The usefulness of measuring n-butyric acid concentration as a new indicator of blood decomposition in forensic autopsy

In forensic medicine, although various alcohols have been reported as indicators of decomposition in collected blood, no studies have examined short-chain fatty acids as indicators. In this study, the blood n-butyric acid concentration was quantified, and the association between n-butyric acid and decomposition was investigated to determine whether the detection of n-butyric acid could be a new indicator of decomposition. Among the forensic autopsies performed from 2016 to 2018 in our laboratory, the cases were divided into decomposed (n = 20) and non-decomposed (n = 20) groups based on macroscopic findings. Blood samples collected at the time of autopsy were derivatized with 3-nitrophenylhydrazine hydrochloride after solid-phase extraction. The n-butyric acid concentration was measured using liquid chromatography–tandem mass spectrometry. In addition, ethanol and n-propanol were measured using a gas chromatography-flame ionization detector. There was a significant difference (p < 0.01) in the concentrations of n-butyric acid between the decomposed and non-decomposed groups (0.343 ± 0.259 [0.030–0.973] and 0.003 ± 0.002 [0.001–0.007] mg/mL, respectively). In the decomposed group, n-butyric acid was detected at high concentrations, even in cases where n-propanol was low. These results suggest that n-butyric acid is more likely to be an indicator of blood decomposition than n-propanol.     

Development and validation of the EUROFORGEN NAME (North African and Middle Eastern) ancestry panel

Inference of biogeographic origin is an important factor in clinical, population and forensic genetics. The information provided by AIMs (Ancestry Informative Markers) can allow the differentiation of major continental population groups, and several AIM panels have been developed for this purpose. However, from these major population groups, Eurasia covers a wide area between two continents that is difficult to differentiate genetically. These populations display a gradual genetic cline from West Europe to South Asia in terms of allele frequency distribution. Although differences have been reported between Europe and South Asia, Middle East populations continue to be a target of further investigations due to the lack of genetic variability, therefore hampering their genetic differentiation from neighboring populations. In the present study, a custom-built ancestry panel was developed to analyze North African and Middle Eastern populations, designated the ‘NAME’ panel. The NAME panel contains 111 SNPs that have patterns of allele frequency differentiation that can distinguish individuals originating in North Africa and the Middle East when combined with a previous set of 126 Global AIM-SNPs.     

Genetic variation of 17 autosomal STR loci in the Lahu population from Yunnan and phylogenetic structure exploration among 28 populations

This study evaluated the genetic variation of 17 autosomal short tandem repeat (STR) loci included in the PowerPlex® 18D Kit. Samples of 562 unrelated healthy Lahu individuals living in Yunnan Province in southwestern China were investigated. The data were analyzed to provide information on allele frequencies and other statistical parameters relevant to the forensic population. Of the 17 loci, 16 reached the Hardy–Weinberg equilibrium after Bonferroni correction. A total of 176 alleles were identified in 17 STR loci, and allele frequencies ranged from 0.000 890 to 0.578 292. The combined discrimination power (CPD) and probability of excluding paternity (CPE) of the 17 STR loci were 0.999 999 999 999 999 999 489 and 0.999 998 301 753 122. The genetic relationships among 28 populations were also estimated.     

Development of the decision tree model for distinguishing individuals of Chinese four surnames from Zhanjiang Han population based on Y-STR haplotypes

Co-separation studies between surnames and Y chromosome genetic markers are beneficial to revealing population migrations, surname origins, population formation histories and forensic familial searching. Genetic distributions of 27 Y-STRs in Chinese four surnames (Li, Lin, Chen and Huang) from Zhanjiang Han population were investigated. Meanwhile, we tried to develop a decision tree model for surname predictions based on Y-STR haplotypes. Allelic frequencies of 27 Y-STRs showed that unique alleles were only observed in a certain surname; besides, some alleles displayed higher frequencies in a certain surname than those in other surnames, implying these alleles might be employed as the useful indicators for surname predictions. Haplotype match probability values of 27 Y-STRs in these surnames revealed that the system could be used as a valuable tool for forensic male identification. The developed decision tree model performed well for the training set with the accuracy of 0.9860 and obtained the relatively high accuracy (>0.70) for surname predictions of the testing set. To sum up, we explored the power of the machine learning to the surname predictions based on obtained Y-STR haplotypes, which showed promising application values in forensic familial searching.     

Eurasiaplex: A forensic SNP assay for differentiating European and South Asian ancestries

We have selected a set of single nucleotide polymorphisms (SNPs) with the specific aim of differentiating European and South Asian ancestries. The SNPs were combined into a 23-plex SNaPshot primer extension assay: Eurasiaplex, designed to complement an existing 34-plex forensic ancestry test with both marker sets occupying well-spaced genomic positions, enabling their combination as single profile submissions to the Bayesian Snipper forensic ancestry inference system. We analyzed the ability of Eurasiaplex plus 34plex SNPs to assign ancestry to a total 1648 profiles from 16 European, 7 Middle East, 13 Central-South Asian and 21 East Asian populations. Ancestry assignment likelihoods were estimated from Snipper using training sets of five-group data (three Eurasian groups, East Asian and African genotypes) and four-group data (Middle East genotypes removed). Five-group differentiations gave assignment success of 91% for NW European populations, 72% for Middle East populations and 39% for Central-South Asian populations, indicating Middle East individuals are not reliably differentiated from either Europeans or Central-South Asians. Four-group differentiations provided markedly improved assignment success rates of 97% for most continental Europeans tested (excluding Turkish and Adygei at the far eastern edge of Europe) and 95% for Central-South Asians, despite applying a probability threshold for the highest likelihood ratio above ‘100 times more likely’. As part of the assessment of the sensitivity of Eurasiaplex to analyze challenging forensic material we detail Eurasiaplex and 34-plex SNP typing to infer ancestry of a cranium recovered from the sea, achieving 82% SNP genotype completeness. Therefore, Eurasiaplex provides an informative and forensically robust approach to the differentiation of European and South Asian ancestries amongst Eurasian populations.     

Ion Torrent ™ Genexus ™ Integrated Sequencer and ForeNGS Analysis Software—An automatic NGS-STR workflow from DNA to profile for forensic science

The Ion Torrent ™ Genexus ™ Sequencer (Genexus) is a highly integrated instrument that can automate library construction, templating, and sequencing in a single-instrument run. By programing the ForeNGS Analysis Software (FNAS), we bridged the gap between sequencing and genotyping without manual intervention. FNAS can automatically transfer sequencing output files from Genexus, analyze the repeat and flanking regions aligned to the GRCh38 assembly, name the alleles according to the ISFG guidelines, and generate user-friendly interactive profiles. Genexus and FNAS can accomplish the fully automatic DNA-to-Profile workflow in forensics. Based on our experiences, the optimal assay parameters on Genexus were validated as follows: 24 cycles of target amplification for library construction; 40 μL of library and 400 bp of template size for templating; 852 flows of dNTPs by order of Ion samba HID2 for sequencing; and 750,000 reads per sample at minimum for 16 samples multiplexed on a lane. By developmental validations of the Precision ID Globalfiler ™ NGS STR Panel v2, Genexus presented competitive performance at the optimal assay parameters qualified to detect commonly used forensic STR markers. It could produce repeatable and reproducible results, and human profiles could be easily separated from nonhuman profiles. Additionally, Genexus was sensitive enough to detect samples with 100 pg of input DNA, and it was suitable for various types of case samples, especially for low copy number samples and degraded samples. Moreover, minor contributors could be detected between the 4:1 and 1:4 mixtures with an analysis threshold of 50 × . The Genexus workflow is a robust and labor-effective solution enabling forensic scientists to obtain NGS-STR profiles within a single day and with only the need to prepare DNA extracts, then set up Genexus, and finally interpret profiles on FNAS.     

Optimized variant calling for estimating kinship

One of the fundamental goals of forensic genetics is sample attribution, i.e., whether an item of evidence can be associated with some person or persons. The most common scenario involves a direct comparison, e.g., between DNA profiles from an evidentiary item and a sample collected from a person of interest. Less common is an indirect comparison in which kinship is used to potentially identify the source of the evidence. Because of the sheer amount of information lost in the hereditary process for comparison purposes, sampling a limited set of loci may not provide enough resolution to accurately resolve a relationship. Instead, whole genome techniques can sample the entirety of the genome or a sufficiently large portion of the genome and as such they may effect better relationship determinations. While relatively common in other areas of study, whole genome techniques have only begun to be explored in the forensic sciences. As such, bioinformatic pipelines are introduced for estimating kinship by massively parallel sequencing of whole genomes using approaches adapted from the medical and population genomic literature. The pipelines are designed to characterize a person’s entire genome, not just some set of targeted markers. Two different variant callers are considered, contrasting a classical variant calling algorithm (BCFtools) to a more modern deep convolution neural network (DeepVariant). Two different bioinformatic pipelines specific to each variant caller are introduced and evaluated in a titration series. Filters and thresholds are then optimized specifically for the purposes of estimating kinship as determined by the KING-robust algorithm. With the appropriate filtering and thresholds in place both tools perform similarly, with DeepVariant tending to produce more accurate genotypes, though the resultant types of inaccuracies tended to produce slightly less accurate overall estimates of relatedness     

Sequence polymorphisms of forensic Y-STRs revealed by a 68-plex in-house massively parallel sequencing panel

Sequence polymorphisms of Y chromosome short tandem repeat (Y-STR) markers can be unveiled using next generation sequencing (NGS). Compared to capillary electrophoresis, NGS has the advantage of distinguishing between some alleles of the same length. Here, a 68-plex in-house panel covering 67 Y-STR loci and the sex determinant Amelogenin locus, was developed. The accuracy of this panel was 100% concordant with three standard reference samples. The sensitive was as low as 250 pg. A total of 466 length-based alleles, 806 sequence-based alleles, and 149 haplotypes were observed across 149 Chinese Han individuals. The total haplotype diversity and discrimination capacity was 1.0000 in detected samples. The DYS710 locus possessed the highest diversity by sequence among these Y-STRs, with 109 sequence-based alleles observed. Micro-variant alleles with the same length were observed in 39 Y-STR loci, with their sequence variations mainly attributable to repeat pattern variations. While the number of sequence-based alleles identified for DYS447, DYS449, DYS710, DYS720 and DYF387S1a/b was approximately three times that of their length-based alleles, flanking sequence variations were observed in 18 alleles. In addition, 201 sequence-based alleles in 42 loci were newly discovered. This significantly expanded the knowledge of human Y-STR sequence polymorphisms. Collectively, the 68-plex panel provided reliable Y-STR results as well as higher resolution for paternal lineage analysis.     

A fatal case of accidental oral formaldehyde poisoning and its pathomorphological characteristics

International Journal of Legal Medicine - Formaldehyde is a colourless irritating gas at room temperature, which, therefore, is usually stored in liquid form. This compound is often used as an...