Interpreting NUMTs in forensic genetics: Seeing the forest for the trees

Abstract

Nuclear mitochondrial DNA (mtDNA) segments (NUMTs) were discovered shortly after sequencing the first human mitochondrial genome. They have earlier been considered to represent archaic elements of ancient insertion events, but modern sequencing technologies and growing databases of mtDNA and NUMT sequences confirm that they are abundant and some of them phylogenetically young. Here, we build upon mtDNA/NUMT review articles published in the mid 2010 s and focus on the distinction of NUMTs and other artefacts that can be observed in aligned sequence reads, such as mixtures (contamination), point heteroplasmy, sequencing error and cytosine deamination. We show practical examples of the effect of the mtDNA enrichment method on the representation of NUMTs in the mapped sequence data and discuss methods to bioinformatically filter NUMTs from mtDNA reads.

From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel

“…On the other hand, mtDNA differences between the same sample are not unheard of, as DNA polymerases, amplification protocols, sequencing runs, and variant callers are frequent sources of disparity in genomics [ 35 , 44 ]. Discerning sequencing errors/false positives from true heteroplasmic variants is a challenging task, usually achieved through post-sequencing curation, looking for signs of poor amplification, strand bias, mutations in LCR, and the presence of NUMTs [ 41 , 45 , 46 , 47 , 48 , 49 ].…”

The time is now for ubiquitous forensicmtMPSanalysis

“…Artifacts in mtMPS data from background noise, damage, and NUMTs will occur and can negatively impact interpretation, including the interpretation of heteroplasmy. ISFG and recent published studies have provided recommendations and guidelines for navigating the interpretation and reporting process of mtMPS data (Crysup et al, 2021; Kim et al, 2018; Marshall & Parson, 2021; Parson et al, 2014; Sukser et al, 2021; SWGDAM Guidelines, 2019).…”

“…Smart et al (2019) formalized a phylogenetic‐based approach to identify potential NUMTs and provide more objective criteria for determining thresholds. These and other strategies to discern NUMTs from artifacts, mixtures, heteroplasmy and damage patterns have been summarized (Marshall & Parson, 2021).…”

The Value of Whole-Genome Sequencing for Mitochondrial DNA Population Studies: Strategies and Criteria for Extracting High-Quality Mitogenome Haplotypes

“…NUMTs are portions of mtDNA that have been inserted into the nuclear genome, most of them within the last 100,000 years [2]. There are more than 1090 documented NUMTs to date, ranging in size from 39 to 18,649 base pairs (bp), including three that are larger than the entire mitogenome (16,569 bp) [3,4]. In addition, the existence of entire copies of the mitogenome that were tandemly inserted as many as 50 times into a single location in the nuclear genome (referred to as a "mega-NUMT" by [5]) has been recently reported [6].…”

“…NUMT variants are often difficult to distinguish from authentic heteroplasmy and other sources of mixed data such as contamination from exogenous mtDNA, PCR or sequencing errors, and DNA damage (e.g., cytosine deamination) [4]. This is especially true when applying low variant detection thresholds.…”