Forensic science international. Genetics最新文献

With the advent of commercial DNA databases, investigative genetic genealogy (IGG) has emerged as a powerful forensic tool, rivalling the impact of STR analyses, introduced four decades ago. IGG has been frequently applied in the US and tested in other countries, but never in Norway. Here, we apply IGG to three cold criminal cases and successfully identify the donor of the DNA in two of these cases. Our findings suggest that when combined with phenotypic prediction and case information, IGG holds substantial potential for resolving both active and cold cases in Norway. This potential is amplified by the digitalization of archives and the transparent and structured nature of society in Norway. Additionally, the databases exhibit sufficient representation to yield matches with distant relatives. Moreover, this work has uncovered a series of lingering research questions spanning the entire workflow from DNA extraction to genealogy research. Finally, we highlight the possibility that more insights can be gleaned from genetic profiles, for instance using an accurate age prediction method. The results show that IGG can be successfully applied in Norway, having reached a level of maturity that enables identification of unknown individuals in cases where DNA is accessible.

Interpreting postmortem concentrations of 3,4-Methylenedioxymethamphetamine (MDMA) remains challenging due to the wide range of reported results and the potential idiosyncratic nature of MDMA toxicity. Consequently, forensic pathologists often rely on a body of evidence to establish conclusions regarding the cause and the manner of death in death involving MDMA. Given these issues, implementing pharmacogenetics' (PGx)' testing may be beneficial. Here, this report discusses an MDMA-related fatality and explores the benefits and limitations of implementing pharmacogenetics (PGx) analysis in such cases. A 34-year-old white European male was found dead at home, lying naked on his bed in a state of marked rigor mortis. MDMA and methylenedioxyamphetamine were quantified using liquid chromatography coupled to tandem mass spectrometry at respectively 3800 and 170 µg/L in femoral blood. PGx analysis was performed on a peripheral blood sample collected in EDTA tube. Deep analysis of cytochrome P450 (CYP) 2D6, 1A2, 2B6, 2C19, 3A4 and catechol-O-methyltransferase (COMT) genes (including copy number variations analysis) was performed by Next Generation Sequencing (NGS) on an Illumina MiSeq® sequencer using the Pharmacogenomics community panel (SOPHIA genetics® x RNPGx). The data obtained was analyzed using Sophia DDM® software. PGx analysis revealed three variants in CYP2C19 (rs75087398, rs12248560 and rs11188072) resulting in a CYP2C19 * 1/* 17 genotype, predictive of a rapid metabolism phenotype, implying greater MDMA elimination. Additionally, two variants were found in the COMT gene (rs4633TT, rs4680AA). In the literature, carriers of rs4680AA or rs4680GA genotypes exhibit lower enzyme activity compared to those homozygous for the G allele. Low COMT activity level has been associated with increased MDMA cardiovascular effects and biological changes, including an increased risk of hyponatremia which is particularly relevant here regarding the potential mechanism of death. Despite these findings, there are currently too few available studies to draw any definitive conclusions, indicating a need for further research in this area to fully understand all the implications. Moreover, focusing solely on metabolic enzymes may not fully explain all the variability in MDMA toxicity. A holistic genetic approach is necessary, incorporating both metabolic enzymes and pharmacological targets, including serotonin, dopamine, and norepinephrine transporters and receptors.

DNA methylation at age-related CpG (AR-CpG) sites holds significant promise for forensic age estimation. However, somatic models perform poorly in semen due to unique methylation dynamics during spermatogenesis, and current studies are constrained by the limited coverage of methylation microarrays. This study aimed to identify novel semen-specific AR-CpG sites using double-enzyme reduced representation bisulfite sequencing (dRRBS) and validate these markers, alongside previously reported sites and neighboring CpGs, using bisulfite amplicon sequencing (BSAS) to develop robust age estimation models. A methylome-wide association study was conducted on semen samples from 21 healthy Chinese men across three age groups, generating over 4 million CpG sites per sample at ≥ 5 × depth. Analysis of 721,840 shared CpG sites revealed that more than 95 % were not covered by conventional methylation microarrays. Differential methylation and correlation analyses identified 139 AR-CpG sites. A two-stage validation process using multiplex PCR-based BSAS was performed. In the first stage, 47 top dRRBS-identified AR-CpG sites, 26 literature-reported sites, and 242 neighboring CpGs were assessed in 129 semen samples (22-64 years), validating 31 dRRBS, 26 literature-reported, and 152 neighboring CpGs as age-related. The second stage examined 154 CpG sites in 247 samples (22-67 years), confirming 71 AR-CpG sites with |rho| > 0.50. Among these, chr2:129071885 (cg19998819) emerged as the strongest age-associated marker (rho = 0.81). Using the second BSAS dataset, age estimation models were developed with multiple linear regression and random forest (RF) algorithms within a repeated nested cross-validation (CV) framework (10-fold outer CV with 10-fold inner CV, repeated 10 times). The RF models demonstrated superior accuracy across feature subsets of 5-25 CpGs. The optimized 9-CpG RF model achieved an average root mean square error of 4.73 years (4.62-4.96, SD=0.10) and an average mean absolute error of 3.30 years (3.23-3.43, SD=0.06). This study demonstrates the utility of dRRBS for large-scale AR-CpG discovery and provides a robust age estimation model and a comprehensive reference database of semen-specific AR-CpG sites for forensic applications.

Genetic polymorphism can cause variation in tramadol (TR) pharmacokinetic characteristics and the expected clinical response. In forensic toxicology, the data about parent and metabolite concentrations (MRs; metabolic ratios) could facilitate to determine the cause of death and to assess time between drug intake and death. In this study, the aim was to investigate if UGT1A8, UGT2B7, ABCC2, and SLC22A1 genotyping can facilitate interpretation by investigating the frequency of UGT1A8, UGT2B7, ABCC2, and SLC22A1 genotypes in forensic autopsy cases positive for TR and to assess whether there is a correlation between these genetic variants and MRs. Cases positive for TR (n = 48) were genotyped by HaloPlex Target Enrichment system for UGT1A8, UGT2B7, ABCC2, and SLC22A1 sequencing, in order to identify single nucleotide polymorphisms (SNPs) and/or insertion deletion (INDELs). In addition to, the concentrations of TR and its metabolites (M1 & M2) were determined by LC-MS/MS. Cases were categorized by cause of death. The investigated SNPs/INDELs were not overrepresented in any group. We found significant correlations between several loci (12 out of 73) in UGT1A8, ABCC2, and SLC22A1 genes and MRs (M2/M1, TR/M2, and TR/M1) in post-mortem TR cases. These results indicate these polymorphisms in the 4 investigated genes might influence TR pharmacokinetics leading to an unsatisfactory therapeutic effect or increasing the risk of toxicity. However, these findings should be supported in future studies with larger groups of cases.

Latin American countries are distinguished by their highly admixed populations, characterized by a significant preservation of Native American matrilineal ancestry. This contrasts with the paternal lineages, which exhibit different patterns due to pronounced sex-biased mating practices during the colonial period. Uniparental genetic markers have been instrumental in population genetics, facilitating the reconstruction of human settlement histories and serving forensic identification purposes. The primary objective of this study was to investigate the diversity and structure of lineage markers in Argentina and compare them with other admixed populations in South America. For this study, we analyzed Y-STR and mtDNA haplotypes from 5202 unrelated individuals, providing a detailed description of the observed variability in both markers. Additionally, we conducted a genetic distance analysis, incorporating data from bibliographic sources across Argentina and South America. In pairwise comparisons among provinces, higher FST values were found in mtDNA haplotypes than in Y-STR haplotypes. This allows for more provinces to be grouped by similarity when using Y-STR data. These differences were also evident in the multidimensional scaling (MDS) analysis between South American countries. Y-STR haplotypes showed greater similarity to European haplotypes, whereas mtDNA haplotypes exhibited greater dispersion. Thus, the comprehensive compilation of haplotypes in this study, including those integrated from our research and those cited in existing literature, provides an in-depth understanding of the inherent genetic complexities within Argentina.

The utilisation of massively parallel sequencing (MPS) in forensic DNA analysis is on the rise, driven by the expansion of targeted MPS panels in the market and the introduction of forensic investigative genetic genealogy. The MPS library preparation process, integral to both whole-genome sequencing (WGS) and targeted MPS panel data generation, is largely based on converting double-stranded DNA (dsDNA) into sequencing libraries. In the current study, we examined the effect of seven routinely used forensic DNA extraction methods on the strandedness (single-stranded or double-stranded) and the fragment size of the DNA extracted from buccal swab, blood, bone and tooth samples. Our findings reveal a variation in the proportion of dsDNA and single-stranded DNA (ssDNA), with the phenol-chloroform and silica column-based extraction methods tested predominantly yielding dsDNA, while the tested Chelex and magnetic bead-based extraction methods predominantly yielded ssDNA. Additionally, fragment size analysis showed that high molecular weight dsDNA was recovered from buccal swab samples with all of the extraction methods except Chelex, which yielded relatively short dsDNA fragments. DNA extracted from tooth samples with tested magnetic bead-based extraction methods resulted in longer dsDNA fragments compared to the silica column-based extraction protocol.

DNA methylation patterns have emerged as reliable markers for age estimation, offering potential applications in forensic investigations, namely, in cases where there is no information about a possible suspect, in the identification of victims of mass disasters, or in immigration cases when assessing the age of individuals seeking asylum. This study aimed to evaluate the 11-CpG panel proposed by Aliferi et al. (2022) for age estimation. During the implementation phase, the ELOVL2 amplicon from the original work was replaced with a shorter fragment, and the two PCR multiplexes were optimized by changing the amplicons and primer conditions of each multiplex. The technical performance of the optimised assay was assessed using artificially methylated DNA standards. Robust quantification of the methylation levels at the 11 CpG sites was observed. Sensitivity tests demonstrated that DNA inputs down to 10 ng could produce reliable methylation quantification. Using the optimised panel, 148 Danish blood samples (18 - 68 years of age) were typed for their methylation status at the 11 CpG sites. Results showed that the DNA methylation at the 11 CpG loci was significantly correlated with age (0.68 ≤ r ≤ 0.88) in the Danish sample set, confirming the potential of the 11 CpGs in age prediction. A Danish age prediction model was constructed using 108 of the Danish blood samples and a support vector machine with polynomial function (SVMp). The performances of the new model and the original model based on UK individuals were compared using the remaining 40 Danish blood samples. Comparing the published model to the one developed in this study gave similar results with mean absolute errors (MAE) of 3.28 and 3.35, respectively. However, the original model showed a bias in the age predictions, underestimating the age by an average of 1.53 years in the Danish samples. This bias towards underestimation was not observed in the newly developed age prediction model based on Danish individuals. In summary, this assay provides a reasonably accurate age estimation of a single-source donor, if the sample material is blood and more than 10 ng of nuclear DNA can be extracted from the sample.

Human identification by forensic DNA profiling primarily relies on the analysis of short tandem repeat markers (STRs) and Amelogenin or other sex determining markers. The resultant DNA profiles can be compared directly between evidence and reference samples or indirectly (i.e., kinship) between human remains and family reference samples. Although Amelogenin serves as a phenotypic marker for biological sex, it is often considered innocuous, and the biological sex derived from this marker is routinely reported and/or uploaded to national DNA databases. However, biological sex does not necessarily align with gender identity, and chromosomal anomalies may affect the presentation of biological sex. Biological sex is genetically determined and assigned at birth based on anatomical features, whereas gender identity is an individual expression that may change over time and may not correspond with biological sex. This paper highlights how the differences between biological sex and gender identity can potentially impinge on individual privacy. Beyond gender differences, genetic anomalies related to the presentation of biological sex can occur, and the consequences of revealing such anomalies may have far-reaching implications for the individuals involved. Disclosing biological sex in a forensic DNA profile does not take into account the ramifications for persons or their families with genetic anomalies related to sex chromosomes (which may or may not be known to the individual), transgender transformations (which may or may not have been disclosed by the individual), or gender-identity expressions that do not correspond with biological sex. Through the presentation of case scenarios, while knowledge of biological sex may be important for operational forensic DNA laboratories and critical in certain cases, it is often not relevant to criminal investigations, courtroom deliberations, or public disclosure. It behoves all of us to understand that the dissemination of biological sex data in the public domain, especially in contexts where disclosure is unnecessary, may impact individual privacy. In light of the current understanding and growing impact of gender identity, it is recommended with all due speed that (1) 'biological sex' and 'gender' be recognised as separate concepts, not to be used synonymously nor interchangeably; (2) definitions of a DNA profile be reviewed and more clarity added; and (3) policies and protocols be developed to restrict such information from reports and court proceedings (i.e., public arenas), when not relevant, thereby reducing unwarranted intrusions into individual privacy and acknowledging the right to keep biological sex private and control how and when this personal information is shared.

While skeletal remains are known for their resilience and often serve as the final source of information for unidentified human remains (UHRs), the traditional downstream processing of these samples is challenging due to their low template nature, DNA degradation, and the presence of PCR inhibitors, typically resulting in limited probative information. To address this issue, advanced genotyping methods can be explored to retrieve additional genetic information from these challenging samples to maximize investigative leads. Therefore, this study investigated the effectiveness of three advanced genotyping methods and assessed their suitability with compromised skeletal samples: 1) targeted next generation sequencing (NGS) of both STRs and SNPs using the ForenSeq® DNA Signature Prep chemistry, 2) targeted NGS of SNPs using the ForenSeq® Kintelligence kit, and 3) SNP genotyping using a microarray via the Infinium Global Screening Array. The genotype recovery and added investigative leads were compared across all methods. All three approaches demonstrated success with the challenging skeletal samples used in this study. Specifically, the ForenSeq® DNA Signature Prep chemistry outperformed traditional STR typing by improving the recovery of CODIS core loci. Additionally, the ForenSeq® Kintelligence kit and Infinium Global Screening Array provided eligible results for forensic investigative genetic genealogy (FIGG) searching. Based on these successes, we have developed a proposed workflow for downstream processing of challenging skeletal samples. Following the guidelines of the US Department of Justice, the recovery of the CODIS core loci should be attempted through traditional CE-based methods or a NDIS-approved NGS chemistry, such as ForenSeq® DNA Signature Prep. Alternatively, a mitochondrial DNA profile may be uploaded to CODIS for comparisons in UHR cases. However, if no probative information is developed from the forensic profile uploaded to CODIS, then FIGG methods can be implemented using the Infinium Global Screening Array for high-quality skeletal samples (DNA concentrations ≥ 0.5 ng/µL) or the ForenSeq® Kintelligence chemistry for low-template skeletal remains (DNA concentration ≤ 0.5 ng/µL). These findings provide valuable insight into the suitability and efficacy of advanced genotyping methods, offering promising opportunities for enhancing the investigation of cases involving UHRs.