Weight of evidence of Y-STR matches computed with the discrete Laplace method: Impact of adding a suspect’s profile to a reference database

The discrete Laplace method is recommended by multiple parties (including the International Society for Forensic Genetics, ISFG) to estimate the weight of evidence in criminal cases when a suspect’s Y-STR profile matches the crime scene Y-STR profile. Unfortunately, modelling the distribution of Y-STR profiles in the population reference database is time-consuming and requires expert knowledge. When the suspect’s Y-STR profile is added to the database, as would be the protocol in many cases, the parameters of the discrete Laplace model must be re-estimated. We found that the likelihood ratios with and without adding the suspect’s Y-STR profile were almost identical with 1,000 or more Y-STR profiles in the database for Y-STR profiles with 8, 12, and 17 loci. Thus, likelihood ratio calculations can be performed in seconds if an established discrete Laplace model based on at least 1,000 Y-STR profiles is used. A match in a population reference database with 17 Y-STR loci from at least 1,000 male individuals results in a likelihood ratio above 10,000 in approximately 94% of the cases, and above 100,000 in approximately 82% of the cases. We offer free software accessible without restrictions to estimate a discrete Laplace model using a Y-STR reference database and subsequently to calculate likelihood ratios.     

Polymorphisms and microvariant sequences in the Japanese population for 25 Y-STR markers and their relationships to Y-chromosome haplogroups

Y-chromosomal short tandem repeat (Y-STR) markers have been used for forensic purposes such as kinship analysis of male-linage and detection of a male DNA component in a mixture of male and female DNA. Recently, rapidly mutating Y-STR (RM Y-STR) markers were reported that are expected to help distinguish close male relatives. This study provides data of Y-chromosomal haplotypes for 25 Y-STR markers, including six RM Y-STR markers (DYS576, DYS627, DYS518, DYS570, DYS449 and DYF387S1) typed with the Yfiler™ Plus kit in 1299 males of the Japanese population. Discrimination capacity increased from 87.2% for 16 Y-STR markers with the Yfiler™ kit to 99.6% for 25 Y-STR markers with the Yfiler™ Plus kit. We characterized sequences of observed microvariant alleles of eight Y-STR markers and a low-amplified allele of DYS390 by Sanger sequencing. DYF387S1, a multi-locus Y-STR marker that is located at two positions on the human Y-chromosome, was observed in tri-allelic patterns in 51 of 1299 samples (3.9%) and we found an extremely high frequency of the tri-allelic pattern of DYF387S1 in haplogroup C-M131. We also analyzed Y-STR gene diversity in each haplogroup and its relevance to mutation rates.     

Improving the regional Y-STR haplotype resolution utilizing haplogroup-determining Y-SNPs and the application of machine learning in Y-SNP haplogroup prediction in a forensic Y-STR database: A pilot study on male Chinese Yunnan Zhaoyang Han population

Improving the resolution of the current widely used Y-chromosomal short tandem repeat (Y-STR) dataset is of great importance for forensic investigators, and the current approach is limited, except for the addition of more Y-STR loci. In this research, a regional Y-DNA database was investigated to improve the Y-STR haplotype resolution utilizing a Y-SNP Pedigree Tagging System that includes 24 Y-chromosomal single nucleotide polymorphism (Y-SNP) loci. This pilot study was conducted in the Chinese Yunnan Zhaoyang Han population, and 3473 unrelated male individuals were enrolled. Based on data on the male haplogroups under different panels, the matched or near-matching (NM) Y-STR haplotype pairs from different haplogroups indicated the critical roles of haplogroups in improving the regional Y-STR haplotype resolution. A classic median-joining network analysis was performed using Y-STR or Y-STR/Y-SNP data to reconstruct population substructures, which revealed the ability of Y-SNPs to correct misclassifications from Y-STRs. Additionally, population substructures were reconstructed using multiple unsupervised or supervised dimensionality reduction methods, which indicated the potential of Y-STR haplotypes in predicting Y-SNP haplogroups. Haplogroup prediction models were built based on nine publicly accessible machine-learning (ML) approaches. The results showed that the best prediction accuracy score could reach 99.71% for major haplogroups and 98.54% for detailed haplogroups. Potential influences on prediction accuracy were assessed by adjusting the Y-STR locus numbers, selecting Y-STR loci with various mutabilities, and performing data processing. ML-based predictors generally presented a better prediction accuracy than two available predictors (Nevgen and EA-YPredictor). Three tree models were developed based on the Yfiler Plus panel with unprocessed input data, which showed their strong generalization ability in classifying various Chinese Han subgroups (validation dataset). In conclusion, this study revealed the significance and application prospects of Y-SNP haplogroups in improving regional Y-STR databases. Y-SNP haplogroups can be used to discriminate NM Y-STR haplotype pairs, and it is important for forensic Y-STR databases to develop haplogroup prediction tools to improve the accuracy of biogeographic ancestry inferences.     

Using probabilistic theory to develop interpretation guidelines for Y-STR profiles

Y-STR profiling makes up a small but important proportion of forensic DNA casework. Often Y-STR profiles are used when autosomal profiling has failed to yield an informative result. Consequently Y-STR profiles are often from the most challenging samples. In addition to these points, Y-STR loci are linked, meaning that evaluation of haplotype probabilities are either based on overly simplified counting methods or computationally costly genetic models, neither of which extend well to the evaluation of mixed Y-STR data. For all of these reasons Y-STR data analysis has not seen the same advances as autosomal STR data. We present here a probabilistic model for the interpretation of Y-STR data. Due to the fact that probabilistic systems for Y-STR data are still some way from reaching active casework, we also describe how data can be analysed in a continuous way to generate interpretational thresholds and guidelines.     

The impact of additional Y-STR loci on resolving common haplotypes and closely related individuals

Commercial Y-STR kits have permitted laboratories to go beyond the original nine minimal haplotype loci (MHL) and to discover the advantage of additional Y-STR loci in resolving common haplotypes. In an effort to examine the impact of Y-STR markers beyond the 17 loci now available in commercial kit form, new Y-STR loci are being investigated on a common set of samples representative of the major U.S. population groups. Additional Y-STRs can also increase the power of discrimination between closely related male individuals, which is important not only in forensics but also in the paternity and genetic genealogy communities.     

Y-chromosomal STRs haplotypes in the Taiwanese Paiwan population

The distribution of Y-chromosomal short tandem repeat (Y-STR) haplotypes was determined in a population of Taiwanese Paiwan aboriginals. Using 17 Y-STR markers, a total of 135 haplotypes were observed, 102 of which were unique. The overall haplotype diversity for the 17 Y-STR loci tested was 0.9922 and the discrimination capacity was 0.6490. In addition, three novel intermediate alleles at the DYS448 locus were also found.     

Improved specificity of Y-STR typing in DNA mixture samples

Y-STR loci are beneficial for the analysis of forensic samples especially in sexual assault cases or other situations where mixtures of male and female cells are present. However, the amplification of Y-chromosomal STRs is also known to result in the formation of artefactual amplification products, mainly due to insufficient PCR specificity. This is a major drawback of the method, as the sensitivity as well as the correct Y-STR interpretation are affected. In our study, the addition of a PCR enhancer to the reaction master-mix resulted in a significant increase of specificity of Y-STR typing. This was clearly demonstrated by a loss of artefactual signal with increasing enhancer concentration, while the peak heights of the Y-STR alleles were not significantly affected by the enhancer. Mixtures of up to 1:500 (200 pg male and 100 ng female DNA) gave correct Y-STR profiles when the PCR enhancer was added to the reaction, while artefactual amplification succeeded over Y-specific amplification when no PCR enhancer was present.     

Mutation analysis for 25 Y-STR markers in Japanese population

In this study, we analyzed DNA samples from 213 Japanese father son pairs with 25 Y-chromosome short tandem repeat (Y-STR) (DYS576, DYS389I, DYS635, DYS389II, DYS627, DYS460, DYS458, DYS19, YGATAH4, DYS448, DYS391, DYS456, DYS390, DYS438, DYS392, DYS518, DYS570, DYS437, DYS385, DYS449, DYS393, DYS439, DYS481, DYF387S1, and DYS533) markers using the Yfiler™ Plus PCR amplification kit. We calculated Y-STR mutation rates for each locus to evaluate the efficacy of the 25 Y-STR markers for paternity testing and forensic identification using samples from male relatives. Six rapidly mutating Y-STR markers (DYS576, DYS627, DYS518, DYS570, DYS449 and DYF387S1), previously reported to have high mutation rates (>1.0 × 10⁻²), are included in the 25 Y-STR markers, but our findings revealed that the mutation rates for all Y-STR markers except for DYS576 and DYS458 were lower than 1.0 × 10⁻². Therefore, the use of these 25 Y-STR markers may be useful for forensic identification in the Japanese population.     

The validation study of a novel assay with 30 slow and moderate mutation Y-STR markers for criminal investigation and database applications

International Journal of Legal Medicine - The Y chromosome short tandem repeat (Y-STR) haplotyping method has been widely used in forensic applications. However, the existing Y-STR panels are not...     

Developmental validation study of a 24-plex Y-STR direct amplification system for forensic application

International Journal of Legal Medicine - In the present study, validation data for 24 Y-STR loci from the Microreader™ 24Y Direct ID System was presented. Eight Y-STR loci have PCR product...     

Comprehensive annotated STR physical map of the human Y chromosome: Forensic implications

A plethora of Y-STR markers from diverse sources have been deposited in public databases and represent potential candidates for incorporation into the next generation of Y-STR multiplexes for forensic use. Here, based upon all of the Y-STR loci that have been deposited in the human genome database (>400), we have sequentially positioned each one along the Y chromosome using the most current human genome sequencing data (NCBI Build 35). The information derived from this work defines the number and relative position of all potentially forensically relevant Y-STR loci, their location within the physical linkage map of the Y chromosome and their relationship to structural genes. We conclude that there exists at present at least 417 separate Y-STR markers available for potential forensic use, although many of these will be found to be unsuitable for other reasons. However, from this data, we were able to identify 28 pairs of duplicated loci that were given separate DYS designations and four pairs of loci with overlapping flanking regions. Removing one locus from each set of duplicates reduced the number of potentially useful loci from 417 to 389. The derived information should be useful for workers who are designing novel Y-STR multiplexes to ensure the presence of non-synonymous loci and, if so desired, to avoid loci that lie within structural genes. It may also be useful for forensic casework practitioners (or molecular anthropologists) to aid in distinguishing between chromosomal rearrangements (such as duplications and deletions) and bona fide DNA admixtures or null alleles caused by primer binding site mutations. We illustrate the practical usefulness of the chromosomal positioning data in the design of eight multiplex systems using 94 Y-STR loci.     

Relating two deep-rooted pedigrees from Central Germany by high-resolution Y-STR haplotyping

Y-STR haplotyping is a powerful forensic and anthropological tool for identifying male lineages. We used high-resolution Y-STR haplotyping to evaluate the possibility of a blood relationship between two deep-rooted paternal genealogies with the same surname and originating from the same geographical region in Central Germany. One pedigree comprised 13 generations covering >450 years, the other comprised nine generations covering >300 years. Of the 68 loci tested, 64 (94%) consistently had the same allele in all males in the two pedigrees (except for some unambiguously sporadic mutations within pedigrees). Only four Y-STRs had a consistent allelic difference of exactly one repeat between the two pedigrees. These findings suggested that the two pedigrees were paternally related, and a conservative assessment taking average mutation rates and the available local haplotype frequencies for nine loci into account yielded a likelihood ratio of 8.2:1 in favour of this hypothesis. Our study thus highlights the power of Y-STR haplotyping to identify male lineages. It also shows that families can be linked to common ancestors on the basis of Y-STR data, even if these individuals lived several hundred years ago. However, the potential of Y-STR haplotyping could still not be fully exploited in our case due to a lack of appropriate population frequency data for all analysed Y-STR loci. This shortcoming makes a strong case for more comprehensive haplotype databases, including more samples and larger numbers of loci.     

Genetic analysis of 17 Y-chromosomal STR loci of Chinese Tujia ethnic group residing in Youyang Region of Southern China

Y-STR haplotype data were obtained in a population sample of 197 unrelated healthy male individuals of Chinese Tujia ethnic minority group residing in an autonomous county of Southern China using 17 Y-chromosome STR markers. A total of 197 haplotypes were identified in the set of Y-STR loci. The overall haplotype diversity for the Tujia population at 17 Y-STR loci was 1.0000 ± 0.0005. Genetic distance was estimated between this population and other 14 Chinese populations including Paiwan and Atayal populations of Taiwan, and Southern Han, Dong, Jing, Miao, Yao, Zhuang, Yi, Maonan, She, Hui, Sala, and Tibetan ethnic groups. The results demonstrated that the 17 Y-STR loci analyzed were highly polymorphic in Tujia ethnic group examined and hence useful for forensic cases, paternity testing, and population genetic studies.     

Y-STR DNA analysis of 154 female child sexual assault cases in the Philippines

The laboratory evaluated 154 sexual assault cases from four Child Protection Units in the Philippines involving female child victims aged from 2?years to 18?years old. All child victims sought medical attention within 72?h after sexual contact. In 130 cases, the child victim knew the alleged offender and identified them during the interview with the social worker. Penile ejaculation was reported by 68 child victims with varying reports of washing after contact. Overall, 84 child victims admitted having wiped their genitalia prior to the collection of biological samples for DNA testing. Laboratory personnel examined vaginal smears in only 109 cases using a light microscope and reported 23 samples to be positive for sperm cells. Using the PowerPlex? short tandem repeat of the Y chromosome (Y-STR) DNA multiplex system, male DNA was detected in vaginal swab samples from 63 child victims. In 39 cases, positive amplification at 11 Y-STR DNA markers consistent with a single male DNA profile was observed. Twenty-eight of these full single Y-STR DNA profiles were found to be unique when searched in worldwide Y-STR DNA population databases (~40,000 haplotypes), eight haplotypes matching Filipinos and/or Asian haplotypes and one Y-STR DNA profile only matching European, Caucasian, and Latin American haplotypes. Y-STR DNA profiles generated will be compared with reference DNA profiles of alleged offenders once reference samples are submitted to the laboratory.     

Genetic polymorphisms and haplotypic structure analysis of the Guizhou Gelao ethnic group based on 35 Y-STR loci

In this research, 35 Y-chromosomal short tandem repeat (Y-STR) loci were analyzed in 286 unrelated healthy Gelao male individuals from Guizhou Province, China. Allelic and haplotype frequencies, haplotype diversity (HD), haplotype match probability (HMP), and discrimination capacity (DC) values were computed. Pairwise Rst values were assessed by AMOVA analysis and visualized through multidimensional scaling and neighbor-joining tree construction. A total of 609 alleles were detected at the 35 Y-STR loci, and the allelic frequencies ranged from 0.0035 to 0.8322. The HD, HMP, and DC were 0.9999, 0.0036, and 0.9825, respectively. A total of 281 haplotypes were found at 35 loci in the 286 Gelao individuals, 98.25% of which were unique. Related forensic parameters revealed that this panel of 35 loci had a high level of genetic polymorphisms in the Gelao group. However, the use of multi-copy and rapidly mutating Y-STR loci could significantly increase the discrimination power of a common Y-STR panel. Analyses of population differentiation with the same set of common Y-STR loci demonstrated that the Gelao group has a closer genetic relationship with the Guizhou Han and Guizhou Miao groups than other groups.     

Autosomal and Y-STR analysis of degraded DNA from the 120-year-old skeletal remains of Ezekiel Harper

The 120-year-old skeletal remains of Confederate Civil War soldier Captain Ezekiel “Zeke” Harper were exhumed by court order in January 2011 for DNA analysis. The goal of the DNA testing was to support or refute whether Captain Harper had fathered a son (Earl J. Maxwell) with his Native American maid prior to his murder in 1892. Bones with adequate structural integrity (left tibia, right tibia, right femur, mandible, four teeth) were retrieved from the burial site and sent to the Institute of Applied Genetics in Fort Worth, Texas for analysis. Given the age and condition of the remains, three different extraction methods were used to maximize the probability of DNA recovery. The majority of the DNA isolates from over fifty separate bone sections yielded partial autosomal STR genotypes and partial Y-STR haplotypes. After comparing the partial results for concordance, consensus profiles were generated for comparison to reference samples from alleged family members. Considering the genetic recombination that occurs in autosomal DNA over the generations within a family, Y-STR analysis was determined to be the most appropriate and informative approach for determining potential kinship. Two of Earl J. Maxwell's grandsons submitted buccal samples for comparison. The Y-STR haplotypes obtained from both of these reference samples were identical to each other and to the alleles in Ezekiel Harper's consensus profile at all 17 loci examined. This Y-STR haplotype was not found in either of two major Y-STR population databases (U.S. Y-STR database and YHRD). The fact that the Y-STR haplotype obtained from Ezekiel's skeletal remains and Earl's grandsons is not found in either population database demonstrates its rarity and further supports a paternal lineage relationship among them. Results of the genetic analyses are consistent with the hypothesis that Earl J. Maxwell is the son of Ezekiel Harper.     

Extending STR markers in Y chromosome haplotypes

Two multiplex reactions were developed to amplify 16 Y-STRs (DYS19, DYS385, DYS389 I and II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, GATA A7.1, GATA A7.2, GATA A10, GATA C4, GATA H4). Here we extend previous population studies done in a sample from northern Portugal for the GATA A7.1, GATA A7.2, GATA C4 and GATA H4 loci. A total of 199 different haplotypes identified by the 16 Y-STR markers were observed in a sample of 208 male individuals, of which 190 were unique and 9 were found twice. The overall haplotype diversity was 0.9996. The haplotype diversity of the Y-STR set composed of the 8 new markers is higher than the Y-STR core set included in the Y-STR haplotype reference database. Sequence structure of new alleles for GATA C4 and GATA H4 is reported. The usefulness of the inclusion of this new set of Y-STRs in forensic casework was also assessed. The increase in haplotype diversity with the addition of any new Y-STR marker to the 8 Y-STR core set is dependent not only on the gene diversity (positively) but also (negatively) on the degree of gametic association between the markers and the haplotypes previously defined. For instance, in our sample the addition of the DYS437, DYS438 and GATA A7.2 to a 13-locus set increased haplotype diversity only by 0.0001.     

Y-STR analysis of degraded DNA using reduced-size amplicons

To increase the success rate of Y-STR genotyping for degraded DNA, we have developed two multiplex PCR sets for 21 Y-STR loci. Besides the 17 Y-STR loci of DYS19, DYS385, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, DYS448, DYS456, DYS458, DYS635, and GATA H4.1 contained in a commercial Y-STR kit, AmpFlSTR Yfiler, the other four loci of DYS388, DYS446, DYS447, and DYS449 were also included in the multiplexes to increase the discrimination capacity. Among a total of 21 Y-STR loci, the primers for eight loci (DYS385, DYS390, DYS438, DYS446, DYS448, DYS449, and DYS635) were newly designed in the present study and nine loci (DYS385, DYS390, DYS391, DYS392, DYS438, DYS439, DYS448, and DYS635) have PCR amplicons smaller than those of the AmpFlSTR Yfiler kit. A sensitivity test using serially diluted standard 9948 male DNA showed that all the values of Y-STR loci in the Y-miniplexes are reliable at template concentrations as low as 30 pg. We compared the effectiveness of the two multiplexes with the AmpFlSTR Yfiler kit by using both enzymatically degraded DNA and 30 samples of 50-year-old skeletal remains. This comparison demonstrated that the new Y-miniplex sets can produce a better signal from degraded DNA than the AmpFlSTR Yfiler kit.     

Y-STR profiling in two Afghanistan populations

Afghanistan's unique geostrategic position in Eurasia has historically attracted commerce, conflict and conquest to the region. It was also an important stop along the Silk Road, connecting the far eastern civilizations with the western world. Nevertheless, limited genetic studies have been performed in Afghan populations. In this study, 17 Y-chromosomal short tandem repeat (Y-STR) loci were typed to evaluate their forensic and population genetic applications in 189 unrelated Afghan males geographically partitioned along the Hindu Kush Mountain range into north (N=44) and south (N=145) populations. North Afghanistan (0.9734, 0.9905) exhibits higher haplotype diversity than south Afghanistan (0.9408, 0.9813) at both the minimal 9-loci and 17-loci Yfiler haplotypes, respectively. The overall haplotype diversity for both Afghan populations at 17 Y-STR loci is 0.9850 and the corresponding value for the minimal 9-loci haplotypes is 0.9487. A query using of the most frequent Afghan Yfiler haplotype (7.98%) against the worldwide Y-STR haplotype reference database (YHRD) returned no profile match, indicating a high power of discrimination with 17 Y-STR loci. A median-joining network based on 15 Y-STR loci displays limited haplotype sharing between the two Afghan populations, possibly due to the Hindu Kush Mountain range serving as a natural barrier to gene flow between the two regions.     

Y-STR loci diversity in native Alaskan populations

Y chromosome short tandem repeat (Y-STR) loci are important genetic markers for forensic biological evidence analyses. However, paternal inheritance, reduced effective population size, and lack of independence between loci can reduce Y-STR diversity and may yield greater population substructure effects on a locus-by-locus basis compared with the autosomal STR loci. Population studies are necessary to assess the genetic variation of forensically relevant markers so that proper inferences can be made about the rarity of DNA profiles. This study examined 16 Y-STRs in three sampled populations of Native Americans from Alaska: Inupiat, Yupik, and Athabaskan. Population genetic and statistical issues addressed were: (1) the degree of diversity at locus and haplotype levels, (2) determination of the loci that contribute more so to haplotype diversity, and (3) the effects of population substructure on forensic statistical calculations of the rarity of a Y-STR profile. All three population samples were highly polymorphic at the haplotype level for the 16 Y-STR markers; however, the Native Americans demonstrated reduced genetic diversity compared with major US populations. The degree of substructure indicated that the three populations were related and admixed in terms of paternal lineage. The examination of more polymorphic loci may be needed to increase the power of discrimination of Y-STR systems in these populations.