of heteroplasmy will be required in order to effectively report casework findings.  The rate of heteroplasmy in the control region (CR) of the mtgenome has been measured in a European population (NIJ 2014 DN-BX-K022), and in individuals of African, East Asian and Latino decent (2016 DN-BX-0171).

We have measured the rate of heteroplasmy in the control region (CR) of the mtgenome using an MPS approach on the MiSeq from Illumina.  Rates were assessed on an individual and per nucleotide basis for ~1301 individuals of European, African, East Asian, and Latino ancestry, across different age groups and for each gender.  To address a previous limitation, we worked with a local company (SoftGenetics, Inc., State College, PA) to develop a software tool for researchers and practitioners to better align MPS data associated with mtDNA sequence, and to provide reporting practices consistent with forensic interests (Holland et al., 2017).  Given our understanding of the diverse nature of mtDNA haplotypes across different population groups, and that mutations resulting in low-level heteroplasmy may be associated with local sequence and their effects on the replication process, the goals of our research study were to measure the rate of heteroplasmy and assess whether or not the sites of observed heteroplasmy were linked to the primary haplotype.  Findings are in the process of being prepared for publication and once published will be a helpful guide for the reporting of mtDNA heteroplasmy; including statistical analysis.  Below is a figure reflecting the posterior mean frequency of observed heteroplasmy depending on the haplogroup from which the primary haplotype was observed.  As expected, and using a conservative approach, as the number of individuals in the haplogroup increases, the mean frequency decreases.