Search Results (9)

Background/Objectives: Understanding and predicting human pigmentation traits is crucial for individual identification. Genome-wide association studies have revealed numerous pigmentation-associated SNPs, indicating genetic overlap among pigmentation traits and offering the potential to develop predictive models without the need for analyzing large numbers of SNPs. Methods: In this study, we assessed the performance of the HIrisPlex-S system, which predicts eye, hair, and skin color, on 412 individuals from the Spanish population. Model performance was calculated using metrics including accuracy, area under the curve, sensitivity, specificity, and positive and negative predictive value. Results: Our results showed high prediction accuracies (70% to 97%) for blue and brown eyes, brown hair, and intermediate skin. However, challenges arose with the remaining categories. The model had difficulty distinguishing between intermediate eye colors and similar shades of hair and exhibited a significant percentage of individuals with incorrectly predicted dark and pale skin, emphasizing the importance of careful interpretation of final predictions. Future studies considering quantitative pigmentation may achieve more accurate predictions by not relying on categories. Furthermore, our findings suggested that not all previously established SNPs showed a significant association with pigmentation in our population. For instance, the number of markers used for eye color prediction could be reduced to four while still maintaining reasonable predictive accuracy within our population. Conclusions: Overall, our results suggest that it may be possible to reduce the number of SNPs used in some cases without compromising accuracy. However, further validation in larger and more diverse populations is essential to draw firm conclusions and make broader generalizations. Full article

It is very important to generate phenotypic results that are reliable when processing chronological old skeletal remains for cases involving the identification of missing persons. To improve the success of pigmentation prediction in Second World War victims, three bones from each of the eight skeletons analyzed were included in the study, which makes it possible to generate a consensus profile. The PowerQuant System was used for quantification, the ESI 17 Fast System was used for STR typing, and a customized version of the HIrisPlex panel was used for PCR-MPS. The HID Ion Chef Instrument was used for library preparation and templating. Sequencing was performed with the Ion GeneStudio S5 System. Identical full profiles and identical hair and eye color predictions were achieved from three bones analyzed per skeleton. Blue eye color was predicted in five skeletons and brown in three skeletons. Blond hair color was predicted in one skeleton, blond to dark blond in three skeletons, brown to dark brown in two skeletons, and dark brown to black in two skeletons. The reproducibility and reliability of the results proved the multisample analysis method to be beneficial for phenotyping chronological old skeletons because differences in DNA yields in different bone types provide a greater possibility of obtaining a better-quality consensus profile. Full article

Forensic genomic systems allow simultaneously analyzing identity informative (iiSNPs), ancestry informative (aiSNPs), and phenotype informative (piSNPs) genetic markers. Among these kits, the ForenSeq DNA Signature prep (Verogen) analyzes identity STRs and SNPs as well as 24 piSNPs from the HIrisPlex system to predict the hair and eye color. We report herein these 24 piSNPs in 88 samples from Monterrey City (Northeast, Mexico) based on the ForenSeq DNA Signature prep. Phenotypes were predicted by genotype results with both Universal Analysis Software (UAS) and the web tool of the Erasmus Medical Center (EMC). We observed predominantly brown eyes (96.5%) and black hair (75%) phenotypes, whereas blue eyes, and blond and red hair were not observed. Both UAS and EMC showed high performance in eye color prediction (p ≥ 96.6%), but a lower accuracy was observed for hair color prediction. Overall, UAS hair color predictions showed better performance and robustness than those obtained with the EMC web tool (when hair shade is excluded). Although we employed a threshold (p > 70%), we suggest using the EMC enhanced approach to avoid the exclusion of a high number of samples. Finally, although our results are helpful to employ these genomic tools to predict eye color, caution is suggested for hair color prediction in Latin American (admixed) populations such as those studied herein, principally when no black color is predicted. Full article

In the last few years, predicting externally visible characteristics (EVCs) by adopting informative DNA molecular markers has become a method in forensic genetics that has increased its value, giving rise to an interesting field called “Forensic DNA Phenotyping” (FDP). The most meaningful forensic applications of EVCs prediction are those in which, having only a DNA sample isolated from highly decomposed remains, it is essential to reconstruct the physical appearance of a person. Through this approach, we set out to evaluate 20 skeletal remains of Italian provenance in order to associate them with as many cases of missing persons as possible. To achieve the intended goal, in this work we applied the HIrisPlex-S multiplex system through the conventional short tandem repeats (STR) method to confirm the expected identity of subjects by evaluating phenotypic features. To investigate the reliability and accuracy of the DNA-based EVCs prediction, pictures of the cases were compared as they were available to researchers. Results showed an overall prediction accuracy greater than 90% for all three phenotypic features—iris, hair, and skin colour—at a probability threshold of 0.7. The experimental analysis showed inconclusive results in only two cases; this is probably due to the characteristics of subjects who had an intermediate eye and hair colour, for which the DNA-based system needs to improve the prediction accuracy. Full article

Forensic DNA Phenotyping (FDP) can reveal the appearance of an unknown individual by predicting the ancestry, phenotype (i.e., hair, eye, skin color), and age from DNA obtained at the crime scene. The HIrisPlex system has been developed to simultaneously predict eye and hair color. However, the prediction accuracy of the system needs to be assessed for the tested population before implementing FDP in casework. In this study, we evaluated the performance of the HIrisPlex system on 149 individuals from the Turkish population. We applied the single-based extension (SNaPshot chemistry) method and used the HIrisPlex online tool to test the prediction of the eye and hair colors. The accuracy of the HIrisPlex system was assessed through the calculation of the area under the receiver characteristic operating curves (AUC), sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). The results showed that the proposed method successfully predicted the eye and hair color, especially for blue (100%) and brown (95.60%) eye and black (95.23) and brown (98.94) hair colors. As observed in previous studies, the system failed to predict intermediate eye color, representing 25% in our cohort. The majority of incorrect predictions were observed for blond hair color (40.7%). Previous HIrisPlex studies have also noted difficulties with these phenotypes. Our study shows that the HIrisPlex system can be applied to forensic casework in Turkey with careful interpretation of the data, particularly intermediate eye color and blond hair color. Full article

Forensic DNA phenotyping (FDP) provides the ability to predict the human external traits from unknown sample donors, directly from minute amounts of DNA found at the crime scene. We developed a MPS multiplex assay, with the aim of genotyping all 41 DNA markers included in the HIrisPlex-S system for simultaneous prediction of eye, hair and skin colours. Forensic samples such as blood, skeletal remains, touch DNA, saliva swab, artificially degraded samples together with individuals with known phenotypes and a set of 2800 M control DNA were sequenced on the Ion Torrent platform in order to evaluate the concordance testing results and the forensic suitability of the 41-plex MPS assay. The panel was evaluated by testing a different number of PCR cycles and the volume of reagents for library preparation. The study demonstrated that full and reliable profiles were obtained with 0.1–5 ng, even with high degraded DNA. The increment of the number of PCR cycles results in an improvement of correctly genotyping and phenotyping for samples with low amounts of degraded DNA but higher frequencies of artefacts were found. The high DNA degradation level did not influence the correct genotyping and phenotyping and the critical parameter affecting the result is the quantity of input DNA. Eye and hair colour was predicted in 92.60% of individuals and skin colour in 85.15% of individuals. The results suggest that this MPS assay is robust, highly sensitive and useful for human pigmentation prediction in the forensic genetic field. Full article

Single-cell sequencing is a fast developing and very promising field; however, it is not commonly used in forensics. The main motivation behind introducing this technology into forensics is to improve mixture deconvolution, especially when a trace consists of the same cell type. Successful studies demonstrate the ability to analyze a mixture by separating single cells and obtaining CE-based STR profiles. This indicates a potential use of the method in other forensic investigations, like forensic DNA phenotyping, in which using mixed traces is not fully recommended. For this study, we collected single-source autopsy blood from which the white cells were first stained and later separated with the DEPArray™ N×T System. Groups of 20, 10, and 5 cells, as well as 20 single cells, were collected and submitted for DNA extraction. Libraries were prepared using the Ion AmpliSeq™ PhenoTrivium Panel, which includes both phenotype (HIrisPlex-S: eye, hair, and skin color) and ancestry-associated SNP-markers. Prior to sequencing, half of the single-cell-based libraries were additionally amplified and purified in order to improve the library concentrations. Ancestry and phenotype analysis resulted in nearly full consensus profiles resulting in correct predictions not only for the cells groups but also for the ten re-amplified single-cell libraries. Our results suggest that sequencing of single cells can be a promising tool used to deconvolute mixed traces submitted for forensic DNA phenotyping. Full article

A custom plate of OpenArray™ technology was evaluated to test 60 single-nucleotide polymorphisms (SNPs) validated for the prediction of eye color, hair color, and skin pigmentation, and for personal identification. The SNPs were selected from already validated subsets (Hirisplex-s, Precision ID Identity SNP Panel, and ForenSeq DNA Signature Prep Kit). The concordance rate and call rate for every SNP were calculated by analyzing 314 sequenced DNA samples. The sensitivity of the assay was assessed by preparing a dilution series of 10.0, 5.0, 1.0, and 0.5 ng. The OpenArray™ platform obtained an average call rate of 96.9% and a concordance rate near 99.8%. Sensitivity testing performed on serial dilutions demonstrated that a sample with 0.5 ng of total input DNA can be correctly typed. The profiles of the 19 SNPs selected for human identification reached a random match probability (RMP) of, on average, 10⁻⁸. An analysis of 21 examples of biological evidence from 8 individuals, that generated single short tandem repeat profiles during the routine workflow, demonstrated the applicability of this technology in real cases. Seventeen samples were correctly typed, revealing a call rate higher than 90%. Accordingly, the phenotype prediction revealed the same accuracy described in the corresponding validation data. Despite the reduced discrimination power of this system compared to STR based kits, the OpenArray™ System can be used to exclude suspects and prioritize samples for downstream analyses, providing well-established information about the prediction of eye color, hair color, and skin pigmentation. More studies will be needed for further validation of this technology and to consider the opportunity to implement this custom array with more SNPs to obtain a lower RMP and to include markers for studies of ancestry and lineage. Full article

The study of DNA to predict externally visible characteristics (EVCs) and the biogeographical ancestry (BGA) from unknown samples is gaining relevance in forensic genetics. Technical developments in Massively Parallel Sequencing (MPS) enable the simultaneous analysis of hundreds of DNA markers, which improves successful Forensic DNA Phenotyping (FDP). The EU-funded VISAGE (VISible Attributes through GEnomics) Consortium has developed various targeted MPS-based lab tools to apply FDP in routine forensic analyses. Here, we present an evaluation of the VISAGE Basic tool for appearance and ancestry prediction based on PowerSeq chemistry (Promega) on a MiSeq FGx System (Illumina). The panel consists of 153 single nucleotide polymorphisms (SNPs) that provide information about EVCs (41 SNPs for eye, hair and skin color from HIrisPlex-S) and continental BGA (115 SNPs; three overlap with the EVCs SNP set). The assay was evaluated for sensitivity, repeatability and genotyping concordance, as well as its performance with casework-type samples. This targeted MPS assay provided complete genotypes at all 153 SNPs down to 125 pg of input DNA and 99.67% correct genotypes at 50 pg. It was robust in terms of repeatability and concordance and provided useful results with casework-type samples. The results suggest that this MPS assay is a useful tool for basic appearance and ancestry prediction in forensic genetics for users interested in applying PowerSeq chemistry and MiSeq for this purpose. Full article