High-performance deep learning pipeline predicts individuals in mixtures of DNA using sequencing data.

In this study, we proposed a deep learning (DL) model for classifying individuals from mixtures of DNA samples using 27 short tandem repeats and 94 single nucleotide polymorphisms obtained through massively parallel sequencing protocol. The model was trained/tested/validated with sequenced data from 6 individuals and then evaluated using mixtures from forensic DNA samples. The model successfully identified both the major and the minor contributors with 100% accuracy for 90 DNA mixtures, that were manually prepared by mixing sequence reads of 3 individuals at different ratios. Furthermore, the model identified 100% of the major contributors and 50-80% of the minor contributors in 20 two-sample external-mixed-samples at ratios of 1:39 and 1:9, respectively. To further demonstrate the versatility and applicability of the pipeline, we tested it on whole exome sequence data to classify subtypes of 20 breast cancer patients and achieved an area under curve of 0.85. Overall, we present, for the first time, a complete pipeline, including sequencing data processing steps and DL steps, that is applicable across different NGS platforms. We also introduced a sliding window approach, to overcome the sequence length variation problem of sequencing data, and demonstrate that it improves the model performance dramatically.

Massively parallel sequencing analysis of nondegraded and degraded DNA mixtures using the ForenSeq™ system in combination with EuroForMix software.

Massively parallel sequencing (MPS) technologies enable the simultaneous analysis of short tandem repeats (STRs) and single nucleotide polymorphisms (SNPs). MPS also enables the detection of alleles of the minor contributors in imbalanced DNA mixtures. In this study, 59 STRs (amelogenin, 27 autosomal STRs, 7 X-STRs, and 24 Y-STRs) and 94 identity-informative SNPs of 119 unrelated Taiwanese (50 men, 69 women) were sequenced using a commercial MPS kit. Forty-eight nondegraded and 44 highly degraded two-person artificial DNA mixtures with various minor to major ratios (1:9, 1:19, 1:29, 1:39, 1:79, and 1:99) were analyzed to examine the performance of this system for detecting the alleles of the minor contributors in DNA mixtures. Likelihood ratios based on continuous model were calculated using the EuroForMix for DNA mixture interpretation. The STR and SNP genotypes of these 119 Taiwanese were obtained. Several sequence variants of STRs were observed. Using EuroForMix software based on the sequence data of autosomal STRs and autosomal SNPs, 97.9% (47/48) and 97.7% (42/43) of minor donors were accurately inferred among the successfully analyzed nondegraded and degraded DNA mixtures, respectively. In conclusion, combined with EuroForMix software, this commercial kit is effective for assignment of the minor contributors in nondegraded and degraded DNA mixtures.

A single nucleotide polymorphism panel for individual identification and ancestry assignment in Caucasians and four East and Southeast Asian populations using a machine learning classifier.

Single nucleotide polymorphism (SNP) profiling is an effective means of individual identification and ancestry inferences in forensic genetics. This study established a SNP panel for the simultaneous individual identification and ancestry assignment of Caucasian and four East and Southeast Asian populations. We analyzed 220 SNPs (125 autosomal, 17 X-chromosomal, 30 Y-chromosomal, and 48 mitochondrial SNPs) of the DNA samples from 563 unrelated individuals of five populations (89 Caucasian, 234 Taiwanese Han, 90 Filipino, 79 Indonesian and 71 Vietnamese) and 18 degraded DNA samples. Informativeness for assignment (In) was used to select ancestry informative SNPs (AISNPs). A machine learning classifier, support vector machine (SVM), was used for ancestry assignment. Of the 220 SNPs, 62 were individual identification SNPs (IISNPs) (51 autosomal and 11 X-chromosomal SNPs) and 191 were AISNPs (100 autosomal, 13 X-chromosomal, 30 Y-chromosomal, and 48 mitochondrial SNPs). The 51 autosomal IISNPs offered cumulative random match probabilities (cRMPs) ranging from 1.56 × 10-21 to 3.16 × 10-22 among these five populations. Using AISNPs with the SVM, the overall accuracy rate of ancestry inference achieved in the testing dataset between Caucasian, Taiwanese Han, and Filipino populations was 88.9%, whereas it was 70.0% between Caucasians and each of the four East and Southeast Asian populations. For the 18 degraded DNA samples with incomplete profiling, the accuracy rate of ancestry assignment was 94.4%. We have developed a 220-SNP panel for simultaneous individual identification and ethnic origin differentiation between Caucasian and the four East and Southeast Asian populations. This SNP panel may assist with DNA analysis of forensic casework.

High efficiency sperm enrichment from forensic mock samples in bubble-based acoustic filtration devices for short tandem repeat (STR) analysis.

A bubble-based acoustofluidic filtration (BAF) microfluidic device, which employs cross-flow filtration (CFF) and acoustic streaming, separates cells with high efficiency for forensic analysis. Forensic samples are typically complex and contain a substantial number of squamous epithelial cells from the female vagina, which tend to have fouling problems during filtration due to their morphological and cell adhesion differences. To overcome this issue, the BAF device utilizes bubble oscillation by bulk acoustic wave (BAW) to generate acoustic streaming, which offers additional hydrodynamic forces for side flushing cleaning and achieves effective removal within a mere 0.5 seconds. Our device is tested with imbalanced cell mixtures of sperm and epithelial cells with large disparity ratios. By concurrently employing CFF and acoustic streaming, the samples with our sperm-enrichment can achieve 91.72-97.78% for the recovery rate and 74.58-89.26% for the purity in the sperm enrichment. They are further subjected to short tandem repeat (STR) profiling, enabling the identification of perpetrators. Notably, even samples with minimal sperm cells demonstrated a significant increase in the male donor DNA ratio, while the peak heights of female alleles became virtually undetectable. The exceptional cell separation capability demonstrated by our BAF device highlights its potential applications in forensic sciences and other areas of cell biology.

Adult femicide victims in forensic autopsy in Taiwan: A 10-year retrospective study.

BACKGROUND: Violence against women and adult femicides are critical medico-legal issues worldwide. Intimate partner violence is one of the leading contributory risk factors. This study aimed to describe the characteristics of femicides in Taiwan. METHODS: A retrospective analysis of forensic autopsy records of adult femicide victims in Taiwan during a 10-year period was carried out. The age, victim-offender relationship, injury patterns and causes of death were analyzed. RESULTS: Among the 220 adult femicide victims recruited, 114 were killed by intimate partners and 106 were killed by non-intimate partner offenders. The average age of victims killed by intimate partners (40.0 y/o) were younger than those killed by non-intimate partner perpetrators (48.6 y/o). The most common site of injuries in the intimate partner group and the non-intimate partner group was the neck and the upper limbs, respectively. The rates of bruise and intracranial injury of non-intimate partner group were significantly higher than that of the intimate partner group. The most common causes of death in both groups were strangulation and sharp force injury. The heart injury was significantly more frequent in victims offended by intimate partners than by other assailants. CONCLUSIONS: The characteristics of adult femicides, and the patterns of injury in victims killed by intimate partners and non-intimate partner offenders were different. This data is helpful for corpus inspection in forensic casework and for strategic planning of femicides prevention.

Child homicide victims in forensic autopsy in Taiwan: A 10-year retrospective study.

BACKGROUND: Child homicides are critical medico-legal issues worldwide. Data on the characteristics of these cases in Asia are limited. This study aimed to describe the characteristics of child homicides in Taiwan. METHODS: A retrospective analysis of forensic autopsy records of child homicide victims (aged 0-17 years) in Taiwan, during a 10-year period between 2001 and 2010, was carried out. The age, sex, relationship with the perpetrator(s), injury patterns of the victims, and causes of death were analyzed. RESULTS: In all, 193 child homicide autopsies were identified. There were 38 (19.7%), 82 (42.5%), 25 (13.0%), and 48 (24.9%) homicide victims aged under 1, 1-5, 6-12, and 13-17 years, respectively. One-hundred boys (mean age: 8.4±7.0) and 93 girls (mean age: 3.7±4.3) were included. A female predominance was noted among the victims aged 0-5. Blunt force (53.4%) was the most frequent method of injury, followed by suffocation/strangulation (20.2%) and sharp force (13.0%). Bruise (64.8%) and brain injury (45.1%) were the most common types of injuries. The cranium (62.2%) and face (60.6%) were the most frequently injured body regions. The distribution of fatal injuries varied among victims in different age groups. Neurogenic shock, asphyxia, and hemorrhagic shocks were most common in victims aged 0-5, 6-12, and 13-17, respectively. The most frequent causes of death included blunt force head injury (40.4%), suffocation/strangulation (20.2%), and sharp force lung trauma (7.3%). The type of offenders, injury methods, types of injuries, distribution of injuries, mechanism of death, and causes of death were significantly different among victims of different age groups. Eighteen (9.33%) victims displayed no external evidence of trauma. CONCLUSIONS: The patterns of injuries, mechanism of death, and causes of death were different among victims of different age groups. A female predominance was noted among the victims aged 0-5. Complete forensic autopsy is necessary to identify child homicide. This report will help forensic examiners and forensic pathologists recognize the signs of child homicide and serve as a working basis for these professionals.