Assessing zygomatic shape and size for estimating sex and ancestry in a South African sample

Unidentified, decomposed and skeletonised human remains are frequently found in South Africa, therefore, standardised, reliable and relevant sex and ancestry estimation methods are required for forensic identification. This study assessed sex and ancestral variation in zygomatic size and shape in a South African population using geometric morphometric analyses. The zygoma of 158 South African individuals were sampled. Eight zygomatic landmarks were captured in 3-dimensions using a Microscribe G2 digitiser and assessed using procrustean geometric morphometrics. Shape and size differences were analysed using multivariate linear regression, discriminant function and canonical variate analyses. Males had significantly larger zygomas than females. Significant shape variation was found between ancestral groups. Bantu-speaking and Mixed ancestry individuals had narrower, shorter and more anteriorly projecting orbital margins, whilst Europeans had vertically elongated and receded orbital margins. European ancestral groups were most discernible from Bantu-speakers and Mixed ancestral groups. Ancestry estimation accuracies improved when ancestry was aggregated with sex. Pairwise ancestry-linked comparisons in females were as follows; Bantu-speakers (76%) from Europeans (72%), Bantu-speakers (71%) from Mixed ancestry (59%) and European (72%) from Mixed ancestry (63%). Similarly, ancestry-linked comparisons in males were as follows; Bantu-speakers (77%) from Europeans (81%), Bantu-speakers (53%) from Mixed ancestry (59%) and European (72%) from Mixed ancestry (82%). Size differences are putatively linked to variations in hormone-regulated growth and muscular robusticity between males and females. Shape variations between ancestral groups are likely attributable to the heterogenous genetic and ancestral origins of the South African population. It is challenging to distinguish between South Africa Bantu speakers and Mixed ancestry people due to Mixed ancestry individuals having variable genetic contributions from Khoesan, Bantu-speakers, Europeans and Asians. Bantu-speaking and Mixed ancestry people had zygomatic morphologies consistent with historical thermoregulatory adaptations to sub-Saharan climates, reported in African-descendants. Zygomatic morphology in European descendants suggests ancestral origins from colder climatic regions. This study demonstrated the utility of the zygoma in distinguishing between ancestral groups in South Africa, but further research is required to develop population-specific standards to distinguish between South African populations with shared African ancestry. The zygoma shows a promising ability to estimate sex and ancestry in South Africans, suggesting population specific standards for this bone may be of forensic interest.     

Y-chromosomal STR haplotypes in Pakistani populations

16 Y-specific STR loci have been analysed in 711 males from 12 populations in Pakistan. Individual loci showed between 4 and 10 alleles, and diversities ranged from 0.07 to 0.77. A total of 527 different haplotypes were found and the haplotype diversity ranged from 0.92 to 0.99 for the different populations. 446 haplotypes occurred in single individuals, and only 19 haplotypes were present in more than three males, but two striking examples of haplotype sharing were found, one involving 13 individuals, and the other 17. The 13 individuals were all Parsis, and 16 of the 17 were Brahuis, providing evidence for population substructuring.     

Racial variation in the proximal and distal femur: heritability and forensic utility.

The femur has been studied successfully by physical anthropologists for many years. Such traits as femoral head diameter and bicondylar width have been examined extensively and are of great value to forensic anthropologists and other skeletal biologists in sex identification. A number of studies over the past decade by the author and his former students have shown marked racial differences in the shape of the proximal femur and in at least one trait of the distal femur--intercondylar notch height. Anterior-posterior (AP) diameter of the proximal femur is much greater among Whites and Blacks than among East Asians and American Indians. Blacks have slightly greater intercondylar notch height than Whites. Other features, such as torsion, also differ between the major geographic racial populations. Current analysis suggests that the East Polynesians fall close to the American Indians and East Asians in the degree of flatness of the proximal femur. One study has tracked the degree of change in flatness during individual development, and finds little change within major populations from the youngest to the oldest individuals. Temporal changes within populations are likewise minimal. Two studies have examined sex differences within populations, which are also found to be very slight. Racial differences, on the other hand, are quite significant, and individuals of admixed ancestry fall intermediate between the two parental populations. Such suggestions of high heritability in the shape of the proximal and distal femur make these traits very useful in assessing ancestry in forensic contexts.     

Palate Shape and Depth: A Shape‐Matching and Machine Learning Method for Estimating Ancestry from Human Skeletal Remains

In the past, assessing ancestry relied on the naked eye and observer experience; however, replicability has become an important aspect of such analysis through the application of metric techniques. This study examines palate shape and assesses ancestry quantitatively using a 3D digitizer and shape‐matching and machine learning methods. Palate curves and depths were recorded, processed, and tested for 376 individuals. Palate shape was an accurate indicator of ancestry in 58% of cases. Cluster analysis revealed that the parabolic, hyperbolic, and elliptical shapes are discrete from one another. Preliminary results indicate that palate depth in Hispanic individuals is greatest. Palate shape appears to be a useful indicator of ancestry, particularly when assessed by a computer. However, these data suggest that palate shape is not useful for assessing ancestry in Hispanic individuals. Although ancestry may be determined from palate shape, the use of multiple features is recommended and more reliable.     

Development of a deoxyribonucleic acid (DNA) restriction fragment length polymorphism (RFLP) database for Punjabis in East Punjab,India

In response to continuing interest in obtaining reference deoxyribonucleic acid (DNA) analysis data for previously unstudied population groups, blood samples were collected from Punjabi individuals living in East Punjab, India. This first segment of our research is focused on restriction fragment length polymorphism (RFLP) analysis, with future segments anticipated for various polymerase chain reaction (PCR) based techniques. In this study, the samples were subjected to RFLP analysis using HaeIII, followed by hybridization with variable number tandem repeat (VNTR) probes for loci D2S44, D1S7, D10S28, D4S139, D17S79 and D5S110. The band sizes of the resulting patterns were estimated using an FBI imaging system. The resulting data were subjected to statistical analysis for conformity with Hardy-Weinberg expectations, first for the total population of Punjabis, and additionally for the subgroups of Sikhs and Hindus. The loci are highly polymorphic in all sample populations studied. Except for D5S110, there is no evidence for departure from Hardy-Weinberg equilibrium (HWE) for the VNTR loci in the population groups. In addition, there is little evidence of correlation between the alleles at any of the pairs of loci and no evidence of association across the six loci. Finally, the data suggest that a multiple locus VNTR profile would be rare in the Punjabi or either of its subgroups.     

A multivariate statistical approach to for the evaluation of the biogeographical ancestry information from traditional STRs

The capability to achieve biogeographic ancestry (BGA) information from DNA profiles have been largely explored in forensic genetics because of its potential usefulness in providing investigative clues. For law enforcement and security purposes, when genetic data have been obtained from unknown evidence, but no reference samples are available and no hints come out from DNA databases, it would be extremely useful at least to infer the ethno-geographic origin of the stain donor by just examining traditional STRs DNA profiles.Current protocols for ethnic origin estimation using STRs profiles are usually based on Principal Component Analysis approaches and Bayesian methods. The present study provides an alternative approach that involves the use of target multivariate data analysis strategies for estimation of the BGA information from unknown biological traces. A powerful multivariate technique such as Partial Least Squares-Discriminant Analysis (PLS-DA) has been applied on NIST U.S. population datasets containing, for instance, the allele frequencies of African-American, Asian, Caucasian and Hispanic individuals. PLS-DA approach provided robust classifications, yielding high sensitivity and specificity models capable of discriminating the populations on ethnic basis. Finally, a real casework has been examined by extending the developed model to smaller and more geographically-restricted populations involving, for instance, Albanian, Italian and Montenegrian individuals.     

基于38-plex InDels的青海地区三个族群遗传推断

目的 基于38-plex InDels族群推断体系研究青海地区汉族、回族及撒拉族的族群成分与遗传结构.方法 使用38-plex InDels复合扩增体系检测3个族群的220份样本并获取InDels位点分型,利用主成分分析、STRUCTURE聚类分析及系统发育树综合分析族群之间的遗传关系,使用族群推断软件DAA v1.0...     

Forensic Applicability of Femur Subtrochanteric Shape to Ancestry Assessment in Thai and White American Males

Ancestry assessment from the postcranial skeleton presents a significant challenge to forensic anthropologists. However, metric dimensions of the femur subtrochanteric region are believed to distinguish between individuals of Asian and non‐Asian descent. This study tests the discriminatory power of subtrochanteric shape using modern samples of 128 Thai and 77 White American males. Results indicate that the samples' platymeric index distributions are significantly different (p ≤ 0.001), with the Thai platymeric index range generally lower and the White American range generally higher. While the application of ancestry assessment methods developed from Native American subtrochanteric data results in low correct classification rates for the Thai sample (50.8–57.8%), adapting these methods to the current samples leads to better classification. The Thai data may be more useful in forensic analysis than previously published subtrochanteric data derived from Native American samples. Adapting methods to include appropriate geographic and contemporaneous populations increases the accuracy of femur subtrochanteric ancestry methods.     

Microscopical discrimination of human head hairs sharing a mitochondrial haplogroup

In forensic analyses, determining the level of consensus among examiners for hair comparison conclusions and ancestry identifications is important for assessing the scientific validity of microscopical hair examinations. Here, we present data from an interlaboratory study on the accuracy of microscopical hair comparisons among a subset of experienced hair examiners currently analyzing hair in forensic laboratories across the United States. We examined how well microscopical analysis of hair can reliably be used to differentiate hair samples, many of which were macroscopically similar. Using cut hair samples, many sharing similar macroscopic and microscopic features, collected from individuals who share the same mitochondrial haplogroup as an indication of genetic relatedness, we tested multiple aspects that could impact hair comparisons. This research tested the extent to which morphological features related to ancestry and hair length influence conclusions. Microscopical hair examinations yielded accurate assessments of inclusion/exclusion relative to the reference samples among 85% of the pairwise comparisons. We found shorter hairs had reduced levels of accuracy and hairs from populations examiners were not familiar with may have impacted their ability to resolve features. The reliability of ancestry determinations is not yet clear, but we found indications that the existing categories are only somewhat related to current ethnic and genetic variation. Our results provide support for the continued utility of microscopical comparison of hairs within forensic laboratories and to advocate for a combined analytical approach using both microscopical analysis and mtDNA data on all forensic analyses of hair.     

Evaluation of 15 biparental STR loci in human identification and genetic study of the Kannada-speaking groups of India

Thirteen tetranucleotide and 2 pentanucleotide repeat units were analyzed in 167 unrelated Kannada-speaking individuals belonging to 3 important communities, namely, Kuruva, Bhovi, and Christians, residing in different districts of Karnataka, India. Allele frequency data obtained from the analysis of 15 short tandem report (STR) markers of the subpopulation groups included in the study were observed to be similar, indicating a common ancestry or gene flow among these communities. Departures from Hardy-Weinberg equilibrium were observed in Kuruva population at locus D5S818 and D18S51 and at locus CSF1PO in the Christian community. The data of these communities were analyzed with allele frequency data of 4 other populations from Karnataka, India-Iyengar Brahmin, Gowda, Lingayat, and Muslim-to compute the combined power of discrimination, ranged from 0.962 to 0.974, with negligible difference between populations. The combined power of exclusion, however, remained constant at 0.999 for all populations evaluated in the study. Thus, the 15 markers selected for this study were found to be highly suitable in human identification and for providing information on genetic polymorphism.     

Investigating the resolution of ancestry testing in geographic regions characterized by high population admixture

Ancestry testing can provide valuable information in forensic applications, but its efficiency has not yet been properly evaluated when used in highly admixed populations. In this work we tested a commercial ancestry panel containing 165 autosomal SNPs in 30 Dubai residents. Most of the tested individuals displayed, as expected, an admixed profile, but in general the main inferred ancestry was of Southwest Asian extraction, testifying that these tests can provide useful complementary forensic information.     

Informativeness of the CODIS STR loci for admixture analysis

Population admixture (or ancestry) is used as an approach to gene discovery in complex diseases, particularly when the disease prevalence varies widely across geographic populations. Admixture analysis could be useful for forensics because an indication of a perpetrator's ancestry would narrow the pool of suspects for a particular crime. The purpose of this study was to use Fisher's information to identify informative sets of markers for admixture analysis. Using published founding population allele frequencies we test three marker sets for efficacy for estimating admixture: the FBI CODIS Core STR loci, the HGDP-CEPH Human Genome Diversity Cell Line Panel and the set of 39 ancestry informative SNPS from the Shriver lab at Pennsylvania State University. We conclude that the FBI CODIS Core STR set is valid for admixture analysis, but not the most precise. We recommend using a combination of the most informative markers from the HGDP-CEPH and Shriver loci sets.     

Investigation of 74 microhaplotypes for kinship testing in US populations

Microhaplotypes are markers that consist of haplotype blocks of SNPs, which can be analyzed by massively parallel sequencing technologies. These allow determining the haplotype phase at every locus by clonal sequencing each DNA strand. MHs are polymorphic loci with same size alleles, no stutter, and lower mutation rate than STRs. They can provide the same power of discrimination of STR-kits, thus useful for mixture deconvolution, but more accurate ancestry prediction than STRs. In this study we investigated the potential of a recently developed 74plex-MH panel for kinship testing using the Familias software.Samples from families of four major US population groups were collected and genotyped using the 74plex-MH panel. MH allele frequency data from 347 individuals were imported into Familias software along with STR allele frequency data of 29 loci (NIST dataset) from 1036 individuals. Different family scenarios were tested and these included unrelated vs parent-child, unrelated vs full siblings, unrelated vs half siblings, unrelated vs cousin pairs. The prediction of the kinship relation for the four populations of interest was reported as Log10 of the likelihood ratio (LR).Overall, the panel of 74MHs and 29STRs showed similar performance in predicting the correct kinship scenarios tested. Correct prediction was reported for parent-child, full siblings, and half sibling scenarios, but not for the cousin pairs scenario. The panel of 74 MHs showed larger Log10LR values than the 29 STR-assay, thus demonstrating the effectiveness of this biomarker as a tool for kinship testing in addition to mixture deconvolution and ancestry prediction.     

Inference of human geographic origins using Alu insertion polymorphisms

The inference of an individual's geographic ancestry or origin can be critical in narrowing the field of potential suspects in a criminal investigation. Most current technologies rely on single nucleotide polymorphism (SNP) genotypes to accomplish this task. However, SNPs can introduce homoplasy into an analysis since they can be identical-by-state. We introduce the use of insertion polymorphisms based on short interspersed elements (SINEs) as a potential alternative to SNPs. SINE polymorphisms are identical-by-descent, essentially homoplasy-free, and inexpensive to genotype using a variety of approaches. Herein, we present results of a blind study using 100 Alu insertion polymorphisms to infer the geographic ancestry of 18 unknown individuals from a variety of geographic locations. Using a Structure analysis of the Alu insertion polymorphism-based genotypes, we were able to correctly infer the geographic affiliation of all 18 unknown human individuals with high levels of confidence. This technique to infer the geographic affiliation of unknown human DNA samples will be a useful tool in forensic genomics.     

The SNPforID 34-plex—Its ability to infer level of admixture in individuals

Forensic scientists use genetic individualization markers to include or exclude persons of interest in investigations. However, when there are no suspects due to absence of database matches or eye-witness information, prediction of biogeographical ancestry can be a valuable investigative tool. The SNPforID 34-plex uses 34 autosomal markers to predict ancestry from three geographic regions, Africa, Europe and East Asia. However, its ability to identify levels of admixture within individuals is unclear. We tested the 34-plex assay in 56 individuals from 15 families with varying levels of self-declared Asian/European admixed ancestry. STRUCTURE 2.3.4 was used for population structure analysis and cluster information provided inferences on levels of admixture. Chi-square tests were performed to evaluate the ability of the SNPforID 34-plex to predict level of admixture. The average/SD Asian and European contribution for individuals self-declared as first generation since admixture was 0.46/0.13 and 0.54/0.13, respectively. As expected, the average European contribution increased for individuals of 1/4, 1/8 and 1/16 Asian/European ancestries – 0.78/0.13, 0.89/0.05 and 0.91/0.03, respectively. There were no significant differences between observed and expected average contribution from each ancestry. However, individual outliers were observed, which could have been misclassified if analyzed separately. These results suggest the 34-plex can be a reliable tool to predict levels of admixture; however caution is required when an individual sample is investigated. A larger number of markers, combined with increased sample sizes comprising varying levels of admixture of different biogeographical ancestries, are required to enhance the ability to predict an individual's level of biogeographical ancestry.     

Quantification of Maxillary Dental Arcade Curvature and the Estimation of Biological Ancestry in Forensic Anthropology

Previous studies suggest that palate shape is a useful indicator of biological ancestry in human remains. This study evaluates interobserver error in ancestry estimation using palate shape and explores palate shape variation in Gullah (descendants of West Africans) and Seminole (Indigenous American) population samples using geometric morphometric analysis. Ten participants were asked to ascribe biological ancestry and shape to 28 dental casts based on a classification scheme employed in previous studies. The mean correct classification was 42.0%, indicating that the likelihood of assigning the correct ancestry is very poor and not significantly different from random assignment (p = 0.12). The accuracy analysis based on categorical classification of the casts was complemented by geometric morphometric analysis of nine 3D landmarks reflecting palate shape of 158 casts. Principal component analysis results show no difference between populations regarding palate shape, and cross‐validated discriminant function analysis correctly classified only 62.0% of the specimens. Combined, these results show that previous methods to estimate ancestry are inaccurate and that this inaccuracy is probably due to a lack of palate shape differences between groups, rather than limitation of the analytical method per se. Therefore, we recommend caution should be used when choosing to apply the analysis of palate shape in forensically relevant contexts.     

"Biological identikit": Development of a SNPs-panel for the analysis of forensic DNA phenotyping and ancestry

Personal identification in mass disasters and in crimes is essential for humanitarian, ethical and legal reasons. In these contexts, when individuals cannot be identified by standard forensic DNA analysis, the Forensic DNA Phenotyping and the analysis of the biogeographical ancestry could help. The aim of this study was to evaluate the potential of a new panel of 891 SNPs in predicting phenotypic traits and biogeographical origin to create a “biological identikit”. In addition to fresh biological material, old evidence found at the crime scene or extracted and long-term stored DNA were tested with 41 SNPs for phenotyping and 850 SNPs for ancestry. All the SNPs were successfully incorporated into a single two-step multiplex PCR reaction using the IonAmpliSeq ™ Library Plus and applied for massive parallel sequencing with the Ion S5 platform using up to 0.05 ng/µL of DNA. The analysis of the results was carried out with an in-house predictive algorithm and consulting 20 population databases. By comparing the results obtained with identikit or video-photographic surveys, it was possible to predict phenotype and ancestry with an accuracy greater than 90%. While these new markers cannot identify a specific individual, they can be a valuable investigative tool.     

Morphometric Analysis of the Neurocranium in an Adult South African Cadaveric Sample,

Craniometric studies of South Africans yield high accuracies of sex and ancestry classification, but most assess only inter‐group variation of Black and White individuals, excluding the highly heterogeneous Colored group, which constitute a significant proportion of the population. This study applied a geometric morphometric approach to the neurocrania of 774 Black, Colored, and White individuals to assess sex and ancestry estimation accuracy based on the detected morphological variation. Accuracies of 70% and 83% were achieved for sex and ancestry, respectively, with ancestry‐related variation contributing the largest proportion of overall observed variation. Even when comparing the closely related Black and Colored groups, relatively high accuracies were obtained. It is thus recommended that a similar approach be used to develop a contemporary three‐dimensional database, which can be used to objectively, reliably, and accurately classify unknown remains in the South African forensic context.     

Cranial Nonmetric Variation and Estimating Ancestry*

Abstract: Historically, when predicting the ancestry of human skeletal remains, forensic anthropologists have not fully considered the variation within human populations, but instead have relied on a typological, experience‐based approach. Unfortunately, reliance on observer experience has produced a method that is as much an art as it is a science. This research focuses on the frequency distribution and inter‐trait correlations of 11 common morphoscopic traits to demonstrate that the experience‐based approach to ancestry prediction is indeed an art that is unscientific, because it is unreplicable, unreliable, and invalid. Ten of 11 traits examined had frequency distributions with significant differences (p < 0.001) between groups, but the range in variation of these traits far exceeds previous assumptions. Such within group variation clearly demonstrates that extreme trait expressions are not reliable for estimating ancestry through visual observation alone, but instead that these traits should be analyzed within a statistical framework.     

Discrimination of half-siblings when maternal genotypes are known

Given the DNA profiles of two individuals and one parent (say the mother) of each, we present likelihood ratios (LRs) comparing the hypothesis that they have the same father with the hypothesis of unrelated fathers. If the individuals have the same mother, the problem is to distinguish full- from half-siblings, otherwise we are comparing a half-sibling relationship with unrelated. We simulate STR profiles at up to 60 loci, based on allele proportions observed at 15 loci in three populations, and use them to approximate misclassification rates both for binary classification (e.g. "half-sib" versus "unrelated"), and when a third "cannot say" category is included. We find that reliable inferences in the absence of the mothers' profiles require many more STR loci than the 10-25 loci that are currently routinely available. However, profiling the two mothers conveys more discriminatory power than profiling the same number of additional loci in the individuals themselves. Our likelihood ratio formulas include a theta (or Fst) adjustment to allow for the individuals concerned to have recent shared ancestry (coancestry), relative to the population from which the allele frequency database is drawn. We illustrate that using an appropriate value of theta can reduce the average misclassification rate.